Cell suspensions

Biol Reprod, 1983 Mar, 28(2), 350 - 62
Prostaglandin release by zygotes and endometria of pregnant rabbits; Harper MJ et al.; When 4-day rabbit zygotes were incubated for 1 h at 37 degrees C in vitro, very little prostaglandin (PG) was released into the medium, and the concentration of PGs in the zygotes after incubation was also low . The release of prostaglandin E (PGE) and prostaglandin F (PGF) into the medium, and their concentration in the zygotes after incubation, increased sharply on Days 6 and 7 of pregnancy, reaching, by Day 7, values close to 200 ng of each PG released in 1 h per mg of protein . By contrast, endometrial samples on Days 4 and 5 of pregnancy released more PGF and less PGE than the zygotes of the same ages on a per mg of protein basis, and on Days 6 and 7, less of both PGs . Furthermore, endometrial concentrations of PGs after incubation, except for PGF on Day 4, were always lower than values for zygotes . Endometrial concentrations of PGs on Day 6 were lower before than after incubation . Although there was a slight upward trend in PG release by endometrial samples with increasing length of pregnancy, the changes were minimal and, in the case of PGE, none of the mean values exceeded 1 ng per mg of protein . In 7-day blastocysts, high levels of both PGF and PGE were found in the blastocoelic fluid, and these did not change during the 1-h incubation . The release of PGF and PGE during in vitro incubation of ruptured and washed Day 6 blastocysts was stimulated by arachidonic acid, and that of PGF, but not PGE, inhibited by indomethacin . The release of PGE, but not of PGF, from Day 6 blastocysts was inhibited by low temperature, and the same conditions inhibited release of both PGF and PGE from endometrial cell suspensions . It seems that both blastocysts and endometria have capability to synthesize PGs, the blastocysts being particularly active in this regard on Days 6 and 7 of pregnancy . It is hypothesized that, in vivo, Day 6 and 7 blastocysts release large quantities of PGs which trigger some of the local endometrial changes associated with pregnancy.

Mutat Res, 1983 Mar, 108(1-3), 395 - 404
UV- and X-ray-sensitive double mutants of mouse L5178Y cells are synergistically more sensitive to 4-nitroquinoline-1-oxide than is either of the single mutants; Shiomi T et al.; The X-ray-sensitive mutant M10 and the UV-sensitive mutant Q31 of mouse lymphoma L5178Y cells are both sensitive to killing by 4-nitroquinoline-1-oxide (4NQO) . Since cell hybridization experiments showed that the 4NQO sensitivities in M10 and Q31 cells behaved as codominant traits (Shiomi et al., 1982c), it is not possible to determine by complementation test whether the M10 and the Q31 mutations responsible for 4NQO sensitivities are allelic . We have obviated this difficulty by selecting double mutants that are sensitive to both X-rays and UV . From X-ray-sensitive M10 cells, two UV-sensitive mutants (XU 1 and XU 2) were isolated by a cell-suspension spotting method . XU 1 and XU 2 were found to belong to the same complementation group as Q31 (group I) . Double mutants XU 1 and XU 2 were 30-37-fold more sensitive to 4NQO than parental L5178Y cells, whereas the single mutants M10 and Q31 were only 6-8-fold more sensitive to 4NQO than L5178Y cells in terms of D10 values (dose required to reduce survival to 10%) . These results show that the M10-Q31-double mutations enhance 4NQO sensitivity synergistically, indicating that the M10 and the Q31 mutations relevant to 4NQO sensitivities are non-allelic . The implications of this finding are discussed.

Int J Lepr Other Mycobact Dis, 1983 Mar, 51(1), 64 - 71
Respiratory activities of in vitro grown Mycobacterium lepraemurium; Ishaque M; Mycobacterium lepraemurium was cultivated on Ogawa egg-yolk medium and its respiratory activities using several substrates were investigated . Glycerol and succinate were oxidized at a slow rate by the cell-free extracts prepared from in vitro grown Hawaiian and Keishicho strains of M . lepraemurium . None of the other intermediates of the glycolysis cycle as well as of the tricarboxylic acid cycle was oxidized by the whole cell suspensions or cell-free extracts . Likewise, many sulfur compounds such as cystine, mercaptosuccinate, monothioglycerol, thioacetate, etc., were inactive . However, sulfhydryl compounds such as L-cysteine, D-cysteine, DL-cysteine, dithioerythritol, dithiothritol, and DL-penicillamine were actively oxidized . Yeast extract was also readily oxidized by cell suspensions of in vitro grown M . lepraemurium . Tween 80 was very poorly oxidized by whole cell suspensions but the cell-free preparations catalyzed an active oxidation of Tween 80 . While bovine serum albumin was oxidized at a slow rate by cell-free extracts, egg albumin was inactive . The thiol-binding agents, p-hydroxymercuribenzoate and N-ethylmaleimide were effective inhibitors of succinate and NADH oxidation, thus indicating the involvement of sulfhydryl compounds in the metabolism of M . lepraemurium.

Immunology, 1983 Mar, 48(3), 461 - 7
Large mononuclear Ia-positive veiled cells in Peyer's patches . II . Localization in rat Peyer's patches; Wilders MM et al.; Dendritic Ia-positive veiled cells were seen in frozen sections of rat Peyer's patches within the lymphoepithelium, in the reticular area just underneath and in the T-dependent interfollicular area . From immunohistochemical stainings with anti-IgA, IgE, IgG, IgM and monoclonal anti T lymphocyte sera it is concluded that these Ia-positive cells do not belong to lymphocyte series . The ultrastructure of these Ia-positive veiled cells resembled the morphology of the veiled cells found in Peyer's patch cell suspensions and in skin lymph . It is suggested that these cells form an antigen presenting cell system in mucosa-associated lymphoid tissue, similar to the antigen presenting cell system in the skin.

Chem Biol Interact, 1983 Mar, 43(3), 253 - 61
Biochemical evidence for chemical and/or topographic differences in the lipoperoxidative processes induced by CCl4 and iron; Poli G et al.; Isolated rat hepatocytes, treated with CCl4 or ADP-Fe3+ complex show an enhanced lipid peroxidation and a decreased glucose 6-phosphatase activity . Lipid peroxidation is much more stimulated by ADP-Fe3+ or Fe3+ than by CCl4, when the metal and the haloalkane are used at a similar concentration . Increasing rates of lipid peroxidation in the different experimental conditions do not correlate with the degree of glucose 6-phosphatase inactivation, which is produced by CCl4 and not by a similar amount of ferric iron . In the case of iron, its intracellular concentration must be higher to give the enzyme inactivation exerted by CCl4 . Higher intracellular levels of iron are reached when the metal is added to the cell suspension together with ADP . Under these conditions there is inactivation of glucose 6-phosphatase . Possible mechanisms accounting for a different enzyme sensitivity to iron and CCl4 are discussed.

Kidney Int, 1983 Mar, 23(3), 448 - 57
Characterization of inflammatory cells in autoimmune tubulointerstitial nephritis in rats; Mampaso FM et al.; Brown Norway rats immunized with bovine tubular basement membrane (TBM) antigens develop tubulointerstitial nephritis . The composition of the inflammatory cell infiltrate was characterized in kidney tissue sections and cell suspensions obtained from affected kidneys . Anti-TBM antibody deposition in the kidney began 8 days after immunization and was followed on days 8 to 10 by C3 deposits and infiltration of polymorphonuclear leukocytes (PMN) . After day 13, the infiltrate became almost exclusively mononuclear in character . On day 13, the inflammatory mononuclear cells recovered by Ficoll-Hypaque centrifugation contained 10% Ig+ cells (B cells), 60% W3/25+ cells (T helper cells), 9% OX8+ cells (T suppressor cells), 9% esterase+ cells (monocytes/macrophages), 4% renal cells, and 8% other unidentified cells . Monocytes/macrophages were prominent only at the latest stages of the disease . The ratio of W3/25+ to OX8+ cells was higher in the kidney than in the spleen or peripheral blood . The sequential accumulation of T cells and then monocytes/macrophages after an initial antibody, complement, and PMN lesion suggests a role for the T cells (selective prevalence of the helper T cell population over that of suppressor T cells) both as inflammatory cells and in progression and regulation of the subsequent stages of injury.

Immunology, 1983 Mar, 48(3), 439 - 52
Mast cell differentiation depends on T cells and granule synthesis on fibroblasts; Davidson S et al.; Mast cell differentiation was generated in the following three experimental situations: (i) infection of mice with Schistosoma Mansoni or with Nippostrongylus brasiliensis and growth of the lymph node cells in the presence of the corresponding helminth antigen; (ii) immunization with horse serum and growth of blood and lymph node cells in the presence of the horse serum; (iii) exposure of T-cell-depleted suspensions of lymph node cells from unimmunized mice to T-cell factor (TCF) released into medium of the young cultures of (i) and (ii) . This differentiation was also obtained when lymph node cells from athymic nude mice were exposed to TCF . The cell suspensions were plated on X-irradiated fibroblast monolayers prepared from embryonic mouse skin . Screening of the suspensions before plating on the fibroblasts in culture revealed no young forms of mast cells, and none were present in culture of nude mice lymph node cells maintained without TCF . Primordial appearance of metachromatic granules generally in the golgi zone was first seen in many 'large lymphoid cells' as early as 18 hr after plating . This was followed by increase in the cytoplasm volume, increase in granule number and mitosis, ending at 10-18 days with homogeneous populations of mature mast cells . When the mesenteric lymph node cells from mice infected with the helminths were grown in the absence of fibroblasts but in the presence of the antigen, homogeneous populations of cells with extended cytoplasm, filled with unstained vacuoles developed during days 7-13 . These cells did not contain histamine (or at most 0.2 microgram per 10(6) vacuolated cells) . When these cells were plated on fibroblast monolayers clear granule formation in all the vacuoles was seen 2 days later . It increased progressively in size and staining intensity, until the vacuoles transformed into typical mast cell granules . By the fourth day the vacuolated cells attained the typical mast cell morphology and the histamine content greatly increased (from 0.12 microgram per 10(6) vacuolated cells to 3.02 micrograms per 10(6) mast cells) . These mast cells were readily degranulated by monoclonal anti-DNP-BSA IgE, and the antigen, releasing 90% of the histamine . The study shows that mucosal mast cells formation from 'large lymphoid-like' cells present in the blood and in the lymph, is stimulated by TCF . The condensation of the metachromatic material and histamine synthesis depends on other cells, presumably fibroblasts which comprise the principal cell in the embryonic skin monolayers . The mechanism of the fibroblast influence is not yet known.

Exp Hematol, 1983 Mar, 11(3), 243 - 8
Hemopoietic stromal precursors in long-term culture of bone marrow: II . Significance of initial packing for creating a hemopoietic microenvironment and maintaining stromal precursors in the culture; Chertkov JL et al.; Conversion of bone marrow cells to a single cell suspension prevents them from creating a hemopoiesis maintaining adherent cell layer (ACL) in culture . The ACL of such cultures is devoid of hemopoietic stromal precursors capable of transferring the hemopoietic microenvironment on implantation under the renal capsule of syngeneic mice . The regeneration of ACL after injury is possible in 1-wk-old, but not older, Dexter-type bone marrow cultures . The results prove the significance of the initial packing of the bone marrow cells in the differentiation of the hemopoietic stromal precursors . These data contradict previous reports which concluded that the structural orderliness of stromal tissue need not be preserved for the hemopoietic microenvironment to be transferred.

J Immunol Methods, 1983 Feb 25, 57(1-3), 301 - 9
A solid-phase immunoenzymatic technique for the enumeration of specific antibody-secreting cells; Sedgwick JD et al.; A sensitive immunoassay is described for the detection of idiotype- and isotype-specific antibody-secreting cells (ASC), based upon the well established principles of ELISA . Single cell suspensions containing ASC are incubated on solid phase to which specific antigen has been chemically conjugated . Antibody attaches to the latter within the immediate microenvironment of the ASC, producing localized zones of bound antibody which are subsequently developed as visual 'spots' in the ELISA . This assay system has sensitivity and specificity at least equivalent to haemolytic plaque assays.

Biochem J, 1983 Feb 15, 210(2), 509 - 15
Induction by growth factors of polysaccharide synthases in bean cell suspension cultures; Bolwell GP et al.; Suspension cells of bean subcultured into medium that maintains the culture and stimulates cell division but not differentiation brings about an increase in arabinan synthase activity . Subculture into a medium that induces both cell division and xylogenesis brings about in addition an increase in xylan synthase . Both synthases are membrane-bound and are concerned with the formation of neutral pectin or hemicellulose of the cell wall respectively . During the rising phase of the induction of these activities in the appropriate culture medium, the increases in activities were inhibited by either actinomycin D (an inhibitor of transcription) or D-2-(4-methyl-2,6-dinitroanilino)-N-methylpropionamide (an inhibitor of translation) . Thus the control for the induction of the enzyme activities involves transcription and possibly translation . Subculture of the cells brought about an increase, probably non-specific, in total membrane-bound translation, as indicated by increased amounts of bound polysomes and incorporation of {35S}methionine into membrane proteins . If the control of the appearance of specific mRNA molecules is partially effected by growth factors then these are probably operative during the period of the cell cycle that is stimulated by subculture and it is probably at this time that the growth factors act to bring about the changes necessary for differentiation.

Experientia, 1983 Feb 15, 39(2), 210 - 2
Pancreatic islet cell suspensions from newborn rats; different preparation procedures, viability and (pro)insulin biosynthesis; Schroder D et al.; Islet cell suspensions were prepared from neonatal rat pancreatic islets . While mechanical disintegration results in a higher yield, cells prepared by trypsin treatment appear to better preserved . Trypsin treatment of pancreatic islets during the cell preparation procedure does not influence the stimulation by glucose of (pro)insulin biosynthesis in freshly isolated cells.

Eur J Biochem, 1983 Feb 15, 130(3), 575 - 80
The influence of the ionic conductance on the relation between electron transport and proton-motive force in intact cells of Rhodopseudomonas capsulata; Clark AJ et al.; 1 . The dependence of membrane potential (delta psi) on the rate of respiration in darkened intact cell suspensions of Rhodopseudomonas capsulata was distinctly non-linear: severe inhibition of respiration with either rotenone or KCN led to only a small drop in delta psi . 2 . In the presence of 0.3 microMs carbonylcyanide p-trifluoromethoxyphenylhydrazone {CF3OPhzC(CN)2} the dependence of delta psi on respiratory rate became linear . Consequently, and particularly at lower concentrations of CF3OPhzC(CN)2, there was a pronounced, synergistic depression of the respiratory delta psi with CF3OPhzC(CN)2 and either rotenone or KCN . 3 . Antimycin A, at a concentration which strongly inhibited the photosynthetic electron transport chain, only slightly lowered the light-induced delta psi in anaerobic cell suspensions . Antimycin and CF3OPhzC(CN)2 synergistically lowered delta psi generated by illumination . 4 . The light-induced delta psi in anaerobic cells was only about 1.5-times larger than the respiratory-induced delta psi in darkened cells . Nevertheless it required approximately 16-times more CF3OPhzC(CN)2 to collapse the photosynthetic delta psi than the respiratory delta psi . 5 . These results are discussed with reference to the ionic current/delta psi relation described in {J.B . Jackson (1982) FEBS Lett . 139, 139-143} . The unifying feature is that the intrinsic conductance of the cell membrane is strongly dependent on delta psi but the conductance due to CF3OPhzC(CN)2 is independent of delta psi.

Biosci Rep, 1983 Feb, 3(2), 141 - 51
13C nuclear magnetic resonance studies of anaerobic glycolysis in Trypanosoma brucei spp; Mackenzie NE et al.; Anaerobic glycolysis in Trypanosoma brucei spp . has been studied by 13C NMR at 50 and 75.5 MHz . The uptake of {U-13C}glucose by cell suspensions of T . b . brucei was monitored by time-course spectroscopy, and while no anomeric specificity was found, the end-products of glycolysis were confirmed as glycerol and pyruvate together with alanine and dihydroxypropionate . The intermediacy of L-glycerol-3-phosphate was also ascertained . The incorporation of C-1 of {1-13C}glucose and of C-6 of {6-13C}glucose into glycerol and pyruvate in T . b . gambiense was quantified by measurement of the longitudinal relaxation times of the species involved . An incorporation to the extent of 66% of each substrate into equimolar amounts of glycerol and pyruvate indicate that Keq for the triosephosphate-isomerase-mediated reaction approaches unity.

Cancer Res, 1983 Feb, 43(2), 617 - 22
Orthotopic implantation of primary N-{4-(5-Nitro-2-furyl)-2-thiazolyl}formamide-induced bladder cancer in bladder submucosa: an animal model for bladder cancer study; Ibrahiem EH et al.; Primary bladder tumors induced in Fischer 344 inbred rats by N-{4-(5-nitro-2-furyl)-2-thiazolyl}formamide were transplanted in syngeneic rats by the intravesical, s.c., i.v., and orthotopic routes . Attempts were made to establish bladder cancer cell lines in vitro . No success was achieved in transplantation by either the s.c., i.v., or intravesical routes when primary tumor cells were transplanted as cell suspensions . Cell suspensions of primary tumors also failed to grow in culture . However, orthotopic implantation into the bladder submucosa gave 45% success . Tumor fragments obtained from either the primary tumor or its lung metastases resulted in 10.6 and 36% tumor takes, respectively, when implanted s.c . However, after one orthotopic passage in the bladder submucosa, the tumor cells injected as cell suspension grew s.c . in 14% and orthotopically in 79% of the animals . Tumor fragments obtained from orthotopic tumors and implanted s.c . resulted in 15% tumor takes . After the second orthotopic passage, tumor cells could be grown in cultures and orthotopically in 100% of animals . The technique of orthotopic implantation as well as the usefulness of this tumor model for bladder cancer studies are described.

Cancer Res, 1983 Feb, 43(2), 536 - 40
Relationship between cellular procoagulant activity and metastatic capacity of B16 mouse melanoma variants; Gilbert LC et al.; The metastatic process is a complex sequence of steps that may involve coagulation and the presence of fibrin . F1 (low incidence of lung colonization) and F10 (high incidence of lung colonization) variants of the B16 mouse melanoma were used to examine the relationship between the level of cellular procoagulant activity and their metastatic potential . Cell suspensions were prepared from cultures of B16-F1 and B16-F10 cell lines . Aliquots (0.2 ml) containing 50,000 cells were assayed for procoagulant activity in recalcified citrated rat plasma and for metastatic capacity by tail vein injection followed by counting of melanotic lung tumor colonies 17 days later . In one series of experiments, procoagulant activity and metastatic capacity were determined at 1, 2, 3, and 4 days after plating . The data showed an almost parallel decrease in both characteristics as the culture density increased . To examine the correlation between cellular procoagulant activity and the metastatic capacity of the B16 variants in two different series of experiments, regression analysis of the level of procoagulant activity and the number of lung tumor colonies gave correlation coefficients of 0.9 (n = 15) and 0.79 (n = 8) . These results suggest that fibrin formation resulting from cellular procoagulant activity may play a role in the metastatic process.

J Gen Physiol, 1983 Feb, 81(2), 283 - 304
Permeability of human red cells to a homologous series of aliphatic alcohols . Limitations of the continuous flow-tube method; Brahm J; Human red cell permeability to the homologous series of methanol, ethanol, n-propanol, n-butanol, and n-hexanol was determined in tracer efflux experiments by the continuous flow tube method, whose time resolution is 2-3 ms . Control experiments showed that unstirred layers in the cell suspension were less than 2 X 10(-4) cm, and that permeabilities less than or equal to 10(-2) cm s-1 can be determined with the method . Alcohol permeability varied with the chain length (25 degrees C): Pmeth 3.7 X 10(-3) cm s-1, Peth 2.1 X 10(-3) cm s-1, Pprop 6.5 X 10(-3) cm s-1, Pbut less than or equal to 61 X 10(-3) cm s-1, Phex 8.7 X 10(-3) cm s-1 . The permeability for methanol, ethanol, and n-propanol was concentration independent (1-500 mM) . The permeability to n-butanol and n-hexanol, however, increased above the upper limit of determination at alcohol concentrations of 100 and 25 mM, respectively . The activation energies for the permeability to methanol, n-propanol, and n-hexanol were similar, 50-63 kJ mol-1 . Methanol permeability was not reduced by p-chloromercuribenzene sulfonate (PCMBS), thiourea, or phloretin, which inhibit transport of water or hydrophilic nonelectrolytes . It is concluded (a) that all the alcohols predominantly permeate the membrane lipid bilayer structure; (b) that both the distribution coefficient and the diffusion coefficient of the alcohols within the membrane determine the permeability, and (c) that the relative importance of the two factors varies with changes in the chain length.

J Gen Physiol, 1983 Feb, 81(2), 239 - 53
Reflection coefficient and permeability of urea and ethylene glycol in the human red cell membrane; Levitt DG et al.; The reflection coefficient (sigma) and permeability (P) of urea and ethylene glycol were determined by fitting the equations of Kedem and Katchalsky (1958) to the change in light scattering produced by adding a permeable solute to a red cell suspension . The measurements incorporated three important modifications: (a) the injection artifact was eliminated by using echinocyte cells; (b) the use of an additional adjustable parameter (Km), the effective dissociation constant at the inner side of the membrane; (c) the light scattering is not directly proportional to cell volume (as is usually assumed) because refractive index and scattering properties of the cell depend on the intracellular permeable solute concentration . This necessitates calibrating for known changes in refractive index (by the addition of dextran) and cell volume (by varying the NaCl concentration) . The best fit was for sigma = 0.95, Po = 8.3 X 10(-4) cm/s, and Km = 100 mM for urea and sigma = 1.0, Po = 3.9 X 10(-4) cm/s, and Km = 30 mM for ethylene glycol . The effects of the inhibitors copper, phloretin, p-chloromercuriphenylsulfonate, and 5,5'-dithiobis (2-nitro) benzoic acid on the urea, ethylene glycol, and water permeability were determined . The results suggest that there are three separate, independent transport systems: one for water, one for urea and related compounds, and one for ethylene glycol and glycerol.

Am J Hematol, 1983 Feb, 14(1), 27 - 36
Mechanism of canine cyclic hematopoiesis: the role of prostaglandin E in feedback regulation; Hammond WP et al.; Prostaglandin E inhibits granulocyte-macrophage colony formation in vitro in man and mouse, suggesting that it plays a role in feedback regulation of granulocyte production in vivo . Therefore, we examined the role of PGE in normal canine hematopoiesis and its potential role in the pathogenesis of cyclic hematopoiesis in grey collie dogs . The prostaglandin synthesis inhibitors indomethacin and ibuprofen (10(-5) M) increased CFU-C growth to 194 and 160% of control, respectively, while PGE2 addition caused a dose-dependent inhibition of bone marrow CFU-C growth in both normal and grey collie dogs . These concentrations of indomethacin and ibuprofen decreased bone marrow cell elaboration of PGE measured by radioimmunoassay to less than 5% of control values . The levels of PGE in leukocyte conditioned medium prepared from grey collies correlated with the number of monocytes in the conditioning cell suspension (r = 0.78, n = 10, p less than 0.05) so that PGE production per monocyte was no different in normal and grey collie dogs . The effect of PGE2 upon CFU-C was to inhibit formation of macrophage, but not neutrophil colony subtypes . These findings make aberrant PGE-mediated inhibition of precursor cells an unlikely mechanism to cause cyclic hematopoiesis, and show that PGE produced by monocytes acts as a feedback inhibitor for precursor cells destined to produce monocytes but not for those destined to form neutrophils.

Scand J Immunol, 1983 Feb, 17(2), 123 - 8
Quantitation and estimation of cooperation between target-binding sites in natural and antibody-dependent cell-mediated cytotoxicity by use of a mathematical transformation . Influence of interferon; Krebs HJ et al.; We have studied the use of a mathematical transformation, the Hill transformation, in natural and antibody-dependent cell-mediated cytotoxicity as a function of effector cell concentration . It can be concluded that when adherent cells are removed from the effector cell suspension, cooperation between binding sites for effector cells on the target cell changes from negative to zero . After stimulation of nonadherent cells with alpha-interferon, the binding sites still remain noninteracting . This is the case both in natural and in antibody-dependent cell-mediated cytotoxicity experiments . The Hill transformation seems to be an easy and easily reproducible way of quantitating cytotoxicity.

Int J Radiat Oncol Biol Phys, 1983 Feb, 9(2), 217 - 20
Response of fibrosarcoma cell subpopulations to small doses of radiation delivered in situ; Thames HD Jr et al.; The survival of cells from 2 tumor subpopulations after gamma-ray doses ranging from 1 to 19 Gy was determined using a lung colony assay . Methylcholanthrene-induced fibrosarcomas grown in the hind legs of C3H/Kam pathogen-free mice were irradiated in situ when the tumors were 8-10 mm in diameter . Single cell suspensions prepared from excised tumors were separated on a linear density gradient, and the clonogenicity of predominantly oxic Band 2 (density 1.08 g/cm3) and predominantly hypoxic Band 4 (density 1.14 gm/cm3) cells was measured . The surviving fraction of cells after doses of 1, 2, and 3 Gy was estimated from that measured after total doses of 5 Gy = 5 X 1 Gy, 10 Gy = 5 X 2 Gy, and 15 Gy = 5 X 3 Gy, under the assumption of equal effect per fraction (checked by estimating survival at 3 Gy after different numbers of fractions) . Very little curvature was evident in the survival curves of Band 2 and Band 4 cells (beta/alpha = .013-.034 Gy-1) . The initial segment of the survival curve of the predominantly oxic Band 2 cells was steeper (1Do = 3.6 Gy) than that of the predominantly hypoxic Band 4 cells (1Do = 5.2 Gy); both remained linear over a large range, to doses in excess of 3 Gy . These results imply that these tumor subpopulations will be insensitive, in their response to multifractionated regimens, to changes in size of dose per fraction in the range 0 to 3 Gy, a trait shared by two acutely responding normal tissues (murine testis and jejunum).

J Bacteriol, 1983 Feb, 153(2), 916 - 20
A voltage clamp inhibits chemotaxis of Spirochaeta aurantia; Goulbourne EA Jr et al.; Anaerobic conditions were employed to study the relationship between membrane potential and chemotaxis in Spirochaeta aurantia . When cells were grown anaerobically and suspended in anaerobic potassium phosphate buffer (pH 5.5), membranes did not appear to be polarized . Nevertheless, motility was supported by a transmembrane pH gradient, and the anaerobic cells exhibited D-xylose taxis . Introduction of trace amounts of air into anaerobic cell suspensions resulted in a transient membrane polarization . The addition of valinomycin to cells suspended under anaerobic conditions did not alter the steady-state value of membrane potential appreciably but served to clamp membrane potential at the preset level . Although there was no detectable effect of valinomycin on the motility of anaerobic cells in potassium phosphate buffer, D-xylose taxis was completely inhibited by this treatment . These data indicate the the action of valinomycin as a voltage clamp serves to inhibit the chemotaxis of S . aurantia and provide evidence to support the suggestion that the mechanism of chemotaxis in this organism involves the transduction of sensory signals in the form of membrane potential fluctuations.

Vet Med (Praha), 1983 Feb, 28(2), 119 - 25
{The effect of different cell concentrations and incubation time of testicular tissue testosterone synthesis}; Mamode MI et al.; The cells of the testes of mice were enzymatically separated and the amount of Leydig cells was determined in different stages of dispersion and after washing the separated tissue . This amount ranged from 2 to 9% . The largest quantity of Leydig cells (9%) was obtained when separated tubuli were washed . The response of cell suspensions at seven concentrations stimulated by human chorion gonadotrophin (HCG) was tested . Cell suspension concentrations from 5.3 x 10(5) to 1.4 x 10(6) cells/ml were found to be satisfactory . After two to three hours of incubation, the values of testosterone could be used for the determination of the activity of gonadotropic preparations.

Toxicol Appl Pharmacol, 1983 Feb, 67(2), 292 - 301
Interactions of aflatoxin B1 and blood components of various species in vitro: interconversion of aflatoxin B1 and aflatoxicol in the blood; Kumagai S et al.; The fate of aflatoxin B1 (AFB1) in the blood of various species of animals was studied in vitro . Examination of the distribution of radioactivity in blood incubated with {14C}AFB1 at 37 degrees C showed that high levels of radioactivity were associated with blood cells . The radioactivity was readily removed from the blood cells by washing with fresh plasma, indicating loose binding of AFB1 to blood cells . Most of the radioactivity in plasma was bound to protein . These results suggest that a large part of the AFB1 in blood in vivo may be carried not only by the plasma proteins but also by the blood cells . When chloroform extracts of plasma of {14C}AFB1-treated mouse, rat, duckling, and hamster blood were developed by thin-layer chromatography, high levels of radioactivity were found in both the AFB1 region and the aflatoxicol (AFL) region . Incubation of blood with nonradioactive AFB1 and AFL showed marked interconversion of AFB1 and AFL in the blood of rats, hamsters, mice, and Mongolian gerbils, but not in the blood of guinea pigs, rhesus monkeys, squirrel monkeys, or humans . Interconversion occurred in red blood cell suspensions but not in plasma, indicating that the red blood cells are responsible for AFB1-AFL interconversion in the blood.

J Immunol, 1983 Feb, 130(2), 951 - 7
Studies of allogeneic tumor transplants: induced rejection of advanced tumors by immune alteration of recipients; Russell PS et al.; In the present experiments, a methylcholanthrene-induced sarcoma (S-702) of B10.D2 origin was found to grow rapidly in B6AF1 mice leading to the death of all recipients in 5 to 9 wk . Nevertheless, immunity to MHC antigens presented by the tumor was readily demonstrable in tumor-bearing mice by their responses to donor strain skin grafts until late in the course of tumor growth, when a nonspecific form of immune suppression developed . In addition, B6AF1 mice preimmunized by exposure to B10.D2 donor strain antigens did not permit tumor growth . Treatment of tumor-bearing B6AF1 mice with CY at 18 days, when the tumors measured over 12-mm in diameter, followed by the i.p . injection of B10.D2 lymphoid cells (at a dosage of from 1.2 to 2.5 X 10(8) cells) resulted in the complete regression of 100% of these large tumors . CY treatment combined with localized immune stimuli in the form of donor strain skin grafts or secondary tumor implants was incapable of producing a sufficiently heightened immune response to cause tumor rejection . A dose of CY temporarily retarded tumor growth in most mice, and in a minority of animals so treated (less than 25%) tumors regressed completely . In syngeneic (B10.D2) animals, CY also temporarily slowed tumor growth, but total regression was never observed . An effective B10.D2 cell inoculum could consist not only of living lymphoid cells but of irradiated (1000 rad) cells as well . Tumor cell suspensions (after irradiation, 10,000 rad) were also effective . These observations suggest local immune factors at the host-tumor interface may have been of importance in the survival of these allogeneic tumor transplants and that CY influenced this state, perhaps through an influence on suppressor cells, allowing subsequent administration of donor strain cellular antigens to induce an effective tumor rejection response.

J Steroid Biochem, 1983 Feb, 18(2), 205 - 8
Angiotensin II potentiates ACTH-stimulated adrenal androgen secretion; Parker LN et al.; Several lines of evidence in animal and human studies, including measurements of elevated adrenal androgens in salt wasting syndromes suggest that adrenal androgen levels may in part be regulated by angiotensin II . This possibility was tested directly by measurement of cortisol and dehydroepiandrosterone (DHA) in canine adrenal cell suspensions after addition of angiotensin II and ACTH, separately and together . Minimal cortisol and no DHA stimulation was obtained using 4 x 10(-10) to 4 x 10(-5) M concentrations of angiotensin II alone . However, increased cortisol and DHA secretion were observed beginning at 4 x 10(-10) M concentrations of angiotensin II when it was combined with 10(-13) MACTH . These are physiological concentrations of both stimuli and may explain the increased adrenal androgens seen in some pathological situations characterized by elevated PRA.

Cell Immunol, 1983 Feb 1, 75(2), 348 - 55
Membrane phenotype of murine effector and suppressor T cells involved in delayed hypersensitivity and protective immunity to herpes simplex virus; Nash AA et al.; The membrane phenotype of T cells involved in delayed hypersensitivity (DH), protective immunity, and suppression of delayed hypersensitivity to herpes simplex virus (HSV) has been determined . T cells from immune lymph nodes transferring DH and antiviral immunity to normal recipients were characterized as Lyt 1+2- . There appeared to be no detectable antiviral role for Lyt 1-2+ cells in the transferred cell suspension . Splenic T cells suppressing the induction of DH to HSV were characterized as being both Lyt 1+2- and Lyt 1-2+ 4 weeks after their induction . At earlier times, i.e., after 7 days, the suppression was mediated solely by the Lyt 1+2- population . Thereafter, a progressive increase in the contribution of the Lyt 1-2+ suppressor was observed . Both the early and later phases of suppression were due to I-J positive cells . The nature of the two suppressor cell types is discussed in relation to suppressor cell "cascades" and to the pathogenesis of herpes simplex virus infection.

J Cutan Pathol, 1983 Feb, 10(1), 33 - 51
Flow cytometry of keratinocytes; Clausen OP; A prerequisite for using flow cytometry (FCM) is the availability of isolated single cells . Procedures for separation and isolation of keratinocytes from animals and man are available, and the resulting single cell suspensions have been subjected to FCM measurements . The major advantage of the method is the accuracy and speed with which a variety of cellular constituents can be quantified . FCM of keratinocytes has, hitherto, been mainly confined to measurements of nuclear DNA for estimation of cell-cycle distributions and for ploidy studies . In mouse epidermis, cell-cycle distributions were estimated from sequentially obtained DNA histograms and evaluated with other cell kinetic measurements, resulting in new information about epidermal cell-cycle progression, not achievable by any of the methods alone . The best way, therefore, to increase our knowledge of keratinocyte proliferation, is the combined use of DNA FCM and other cell kinetic methods . DNA FCM has also been applied to healthy and diseased human epidermis, and may add valuable information to the classification of skin disease in selected cases . It is believed that further progress in the characterization of keratinocyte growth and development will depend on parameters other than DNA alone.

J Biol Chem, 1983 Jan 10, 258(1), 636 - 42
Tissue distribution, structural characterization, and biosynthesis of Mac-3, a macrophage surface glycoprotein exhibiting molecular weight heterogeneity; Ho MK et al.; Mac-3 is a mouse macrophage differentiation antigen defined by a rat anti-mouse monoclonal antibody (MAb),M3/84 . The structure, biosynthesis, quantitative surface expression, and distribution of Mac-3 have been studied by radiolabeling and isolation with MAb-Sepharose, saturation binding, absorption, and immunofluorescence flow cytometry . In sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Mac-3 migrates as a diffuse band with average Mr = 110,000 . Labeling of intact cells with 125I and accessibility to MAb show it is present at least in part on the cell surface . Saturation labeling with 125I-MAb shows 4.2 X 10(4) cell surface sites on thioglycollate medium-elicited peritoneal macrophages . {35S}Methionine and {3H}glucosamine incorporation into Mac-3 by purified macrophages show it is a glycoprotein synthesized by these cells . Absorption shows Mac-3 is strongest in macrophages, present in lower quantities in lung, liver, bone marrow, and spleen, and undetectable in thymus, lymph node, brain, and heart . Immunofluorescent flow cytometry shows surface expression on thioglycollate-elicited macrophages but not bone marrow, spleen, lymph node, or thymus cell suspensions . Similar amounts of Mac-3 are immunoprecipitated from resident macrophages or macrophages elicited by sterile inflammatory agents, intracellular parasites, or immunomodulators, but the average Mr of Mac-3 varies from 92,000 to 110,000 . Mac-3 is synthesized from precursor(s) of Mr = 74,000 and 79,000, identical in the different macrophages . Processing into the mature molecule, which migrates in sodium dodecyl sulfate-polyacrylamide gel electrophoresis as a more diffuse band and varies in Mr among macrophage elicited by different agents and to a lesser degree between different preparations of the same type of macrophage, occurs in 15 to 30 min.

Cell Tissue Res, 1983, 232(3), 539 - 52
Ultrastructural and functional development of macrophages in the dermal tissue of rat fetuses; Takahashi K et al.; After about 12 days of gestation, fetal macrophages begin to appear in the subepidermal mesenchyme of rat fetuses . The macrophages are ultrastructurally characterized by cytoplasmic vacuoles, abundant polyribosomes and long filopodia . Immunocytologically, they possess Fc and complement (C3) receptors on the cell surface and are capable of immune phagocytosis, Latex or carbon phagocytosis, and glass adherence . From 15 days of gestation, lysosomal granules and micropinocytic vesicles gradually develop, together with an enlargement of Golgi complexes, whereas the number of polysomes and the number and size of cytoplasmic vacuoles are gradually reduced when gestation ends . Finally, the macrophages become amoeboid . Non-specific esterase and endogenous peroxidase activities are always absent in these macrophages . In culture experiments with cell suspensions prepared from the mesenchyme, fetal macrophages show a similar maturation process . Autoradiography with 3H-thymidine demonstrates a high proliferative capacity of the macrophages, particularly during the fetal stage.

Hemoglobin, 1983, 7(3), 205 - 26
Characterization and properties of Hb York (beta 146 His leads to Pro); Kosugi H et al.; A second case of Hb York (beta 146 His leads to Pro), was discovered in a patient with polycythemia . The oxygen equilibrium curves (OEC) of red cell suspensions in a buffer (pH 7.4) at 37 degrees C revealed a biphasic curve with a P50 of only 12.5 mm Hg (normal value: 26.5 +/- 1.0 mm Hg) . The purified Hb York had an extremely high affinity for oxygen with diminished cooperativity and decreased Bohr effect . The oxygen affinity was significantly reduced by inositol hexaphosphate . Molecular stability studies by mechanical shaking of various liganded forms of Hb York revealed stabilities between those of Hb A and Hb S . Isolated beta Y-subunits were more unstable than beta A-subunits at every pH examined . Hb York was 1.4 times more unstable than Hb A in 18.9% isopropanol.

Biorheology, 1983, 20(1), 41 - 56
Theoretical modeling of filtration of blood cell suspensions; Skalak R et al.; A theoretical model of filtration of suspensions containing red blood cells (RBCs) and white blood cells (WBCs) has been developed . Equations are written for the pressure drop, the filtration flow and the fractions of filter pores containing RBCs (alpha) and WBCs (alpha*) . Because the relative resistances (ratios of resistance of cell to resistance of suspending fluid) of RBCs (beta) and WBCs (beta*) through the filter pore are greater than one, the transit of these cells (especially WBCs) through the filter is slower than that of suspending fluid; this leads to values of alpha and alpha* higher than those simply expected from the hematocrit and leukocrit, respectively, in the entering and exiting suspensions . In the absence of pore plugging by the cells (steady flow), the pressure drop can be computed from alpha, alpha*, beta and beta* . In order to model unsteady flow, differential equations are written to include pore plugging and the subsequent unplugging by the rising filtration pressure at a constant flow . By specifying the fractions of entering RBCs (epsilon) and WBCs (epsilon*) which would plug the pores and the rate at which the plugged pores would unplug in response to pressure rise (epsilon u), as well as the fractions of entering RBCs (epsilon p) and WBCs (epsilon p*) that would plug the pores permanently, theoretical pressure-time curves can be generated by numerical integration, and the results fit the experimental data well . From such fitting of theoretical curve to experimental data, information can be deduced for epsilon, epsilon*, epsilon u, epsilon p and epsilon* p.

Biorheology, 1983, 20(1), 29 - 40
Influence of red cell concentration on filtration of blood cell suspensions; Schmalzer EA et al.; Pressure-time curves obtained by passing suspensions of blood cells in Ringer solution through a 5 microns polycarbonate filter at constant flow (1.6 ml/min) were evaluated for their ability to reflect the deformability of the erythrocytes . The initial pressure reading (Pi) obtained in a quasi-steady state during the first 1-2 sec of pumping was found to be reproducible for hematocrit values between 10 and 30 percent . This Pi value was normalized by the pressure generated by the cell-free suspending medium (PO) at the same flow rate . The ratio Pi/PO was found to be linearly proportional to hematocrit up to 30 percent but independent of leukocyte concentration up to 12,000/mm3 . Later portions of the curve did vary with leukocyte count . By using the equations developed from theoretical modeling of cells passing through a filter, the experimentally determined relation of Pi/PO to hematocrit, and the known geometry of the filter pores, we were able to calculate parameters reflecting the deformability of red cells . These include beta, the ratio of resistance in a pore containing a red cell to that in a pore containing only the suspending medium, and alpha, the proportion of pores filled by erythrocytes in transit . The application of theoretical analysis to experimental data has provided quantitative insights into the behavior of red cells during filtration tests in normal and disease states.

Biorheology, 1983, 20(1), 11 - 27
Role of white blood cells in filtration of blood cell suspensions; Chien S et al.; The time-dependent filtration pressure curves of cell suspensions pumped through 5 microns polycarbonate filters at a constant flow rate were analyzed with the aid of a theoretical model developed in an accompanying paper . The cell suspensions contained mixtures of erythrocytes and leukocytes, the concentrations of which were systematically varied . The pressure-time (P-t) curves generally showed multiphasic components . Following the attainment of a quasi-steady state level, the pressure rose first rapidly and then more slowly . The rates of pressure rise in the fast and slow phases were normalized by using the steady state pressure reading (PO) obtained with Ringer solution at the same flow rate, and are designated k1 and k2, respectively . Both k1 and k2 increased with rising concentrations of leukocytes, {WBC}, or erythrocytes, {RBC} . {WBC} is 700-1000 times more effective than {RBC} in affecting k1 and k2 . k1 is related to the dynamic plugging and unplugging of filter pores, primarily by leukocytes . k2 is attributable to the "permanent" plugging of filter pores, again predominantly by leukocytes . The experimental P-t curves can be fitted with the theoretical model by using appropriate constants for leukocyte plugging . The results indicate that nearly 2/3 of the entering leukocytes cause transient plugging of pores, with an unplugging rate of 4.1 percent/sec/unit pressure rise, and that approximately 2.2 percent of the entering leukocytes are "permanently" lodged . These results underscore the important role of leukocytes in determining the later phase of the P-t curve and support the concept that leukocyte plugging may have pathophysiological significance in causing microvascular occlusion in disease states.

Nauchnye Doki Vyss Shkoly Biol Nauki, 1983, (1), 85 - 8
{Effect of carbon monoxide on the growth of sulfate-reducing bacteria and their oxidation of this substrate}; Karpilova IIu et al.; The effect of carbon monoxide on the growth of six strains of sulfatee-reducing bacteria have been studied as well as the ability of bacterial suspensions and extracts to oxidize CO . It was shown that sulfate-reducing bacteria possess a comparably high resistance to carbon monoxide . There are difference in the sensitivity of certain species and strains of sulphat-reducing bacteria to the content of CO in the gas phase . The cell suspensions and extracts are capable of oxidizing 100% CO in gaseous phase . The rate of carbon monoxide oxidation by extracts is much higher than by suspensions.

Invasion Metastasis, 1983, 3(4), 243 - 8
Plasminogen activators as markers of tumor colonization potential; Ng R et al.; Cell suspensions from the R 3230 AC rat mammary adenocarcinoma, when injected intravenously into F344 rats, invariably produce multiple lung foci within 10 days . We compared the colonization potential of cultures obtained from these foci and from cell populations exposed to 100 micrograms/ml medium of both concanavalin A and wheat germ agglutinin for 5 passages with the original cell line . Plasminogen activator activity (PAA) was determined in all three cell subpopulations, using S2251 (KABI) as chromogenic substrate . All cell lines retained their ability to grow after subcutaneous implant . The lectin resistant variant was found to have lost its capacity to nidate in the lung completely and also had the lowest PAA . In contrast, the cell population derived from the lung foci ranked highest in PAA.

Biorheology, 1983, 20(5), 557 - 67
Shear viscoelasticity of suspensions of biological cells with fluid membrane; Takano Y et al.; In order to consider the effect of membrane fluidity upon the mechanical property of biological cell suspensions, we have calculated the complex intrinsic viscosity {eta*} = {eta'} - i{eta"} of spherical shell structures with material incompressibility in suspension as a function of the dimensionless frequency x = omega eta'a/gamma' together with the parameters of hm = eta m/eta', g = gamma/gamma', h = eta/eta', delta = d/a, where a is the radius of the cell, d is the width of the membrane, eta, eta m, and eta' are the viscosities of the medium, of the membrane and of the internal region of the cell, gamma and gamma' the surface tensions at the outer and the inner side of the membrane respectively, and omega the angular frequency . The result is simply represented by two dispersions as follows: {eta*}/{eta} = A1 + B1/(1 + i omega tau 1) + B2/(1 + i omega tau 2) . Here i is the imaginary unit, A1 = 2(1 - h)/(2 + 3h) + O(delta), B1 = 3h/(5 + 5h) + O(delta), B2 = h (19 + 16h)/{5(1 + h) (2 + 3h)} + O(delta), tau 1 = {(5/24) (1 + h) (1 + 1/g) delta-2 + O(delta-1)} a eta'/gamma', tau 2 = {(2 + 3h) (19 + 16h)/{40 (1 + h) (1 + g)} + O(delta)} a eta'/gamma', and {eta} = (5/2) {96hmg + 32g (5 + 5h - 12hm)delta + O(delta 2)}/{96hmg + 32g (5 + 2h - 12hm) delta + O(delta 2)}.

Biorheology, 1983, 20(5), 471 - 83
Theory of non-Newtonian viscosity of red blood cell suspension: effect of red cell deformation; Murata T; The effects of the deformation of red blood cells on non-Newtonian viscosity of a concentrated red cell suspension are investigated theoretically . To simplify the problem an elastic spherical shell filled with an incompressible Newtonian fluid is considered as a model of a normal red cell . The equation of the surface of the shell suspended in a steady simple shear flow is calculated on the assumption that the deformation from a spherical shape is very small . The relative viscosity of a concentrated suspension of such particles is obtained based on the "free surface cell" method proposed by Happel . It is shown that the relative viscosity decreases as the shear rate increases.

Biomed Biochim Acta, 1983, 42(11-12), S332 - 6
CDS-AG medium for red blood cell preservation; Strauss D; Buffy coat-free red cell concentrates (RCC) from ACD, ACD-AG, and EDTA blood were prepared at the day of collection and resuspended with about one half of the packed red cell volume of a citrate-dextrose-sucrose-adenine-guanosine solution (CDS-AG) . The posttransfusion survival of RBC in red cell suspensions (RCS) amounted to about 80% after 35 days of storage . 50 to 63% of the normal ATP level still remained . About 0.3% of red cells were hemolysed . 10% of red cells existed unchanged as discocytes, about 85% were transformed to echinocytes and 5% to spherocytes . The MCHC increased by about 25% due to a shrinkage of RBC . A much more hypertonic citrate-saline anticoagulant solution (420 mosmol/kg) increased the shrinkage, led to a higher hemolysis and lower posttransfusion survival rates.

Biorheology, 1983, 20(3), 311 - 6
Indices of filterability of red blood cell suspensions; Skalak R et al.; A number of different experimental techniques have been devised in recent years to use microsieving as a test of the filterability of suspensions of red blood cells . Various indices have been proposed to express the results of these tests . In the present paper a correlation is made of the intrinsic increase in resistance at the level of a single pore in the filter to the macroscopically observed pressure and flow through the entire filter . Further it is shown how a number of different tests may be used to derive the same index . The results apply only to situations in which there is no plugging of pores.

Biorheology, 1983, 20(3), 295 - 309
The bulk rheology of close-packed red blood cells in shear flow; Secomb TW et al.; A theoretical analysis is made of the dynamical behavior and bulk rheology of close-packed red blood cell suspensions subjected to simple shear flow . The model for the polyhedral cell shapes and tank-treading membrane motion developed in the companion paper (1) is used . The flow in the thin lubricating plasma layers between cells is analyzed taking into account the mechanical properties of the membrane at the corner regions of sharp membrane curvature . This leads to predictions for the apparent viscosity as a function of hematocrit and shear rate . Good agreement with experimental results is obtained at moderate and high shear rates (above 20 s-1) . At lower shear rates, a rapid rise in apparent viscosity has been found experimentally, and the mechanisms leading to this behavior are examined.

Biorheology, 1983, 20(3), 283 - 94
The motion of close-packed red blood cells in shear flow; Secomb TW et al.; Experimental and theoretical results are presented concerning the motion of close-packed red blood cell suspensions subjected to steady simple shear flow . The behavior of the suspension was observed microscopically using a cone-and-plate rheoscope . At moderate and high shear rates the cells show a fairly orderly arrangement, each appearing polygonal in the field of view . An idealized theoretical model for the suspension is developed, in which each cell is a 14-sided polyhedron of varying shape, but with constant surface area and volume . Tank-treading motion of the membrane is predicted, and an approximation to the motion is calculated which is consistent with the known mechanical properties of the membrane . It is shown that considerably more energy is dissipated in the membrane than in the cytoplasm during tank-treading.

Ann Rech Vet, 1983, 14(2), 163 - 8
A laboratory reference vaccine to titrate immunogenic activity of antibrucella vaccines in mice; Bosseray N et al.; The number of bacteria (CFU) in spleens of mice fifteen days after a standard intraperitoneal challenge of Brucella abortus, is dependent on vaccinal immune status of mice . From this observation, a control method of vaccinal activity was previously proposed, which classified the responses in relation to fixed reference values . A reference vaccine, a lyophilized formalin-killed bacterial cell suspension of B . melitensis strains H38 was prepared and titrated . From the dose response curve, the quantities giving reference values were calculated and expressed in a Unit system . This vaccine makes possible inter-laboratory comparisons of vaccinal activity, that can be expressed on the Units basis.

Arch Immunol Ther Exp (Warsz), 1983, 31(2), 183 - 9
The use of protein beads as immunoadsorbent for the column fractionation of lymphocytes; Eckert R et al.; Beads of calf serum proteins (CSB) with antigen-antibody complexes or antigen by activation with glutaraldehyde were used as immunoadsorbents for the fractionation of mouse lymphocytes . T-lymphocytes could be separated by this method with high purity . The precursors of antibody-forming cells were completely eliminated from spleen and bone marrow cell suspensions . The unspecific binding of lymphocytes to CBS or Sepharose was similar and not selective for T or B lymphocytes.

Int J Immunopharmacol, 1983, 5(4), 283 - 8
The in vitro effect of phenytoin and carbamazepine on subpopulations of human blood mononuclear cells; Gilhus NE; Mononuclear cells from peripheral blood of 10 healthy blood donors were incubated with phenytoin and carbamazepine dissolved in 1% propyleneglycol . Phenytoin 20 mg/1 significantly reduced the percentage of active E rosette-forming T-lymphocytes; to mean 17.6% compared to mean 25.9% in control incubations (P less than 0.05) . Carbamazepine 10 mg/1 reduced this percentage to 21.0% (0.05 less than P less than 0.10) . Phenytoin reduced the percentage of active T-lymphocytes in 9 of the 10 cell suspensions, while carbamazepine reduced the active T-lymphocytes in 8 of the 10 suspensions . The percentage of total E rosette-forming T-lymphocytes was similarly reduced by 20 mg/1 phenytoin and 10 mg/1 carbamazepine; from 65.3 to 59.5% and to 60.3%, respectively (P less than 0.05, 0.05 less than P less than 0.10) . The reduction of active and total T-lymphocytes was concentration-dependent for both drugs . Phenytoin and carbamazepine 5 mg/1 did not influence the percentage of T-lymphocytes . The decrease became apparent at a concentration of 10 mg/1 for both drugs, and grew steadily more marked with increasing drug concentrations up to 80 mg/1 . However, the reduction in the percentage of T-lymphocytes was more pronounced for phenytoin than for carbamazepine at all drug concentrations . The percentage of cells with Fc gamma-receptors and of those with receptors for the activated human complement component C3b, were not influenced by phenytoin and carbamazepine, nor were the cells with phagocytizing capacity and the non-phagocytizing cells with membrane-bound immunoglobulin (i.e . B-lymphocytes).

Z Rechtsmed, 1983, 90(2), 127 - 36
{Enzyme activity of isolated leukocyte populations . II . Cytochemical and zymographic studies of cadaver blood}; Oehmichen M et al.; Blood was taken from the femoral vein of 17 autopsied cadavers with different diagnoses and postmortal intervals (10-120 h) . The granulocytes and lymphocytes were isolated with routine methods . The cell suspensions were subject to morphologic, enzyme-cytochemical (naphthol AS-D chloroacetate esterase) and electrophoretic (PGM1, PGM3, GOTM, PEPA, MEM, FUCA) investigations . The following results were obtained: Erythrocyte-free cell suspensions are only found during short postmortal intervals; Isolation of pure lymphocytes from cadaver blood using the described method is only possible very early in the postmortal interval (within 10 h); The isolated granulocytes contain an impressively high percentage of eosinophilic granulocytes; The percentage of naphthol AS-D-chloroacetate esterase-positive granulocytes is considerably lower in stored, conserved blood than in blood smears; Identification of the above mentioned enzymes from isolated granulocytes from all cadavers is possible by electrophoresis.

Z Rechtsmed, 1983, 90(2), 115 - 25
{Enzyme activity of isolated leukocyte populations . I . Cytochemical and zymographic studies of stored blood under various storage conditions}; Kompf J et al.; Heparinized venous blood was stored under sterile conditions at different temperatures (4 C, 20 C, 37 C) for various intervals (0-7 days) . After storage the granulocytes and lymphocytes were isolated with routine methods . Naphthol AS-D-chloroacetate esterase as a granulocyte marker and acid alpha-naphthyl acetate esterase as a T-lymphocyte marker were identified on smears of the washed cell suspension . Different enzymes were identified in the cell sediment with electrophoresis . Relatively pure lymphocyte suspensions were obtained within the first 24 h . After this time, however, the percentage of these mononuclear cells declined markedly . The percentage of isolated granulocytes varied slightly; there was a marked predominance of granulocytes (more than 70%) at all intervals investigated during the isolation . Cytochemical analysis of the granulocytes and lymphocytes indicated that the decrease in the percentage of enzyme-positive cells depends in each case on the duration of the storage interval . During the first 24 h, only PGM1 and GOTM could be identified in the lymphocyte suspension with horizontal starch gel electrophoresis . The enzymes PGM1, PGM3, PGI, MDH, GOTM, 6-PGD, ADA could always be identified in the granulocyte suspension; AK, FUCA, MEM could be occasionally identified; and GPT and GLO could never be identified.

Leuk Res, 1983, 7(4), 523 - 37
Non-Hodgkin's lymphoma phenotyping: problems in the use of heterologous and monoclonal antibodies; Barcos M et al.; Cell suspensions or frozen sections of lymph node biopsies from 32 patients with non-Hodgkin's lymphoma (NHL) were studied for sheep erythrocyte (E)-binding under three conditions (Estandard, EAET, Egravity), Fc and C receptors, immunoglobulin (Ig) heavy and light chain class and reactivity with heterologous antisera to T cells (T-LCL), HLA-D (Ia-like) and common acute lymphocytic leukemia (c-ALL) antigens . Selected B and T cell lymphomas were also tested for reactivity with the monoclonal antibodies OKT 3, OKT 4, OKT 6, OKT 8, OKT 11A, Leu-1, Leu-2a, Leu-3a, Leu-4 and Leu-7 . There were 26 B and 6 T lymphomas . Most B lymphomas were mu+ (81%), kappa+ (77%) and 31% were mu+ delta+ . One of the T lymphomas arose in a patient with antecedent follicular small-cleaved (B) cell lymphoma . The most accurate marker for characterizing the immunologic phenotype in NHL was the clonal excess of kappa+ or lambda+ cells . Neither Estandard, EAET, Egravity or T-LCL were consistently reliable as sole reagents in identifying T-cell lymphomas, their individual scores often being lower than those of monoclonal pan-T cell reagents . HLA-D (Ia-like) antigen was noted in 89% of B and 50% of T lymphomas . The corresponding values for c-ALL antigen were 12 and 33%, respectively . The comparative scores in T-lymphomas between OKT 4 and Leu-3a for "helper-inducer" (HE) cells and OKT 8 and Leu-2a for "suppressor-cytotoxic" (SU) cells were not uniformly consistent . Four T lymphomas had a mixed HE/SU cell phenotype, one was HE, and another SU . Anti-T reactivity was detected in the neoplastic follicles of six of seven follicular lymphomas . The percentage of anti-T reactive cells within positive neoplastic follicles was usually small (5-15%) and of the same order as that noted within reactive lymphoid follicles (5-30%) . High numbers (50-100%) of cells from five small lymphocytic B, three diffuse small cleaved cell B and six T cell lymphomas were also positive with one or more anti-T reagents, suggesting the presence of cross-reactive antigens that make phenotyping of lymphomas with monoclonal antibodies problematic . Reactivity with the monoclonal antibody Leu-7 (HNK-1), a putative NK-specific reagent, was seen in one of five B and three of five T lymphomas.

J Membr Biol, 1983, 75(1), 65 - 72
Receptor capping in mouse T-lymphoma cells: a Ca2+ and calmodulin-stimulated ATP-dependent process; Bourguignon LY et al.; The roles that Ca2+, calmodulin, and ATP play in the redistribution of concanavalin A (Con A) binding sites on the surface of mouse T-lymphoma cells were examined . Membranes of cells labeled with fluorescein-conjugated Con A (Fl-Con A) were made permeable ("skinned") to ions and proteins by incubation in a solution containing no added Ca2+, 7 mM EGTA, and ATP . The intracellular ionic and protein concentrations could then be varied, and the degree of Con A receptor capping monitored simultaneously . A graded increase (9.0 to 30%) was found in the number of capped cells with increasing Ca2+ concentration from 10(-6)-10(-4.9) M . Increasing concentrations of trifluoperazine, chlorpromazine, and promethazine (1.5 x 10(-6) to 1.0 x 10(-4) M) in cell suspensions containing 10(-4) M Ca2+ produced graded inhibition of capping in the same order that the drugs bind to calmodulin . Removal of extracellular Ca2+ dissociated (reversed) some of the caps into patches, thus reducing their number (12%) . ATP was required for either capping or cap dissociation to occur . Addition of calmodulin (3.9 x 10(-8)-6.3 x 10(-7) M) to the cell suspension increased the Ca2+ sensitivity . These results provide direct evidence that capping of Con A receptors is a reversible process (i) regulated by intracellular Ca2+ concentrations, (ii) requiring ATP as an energy source, and (iii) susceptible to the influence of calmodulin . These findings are consistent with the hypothesis that the collection of surface receptor patches into cap structures is controlled by the interaction of actomyosin filaments, which in turn is regulated by a Ca2+-calmodulin-activated control system.

J Immunol, 1983 Jan, 130(1), 203 - 8
Immunohistologic analysis of lymphoid infiltrates in primary Sjogren's syndrome using monoclonal antibodies; Adamson TC 3rd et al.; The characterization of lymphocytes infiltrating salivary glands in patients with primary Sjogren's syndrome (1 degree SS) yields insights to disease pathogenesis that are not revealed by studies of the corresponding peripheral blood lymphocytes (PBL) alone . We analyzed salivary gland lymphocytes (SGL) and PBL in 14 patients with untreated 1 degree SS using monoclonal antibodies that detect T cells, T cell subsets, B cells, and antigens associated with lymphocyte activation . A four-step biotin-avidin immunoperoxidase technique was used for salivary gland frozen sections; cell suspensions and PBL were stained cytofluorographically . A predominance of T cells (Leu 1 = L17F12; Leu 4 = OKT3) was found in SGL (greater than 75%) and PBL (76 +/- 9%) with the majority belonging to the Leu 3a (OKT4) subset . A minority of B cells (anti-delta, -kappa, -lambda) was present in both SGL and PBL; however, a subset of B cells defined by monoclonal antibody B532 was present in SGL (5 to 20%) but was absent from PBL . An increased prevalence of activation antigens (Ia; OKT10) was found on SGL T cells (greater than 50% positive) compared to PBL T cells (less than 15% positive) . These studies demonstrate that specific antigenic markers on lymphocytes at the site of inflammation in 1 degree SS differ significantly from those of the corresponding PBL . These differences emphasize that theories of disease pathogenesis of 1 degree SS must include studies on SGL.

Acta Physiol Scand Suppl, 1983, 522, 9 - 18
Intracerebral grafting of neuronal cell suspensions . II . Survival and growth of nigral cell suspensions implanted in different brain sites; Bjorklund A et al.; Dissociated dopamine-rich cell suspensions were prepared from the ventral mesencephalon of rat embryos and injected in one or several sites in striatal and non-striatal regions in the dopaminergically denervated brain of adult rats . While the grafts survived well in all sites, the dopamine fibre outgrowth was markedly different depending on whether the grafts occurred in an area normally innervated by the mesencephalic dopamine neurones (i.e . neostriatum or nc . accumbens) or in areas not normally innervated by these neurones (i.e . parietal cortex, lateral hypothalamus or substantia nigra) . Moreover, in grafts placed at different sites along the trajectory of the nigrostriatal pathway the outgrowing fibres remained confined to the graft, and there was little evidence that the implanted neurones could elongate their axons along the pathway of the nigrostriatal tract to reach the striatum from a distance . Thus, the intracerebral suspension grafts provided efficient reinnervation of a denervated target only when placed in the immediate vicinity of the target area . The results of multiple graft placements indicate that a relatively complete restoration of a lost innervation should be possible to achieve in large areas of the brain, such as the striatal complex, with the suspension grafting technique.

Acta Physiol Scand Suppl, 1983, 522, 67 - 75
Intracerebral grafting of neuronal cell suspensions . VIII . Survival and growth of implants of nigral and septal cell suspensions in intact brains of aged rats; Gage FH et al.; Neuronal cell suspensions prepared from the ventral mesencephalon and the septal-diagonal band area of rat embryos were implanted into the depth of the intact neostriatum or hippocampus of 21-23 month old female rats . Graft survival, assessed 3-4 months after grafting, was comparable to that seen in our previous studies of young adult recipients . Fibre outgrowth into the host brain was evaluated in animals which were subjected to lesions of the intrinsic nigrostriatal or septohippocampal system 6-10 days before killing . Dense dopamine fibre outgrowth was seen within a zone of up to about 1 mm radius around the nigral implants, and dense growth of acetylcholine esterase (AChE) positive fibres occurred up to about 2 mm away from the septal implants . The overall magnitude of fibre outgrowth was less than that generally seen in previously denervated targets in young adult recipients, but it appeared to be as extensive as in young recipients when the grafts are placed in non-denervated targets . The distribution of the AChE-positive fibres from the septal implants in the host hippocampus suggested that the pattern found in the non-denervated target of the aged recipients was more diffuse, and partly different, from normal, and that age-dependent synapse loss in intrinsic connections may influence the patterning of the graft-derived innervation.

Acta Physiol Scand Suppl, 1983, 522, 59 - 66
Intracerebral grafting of neuronal cell suspensions . VII . Recovery of choline acetyltransferase activity and acetylcholine synthesis in the denervated hippocampus reinnervated by septal suspension implants; Bjorklund A et al.; The time-course and magnitude of fibre outgrowth from septal suspension grafts injected into the previously denervated hippocampal formation was monitored by measurements of choline acetyltransferase (ChAT), and the activity of the grafted neurons was assessed by measurements of {14C}acetylcholine (ACh) synthesis from {14C}glucose in vitro . Graft-derived ChAT activity was barely detectable 10 days after grafting, but increased sharply between 10 days and 1 month in the areas of the hippocampus located close to the septal implants . By 6 months ChAT activity was restored to near normal levels in all segments of the previously denervated hippocampus . The overall hippocampal {14C}ACh synthesis was also restored to normal levels in the grafted animals, and estimates of the ACh turnover rate suggested that the transmitter machinery of the newly established "septo-hippocampal" connections operated at a rate similar to that of the intrinsic septohippocampal pathway . The intrahippocampal septal suspension grafts, similar to the intrastriatal nigral grafts, thus seem to be capable of maintaining function at a relatively "physiological" level despite their abnormal positions.

Acta Physiol Scand Suppl, 1983, 522, 49 - 58
Intracerebral grafting of neuronal cell suspensions . VI . Survival and growth of intrahippocampal implants of septal cell suspensions; Bjorklund A et al.; The survival and growth of intrahippocampal septal suspension grafts were investigated by acetylcholine esterase (AChE) histochemistry in animals with lesions of the intrinsic septohippocampal cholinergic pathways . AChE was demonstrable in the grafts after the first postoperative week, and AChE-positive fibres were seen to extend into the host hippocampus by 3 weeks . Rapid fibre outgrowth occurred between 3 weeks and 3 months after grafting, and continued at a slower rate thereafter . By 6 months a fairly complete reinnervation of the initially denervated hippocampus was achieved in most specimens, and this persisted at 14 months, the longest postoperative time analysed . A comparison between the development of the AChE-positive neurones in the suspension grafts with that seen during ontogeny in situ suggested that the grafted neurones lagged behind normal development by at least 1 week . Similar to our previous observations on septal grafts implanted as solid tissue pieces, the pattern of the newly-formed AChE-positive innervation in the host hippocampal formation, established from the septal suspension grafts, was remarkably similar to that of the normal AChE-positive septal innervation . This pattern became established as soon as the graft-derived fibres first grew in, suggesting that the ingrowing axons extended and ramified preferentially into those hippocampal subfields which normally receive an AChE-positive innervation from the septal-diagonal band area.

Acta Physiol Scand Suppl, 1983, 522, 1 - 7
Intracerebral grafting of neuronal cell suspensions . I . Introduction and general methods of preparation; Bjorklund A et al.; The steps involved in the grafting of mesencephalic and septal embryonic tissue in the form of dissociated cell suspensions are described in detail . This includes dissection of the donor embryos, incubation in trypsin, mechanical dissociation, and stereotaxic injection into the brains of adult recipient rats . Some of the technical problems and limitations are discussed.

Leuk Res, 1983, 7(6), 735 - 46
ALL masquerading as AUL; Greaves MF et al.; Of 597 cases of acute leukaemia in adults (greater than 16 years) seen at St . Bartholomew's Hospital, London, between May 1973 and January 1982, 412 were diagnosed as AML, 103 as ALL and 58 as Philadelphia chromosome positive blast crisis of CML (13 presenting as acute leukaemia and 45 having a prior chronic phase) . The remaining 24 cases were considered to be acute undifferentiated leukaemia . Twenty-one of the latter were investigated using a panel of immunological markers at diagnosis and/or retrospectively using frozen cell suspensions . Eighteen out of 21 were shown to have a predominantly 'lymphoid' phenotype which comprised 12 cases of common ALL (two of whom were Ph1 positive), three cases of null-ALL, one case with a probable early thymic phenotype, and two cases with a monoclonal B lymphoblast phenotype . One 'common ALL' and one 'null-ALL' had a significant proportion of pre-B (cytoplasmic mu chain+) cells . One other case reacted with anti-myeloid sera . Leukaemic blasts from two patients were unreactive with all markers tested . No cases of glycophorin positive erythroleukaemia or anti-platelet (glycoprotein I) positive leukaemia were detected . These observations suggest that the overwhelming majority of acute leukaemias have an identifiable affiliation to the lymphoid or myeloid lineages and that patients diagnosed haematologically as 'AUL' might benefit by therapy appropriate for their leukaemic cell type.

Cell Tissue Res, 1983, 231(2), 313 - 23
Characterization of the population of phagocytic cells in thymic cell suspensions . A morphological and cytochemical study; Duijvestijn AM et al.; Rat thymic phagocytic cells were characterized in vitro using various light- and electron-microscopical techniques . Thymic cell suspensions were mechanically prepared and enriched for non-lymphoid cells, which were predominantly phagocytic and of three types . Type I showed acid phosphatase (APh) activity in small granules dispersed throughout the cytoplasm and were mostly Ia antigen-positive, although the Ia membrane label varied in intensity and distribution among individual cells . Only a few cells had endogenous peroxidase activity . The type-I cells could not be clearly distinguished morphologically from type-II or -III cells, and most likely comprise precursors of both these cell types . Type-II were large pale cells with many slender cell processes . These cells had APh activity centrally positioned, were strongly positive for Ia on the cell membrane and were negative for endogenous peroxidase . The cytoplasm frequently contained Birbeck granules, which unequivocally classifies these cells as the in vitro equivalent of the interdigitating cells present in the medullary area of the thymus in situ . Type-III cells were rounded with a smooth or ruffled cell membrane and contained vacuoles and many phagolysosomes . They were strongly positive for APh which was present throughout the cytoplasm . About 50% of these cells were positive for endogenous peroxidase in a pattern resembling resident macrophages . The cells were negative for Ia antigens . Type-III cells mostly likely represent the macrophages found in the cortical area of the thymus.

Cancer Drug Deliv, 1983, 1(1), 43 - 58
Inhibition of liver metastases of M 5076 tumor by liposome-entrapped adriamycin; Mayhew E et al.; The toxicity and therapeutic efficacy of free adriamycin (AM) and AM entrapped in standardized liposomes (AM-MLV) were evaluated in normal mice and in mice bearing M 5076 murine tumor, which metastasizes to the liver after i.v . and s.c . transplants of tumor cell suspensions . Acute and chronic toxicity to AM could be reduced by drug encapsulation in liposomes . The data indicated that at approximately equitoxic doses of free AM (10 mg/kg) and AM-MLV (greater than 20 mg/kg), the increase in survival times of mice transplanted i.v . with tumor cells were 25% and 100%, respectively . Furthermore, only in the AM-MLV-treated mice were long-term survivors observed . In contrast AM-MLV were equally effective as free AM in mice transplanted s.c . with tumor cell suspensions . AM-MLV, however, were more effective than free AM against liver metastases in mice bearing s.c . tumor, indicating differential antitumor activities against the same tumor type growing at different locations in the same animals.

Clin Exp Metastasis, 1983 Jan-Mar, 1(1), 71 - 81
Treatment of artificially-induced pulmonary metastases with fractionated doses of vincristine and/or radiation therapy; Grdina DJ et al.; The cytotoxic effects in vivo of vincristine (VC), radiation, or both modalities in combination on murine fibrosarcoma (FSa) cells grown as pulmonary tumors were determined . Fourteen days following the i.v . injection of viable FSa cells, recipient mice developed between 100 and 150 visible pulmonary nodules . At that time, tumor-bearing animals were exposed to either single or combined modality treatments, as well as single and fractionated dose regimens . Animals were sacrificed 1 hour after the last treatment . Tumor nodules were excised and made into a single cell suspension and separated on the basis of cell size by centrifugal elutriation . Flow microfluorometry (FMF) was used to determine the cell-cycle parameters and the relative synchrony of the separated populations, as well as the percentage contamination by normal diploid cells in each of the tumor cell populations . Known numbers of viable cells from each elutriator fraction were injected into recipient mice to determine their colony-forming efficiency (CFE) . Surviving fractions were determined by comparing the CFEs of treated FSa cells from each of the separated elutriator fractions with those of appropriate untreated controls . Following a single dose of VC (1 mg/kg), populations of cells enriched in late S and G2 + M were the most sensitive . However, following the administration of five doses (0.25 mg/kg each) over a 24 hour period, populations of cells most enriched in G1 cells exhibited the lowest percentage of survivors . A single dose of radiation (1000 rad) was most effective in killing S-phase cells . When administered in five fractions (250 rad per fraction), each separated by a 6 hour interval, no phase-specific sensitivity was observed . The combination of both modalities exhibited a marked schedule dependence, with a single dose (1000 rad) of radiation followed by five doses of VC (0.25 mg/kg) being the most effective treatment protocol observed.

Acta Physiol Scand Suppl, 1983, 522, 39 - 47
Intracerebral grafting of neuronal cell suspensions . V . Behavioural recovery in rats with bilateral 6-OHDA lesions following implantation of nigral cell suspensions; Dunnett SB et al.; Bilateral 6-hydroxydopamine-induced lesions of the ascending forebrain dopamine neurones induce a behavioural syndrome in rats which includes profound aphagia, adipsia, akinesia and bilateral sensorimotor neglect . Such animals will die unless maintained by intragastric feeding . Three experiments are reported in which we have attempted to ameliorate this syndrome with single or multiple placements of nigral cell suspensions into the forebrains of rats with bilateral dopamine depletions . Although the grafts were efficient in reversing the sensorimotor and akinetic impairments, and produced a significant increase in eating, the grafted rats remained hypophagic and adipsic . The results indicate that although many components of the bilateral dopamine denervation syndrome can be reversed by intrastriatal nigral suspension grafts, the severe eating and drinking deficits remain unameliorated.

Acta Physiol Scand Suppl, 1983, 522, 29 - 37
Intracerebral grafting of neuronal cell suspensions . IV . Behavioural recovery in rats with unilateral 6-OHDA lesions following implantation of nigral cell suspensions in different forebrain sites; Dunnett SB et al.; Single and multiple implants of nigral cell suspensions were grafted to the forebrains of rats with unilateral 6-hydroxydopamine-induced dopamine denervations . Control lesions alone induced a marked behavioural asymmetry, as assessed by amphetamine- and apomorphine-induced rotation, sensorimotor tests and side bias in an unbaited T-maze, and the animals were hyperactive to a low dose of apomorphine . Single suspension placements into different denervated striatal regions were capable of reversing the behavioural asymmetries dependent upon the specific placement for each test . Multiple suspension grafts were capable of reversing all behavioural asymmetries, and additionally abolished the supersensitive hyperactivity to apomorphine . By contrast, single suspension grafts placed into the substantia nigra or lateral hypothalamus had no detectable effect on any functional measure . The results indicate that nigral suspension grafts can be at least as effective as solid grafts in reversing the functional deficits induced by dopamine denervation, provided that placements are selected within appropriate dopamine terminal regions of the forebrain (e.g . caudate-putamen or nucleus accumbens).

Scan Electron Microsc, 1983, (Pt 4), 1963 - 72
The application of scanning electron microscopy to cells in culture: selected methodologies; Allen TD; Cell culture is one of the most widely used techniques in modern biology . As a result, morphology of culture systems at the ultrastructural level is also becoming more widespread . Whilst offering an optimum in terms of fixation accessibility, cells in tissue culture can present problems in terms of handling for scanning electron microscopy, particularly when they are grown on, or in semi-solid media, or on gels e.g . collagen . The examination of cultured cell suspensions or mixed cell populations may also require careful handling to produce meaningful results in terms of surface morphology and cellular interaction . Cells specifically cultured on carbon films can also be used for information of internal cell structure as well as surface topography, often simultaneously, if a suitable imaging system is available . This communication reviews a wide spectrum of experimental manipulations of cultured material to provide an introduction to the gathering of morphological information from material in tissue culture.

Jpn J Physiol, 1983, 33(4), 635 - 50
Rate of CO2 diffusion in the human red blood cell measured with pH-sensitive fluorescence; Niizeki K et al.; The diffusion rate of CO2, into and out of the red cell, was measured by using a stopped flow method with pH-sensitive fluorescence of 4-methylumbelliferone . A red cell suspension of 15% hematocrit with the PCO2 of 8 Torr (or 65 Torr) was mixed with the same amount of saline solution having 65 Torr (or 8 Torr) . Carbonic anhydrase was added to the external solution at a concentration of 20 mg/100 ml in order to accelerate the hydration and dehydration reactions, so that the PCO2 change in the fluid could be observed instantaneously through pH . In the inward diffusion PCO2 showed a large change, suggesting a lack of HCO3- shift across the red cell membrane . In the outward diffusion, however, the PCO2 change was smaller, suggesting that H+ ions produced in the external solution by CO2 hydration were rapidly buffered by the red cell . The half-times of the inward and outward diffusions were, on an average, 0.08 and 0.13 sec, respectively . The results of the simulation revealed that the above difference in half-times was attributed to the difference in slope between the two dissociation curves with and without the HCO3- shift . The diffusion rate was almost constant and remained independent of the direction of CO2 flux . That is, at a low pH range the permeation of H+ ions across the red cell membrane was much faster than the diffusion rate of CO2.

Vox Sang, 1983, 45(3), 217 - 23
Red cell suspensions in SAGM medium . Further experience of in vivo survival of red cells, clinical usefulness and plasma-saving effects; Hogman CF et al.; Red cells depleted of buffy coat and more than 90% of the plasma were suspended and stored in a medium composed of sodium chloride, adenine, glucose and mannitol (SAGM) . The 24-hour posttransfusion survival of 51Cr-labeled red cells was 83.5 +/- 5.3% (n = 4) after storage for 35 days and 77.4 +/- 4.7% (n = 6) after 42 days . No abnormal in vivo hemolysis occurred as judged from posttransfusion haptoglobin consumption studies . No abnormal body temperature elevation was found at continuous pertransfusion recordings . The frequency of febrile or urticarial transfusion reactions was 0.19% as compared to 0.68% during a whole-blood transfusion period . Since a mean of 280 ml of plasma can be collected from each blood unit the plasma-saving effects of the system are considerable . Favorable large-scale clinical experience is reported.

Immunol Rev, 1983, 73, 35 - 51
Natural cytotoxicity: early killing of allogeneic lymphocytes in rats; Heslop BF et al.; In many strain combinations among inbred rats, intravenously injected 51Cr-labelled lymphocytes are destroyed in substantial numbers by unsensitized allogeneic hosts . Destruction of cells (referred to as natural cytotoxicity (NC)) occurs within a few hours of injection, and is characterised by a decreased accumulation of radioactivity in the lymph nodes and increased renal excretion of label by allogeneic hosts, as compared with the distribution of label in syngeneic recipients of the same cell suspension . An intact spleen is necessary for killing . The level of NC expressed is consistent for a given donor-host combination . Using arbitrary criteria to compare the levels of NC expressed by different donor-host combinations among inbred rats, 13 of 95 strain combinations have been shown to express high NC, 63 intermediate NC and 19 low NC . The level of NC expressed cannot be correlated with the extent to which donor and host differ in respect of known MHC genes . Segregation analysis has shown high NC to be controlled by at least 2 independently segregating genes, one of which is MHC-linked . It is possible to weaken or abrogate NC by the neonatal injection of bone-marrow cells from the donor strain, and to reverse this reduced reactivity by the injection of host strain lymphocytes . The substitution of either the donor (P1) or the host (P2) by the P1 X P2)F1 hybrid reduces or eliminates NC in strain combinations normally expressing high NC . It is currently uncertain whether NC can be augmented . In the single strain combination in which maturation has been studied, NC becomes evident during the 4th week of life and attains adult levels during the 6th-7th weeks . NC is at least partially radio-sensitive . Two groups of reactivities appear to be related to NC: (i) those which have been identified in the context of aberrant lymphocyte homing, and for which allogeneic lymphocytes are the targets; (ii) the group of natural resistance systems which includes NK cells, and whose reactivity is directed against a variety of other target cells.

Ophthalmic Res, 1983, 15(2), 61 - 7
Aqueous humor and serum IgE antibody in experimental ocular Onchocerca infection of guinea pigs; Donnelly JJ et al.; Infection of inbred Strain 2 guinea pigs by subcutaneous or intradermal injection of fresh or cryopreserved living Onchocerca lienalis microfilariae, followed by a challenge intracorneal infection of microfilariae, resulted in serum and aqueous IgE antibody and in significant corneal inflammation . Systemic or intraocular infections given separately were not sufficient to elicit IgE antibody or ocular inflammation . When intravenous transfer of pooled spleen cell suspensions from systemically infected donors to normal syngeneic recipients was substituted for the course of systemic infections, a subsequent intracorneal challenge of cell transfer recipients with microfilariae produced serum and aqueous IgE antibody . Administration of diethylcarbamazine citrate to infected animals following the intracorneal challenge resulted in increased serum IgE antibody and in increased corneal inflammation.

Dev Neurosci, 1983-84, 6(3), 137 - 51
Intracerebral grafting of embryonic neural cells into the adult host brain: an overview of the cell suspension method and its application; Gage FH et al.; An overview is presented of general principles for intracerebral grafting of embryonic brain tissue to the adult mammalian brain . Special reference is made to the development and use of the dissociated neuronal cell suspension method . Examples are drawn primarily from experiments where embryonic ventral mesencephalon is transplanted to adult striatum and embryonic septal-diagonal band area is transplanted to the hippocampal formation . Results related to the cell viability in vitro and in vivo and to axonal outgrowth are the main focuses of this overview.

Arch Immunol Ther Exp (Warsz), 1983, 31(4), 449 - 57
Antimonocyte serum . Preparation and characterization; Goscicka T et al.; Cell suspensions enriched with monocytes were obtained from the blood of Vietnam normal pig and rabbits injected with avirulent Welshimer strain of L . monocytogenes by velocity sedimentation on BSA gradient and separation on Ficoll-Triosil gradient . They were used for immunization of rabbits and a sheep, respectively . The antisera specific for monocytes (RAMS and SAMS) were obtained by scrupulous absorption with lymphocytes and granulocytes.

Neoplasma, 1983, 30(6), 691 - 700
Electrophoretic mobility profiles of mouse leukemias . II . Electrophoresis of thymic lymphoma, myeloid leukemia and reticulum sarcoma cells; Bubenikova D et al.; Electrophoretic mobility of cells from 21 primary mouse leukemias was investigated and compared with that of various normal mouse reference cells . Six thymic lymphomas, nine reticulum cell sarcomas, five myeloid leukemias and one stem-cell leukemia were examined . The mean AEM of cells from the group of primary thymic lymphomas (1.13 microns s-1 V-1 cm) was similar to that of reticulum cell sarcomas (1.15 microns s-1 V-1 cm) and significantly lower than that of the myeloid leukemias (1.20 microns s-1 V-1 cm) . Since the examined cell suspensions were prepared from leukemic lymph nodes, the analysis was performed to investigate the possible admixture of normal LNC in the cell populations . Electrophoretograms of normal and leukemic lymph nodes were compared and the AEM of cells from leukemic lymph nodes was considered separately for two cell populations, one corresponding in size to normal LNC (6-9 microns in diameter) and the other to leukemic blast cells (10-16 microns in diameter) . In the 6-9 microns cell subpopulations from leukemic lymph nodes, electrophoretically slow cells corresponding to B LNC were almost totally depleted (1%); also electrophoretically fast cells with the mean AEM of T LNC were fewer (45-57%) than in normal LNC populations . The leukemic cells 10-16 microns in diameter displayed mean AEM of 1.09 (thymic lymphomas), 1.12 (reticulum cell sarcomas) and 1.21 (myeloid leukemia) micron s-1 V-1 cm . The mean AEM of blast cells from thymic lymphomas and reticulum cell sarcomas was not significantly different; it was similar to that of blast cells prepared from normal mouse thymus (1.09 microns s-1 V-1 cm) on a density gradient . In contrast, the 10-16 microns cell populations from lymph nodes of mice with myeloid leukemias were significantly faster than blast cells from thymic lymphomas and reticulum cell sarcomas.

Ann Med Interne (Paris), 1983, 134(5), 395 - 410
{Blood hyperviscosity syndromes . Classification and physiopathological understanding . Therapeutic deductions}; Larcan A et al.; Blood has a number of rheological properties which partially determine flow, especially at capillary level, and its capacity to deliver oxygen . It is non-Newtonian, pseudoplastic, thixotropic and viscoelastic . Viscosity can be studied with different types of viscosimeters (coaxial cylinder or capillary viscosimeters) . It can be defined by the ratio of stress of deformation to rate of deformation . Viscosity depends on macrorheological parameters: hematocrit, serum proteins, especially fibrinogen and globulins, and also on microrheological parameters: degree of aggregation and red blood cell deformability . Viscosity rises when the temperature falls and decreases with the radius of the tube through which the blood flows (Fahraeus-Linqvist effects) . Blood viscosity is studied clinically at different temperatures, and, above all, at different rates of deformation by carefully recording the hematocrit . Plasma viscosity, fibrinogen, albumia and immunoglobulin levels, the viscosity of blood cell suspensions in normal saline must also be taken into consideration . Special investigations (rheoscopy, filtrability) provide information about red cell aggregation and deformability . Hyperviscosity syndromes are observed with: --raised hematocrit (polycythemia and pseudopolycythemia), --conditions with raised serum proteins or changes in their composition (especially hyperfibrinogenemia, raised immunoglobulins, low albumin levels); inflammatory syndromes, dysglobulinemias (Fahey's syndrome of plasma hyperviscosity), --low temperature (hypothermia), --increased red cell aggregability (shock, fat embolism), --reduced red cell deformability due to various congenital and acquired conditions (sickle cell anemia, renal failure, hyperlipoproteinemia, thrombosis, diabetes) . Conversely, hypoviscosity may occur with a low hematocrit, hypoproteinemia, hypofibrinogenemia, and hyperthermia . Increased viscosity results in a slowing of blood flow, stagnation of its constituents and in ischemia . Therapeutic interventions may be considered on the different components of the hyperviscosity syndrome: hemodilation, plasmapheresis, dispersion of aggregants, agents acting on red cell deformability.

Adv Exp Med Biol, 1983, 159, 419 - 34
Cellular oxygen utilization and radiation response of V-79 spheroids; Durand RE; Chinese hamster V-79-171 cells, when placed in stirred suspension cultures, spontaneously grow as spherical multicell aggregates (spheroids) which eventually contain many thousand tightly-packed cells . These spheroids thus provide a valuable model for the study of oxygen transport, since oxygen concentration at the spheroid periphery can be fairly precisely controlled in the stirred suspension, and the consequences of oxygen diffusion inward through the tissue-like mass can be determined experimentally and compared with theoretical calculations . As in human tumors, appearance of central necrosis can be correlated with severe hypoxia . Additionally, since severely hypoxic cells are considerably more resistant to radiation than are oxygenated cells, determination of the fraction of hypoxic cells by their radioresistance provides a noninvasive technique for monitoring respiratory processes . Using this approach, we have found an excellent correlation between the rate of cellular oxygen utilization by a single cell suspension and the net radio-sensitivity of multicell spheroids when treated with modulating agents, varying from direct control of metabolic activity by changing the ambient temperature, to less direct effects on cellular oxygen consumption induced by exposure of cells to nitroheterocyclic radiosensitizers, chemotherapeutic agents, anesthetics, antibiotics, and even cell growth factors including insulin . Our data from these studies indicate that, at least in the simplistic spheroid system where oxygen delivery to the internal cells is a direct function of the respiratory activity of the external cells, control of cellular oxygen utilization is a convenient and effective method of controlling the net radiosensitivity.

Adv Exp Med Biol, 1983, 159, 399 - 417
A method to measure the radio and chemosensitivity of human spheroids; Carlsson J et al.; A method based on the spontaneous outgrowth of cells from spheroids was tested . Different outgrowth patterns were seen depending on the types of spheroids and on the radiation or drug doses . The method allowed dose-effect relations to be determined . Spheroid survival was defined as when the outgrowing monolayers contained at least thousand cells within five weeks . The method was used as an alternative to cloning of isolated single cells . The glioma and osteosarcoma spheroids could not be disintegrated to single cell suspensions since they resisted enzymatic and mechanical treatments for cell separation . Detection of differences in radio and chemosensitivity between different types of spheroids of human origin might be valuable for the understanding of the large variations in therapeutical response often seen between different types of tumors.

Microsc Acta Suppl, 1983, 6, 21 - 7
{Determination of a significant cell count for positive diagnosis in gynecologic suspension preparations and possible consequences for the analysis of automated preparations}; Schwarz G et al.; Monolayer preparations used in automated cytology are generated from gynecologic cell suspensions . The suspension-derived preparations (s.p.) show some peculiarities compared with conventional cytologic smears . One important feature of the s.p . is a distribution of cells mainly by chance . Therefore it seems to be possible to determine a cell sample size in minimum needed for cytologic diagnosis . In a blind test the needed cell sample size was determined by counting the cells in the microscopic fields of vision (ocular 12,5 x, objective 10 x, phi of the field of vision 2 mm) step by step until it was possible to classify the specimen (diagnosis 1) . To be on the safe side (personal safety requirement) some further cells were assessed (diagnosis 2) . We used s.p . from 50 non-suspicious women and 50 women with invasive squamous cell carcinoma of the uterine cervix and its precursors . To recognize strong positive cases (severe dysplasia to invasive carcinoma) only about 600 cells are needed (mean: 150/147 cells) . The most cells (about 1200 cells) are required in negative cases and cases of mild to moderate dysplasia (mean: 293/236 cells) . The highest personal safety requirement was found in cases of mild to moderate dysplasia (311% compared with a mean sample size of 236 or 100%) . The results support such approaches in automated cytology which analyze only some hundreds or about 1000 cells with high resolution.

Arch Dermatol Res, 1983, 275(2), 130 - 3
Isolation of a viable eccrine sweat gland by dispase; Okada N et al.; Dispase was used to obtain viable eccrine sweat glands from human skin in an intact shape . The full thickness of human skin was soaked in a solution of dispase in Eagle's minimum essential medium at a concentration of 500 units/ml and kept in a refrigerator at 4 degrees C for 24 h . The epidermal sheet with its appendages could then be easily separated from the dermis by lifting the epidermis with fine forceps . Electron-microscopic observation revealed that the eccrine sweat gland was completely separated froM the dermis at the basement membrane zone . The isolated epidermal sheet was scarcely dissociated by mechanical agitation in the presence of Ca2+ and Mg2+ ions . The eccrine sweat gland was cut away from the epidermis by using microscissors under a stereomicroscope . A cell suspension of the isolated eccrine sweat glands was obtained after trypsinization . The cells remained more than 90% viable up to 48 h in the culture medium . The obtained viable eccrine sweat glands will be useful for the study of the biology of sweating.

Pathol Biol (Paris), 1983 Jan, 31(1), 49 - 51
{Test of human basophil degranulation . The interpretation and analysis of variability factors in the absence of antigen}; Mordelet-Dambrine M et al.; The human basophil degranulation test (TDBH) is performed using a concentrated cell suspension enriched for polymorphonuclear basophils . This preparation is placed in wells on a glass slide with an allergen solution of decreasing concentration . After incubation the cells are fixed in the wells and stained with toluidine blue . Preparations containing fewer metachromatic basophils than in control preparation of the same cell suspension without antigen contain degranulated basophils and in this way a degranulation index can be calculated . By considering the variability in the number of basophils between duplicate control wells from the same cell suspension, it can be shown that the test can be considered to be positive only if the number of degranulated basophils equals or exceeds 50% of the total number of basophils . This variance can be attributed to the subjectivity of the reader, and to differences in the differential cell counts from well to well . On the other hand, the presence of atopic allergy had no effect on the basophil counts.

Avian Dis, 1983 Jan-Mar, 27(1), 21 - 35
Application of enzyme-linked immunosorbent assay for detecting antibody to Mycoplasma gallisepticum infections in poultry; Ansari AA et al.; An enzyme-linked immunosorbent assay (MYCO-ELISA) was developed to detect antibodies to Mycoplasma gallisepticum (MG) in chicken sera . The assay was standardized in terms of optimum antigen concentration, serum dilution, conjugate dilution and incubation temperature, and time . The MYCO-ELISA antigen was prepared from MG whole bacterial cell or its disrupted cell suspension . Both preparations showed strong affinity for binding or adsorbing to the surface of polystyrene wells of the microtiter plate . The MYCO-ELISA was more sensitive than the hemagglutination-inhibition test . However, cross-reactions were observed with sera from M . synoviae (MS)-infected birds.

J Immunol Methods, 1983, 56(1), 117 - 24
Comparison of the sensitivity of the protein A plaque assay and the immunofluorescence assay for detection of immunoglobulin producing cells; Van Oudenaren A et al.; This paper reports a comparative study on the sensitivity of the protein A plaque assay and the cytoplasmic immunofluorescence assay for detection of immunoglobulin (Ig) producing cells in cell suspensions of murine lymphoid organs . The protein A plaque assay was found to detect as many or several times more Ig producing cells than the immunofluorescence assay, depending on the age and antigenic load of the mice, and upon the Ig class and organ studied.

Diagn Immunol, 1983, 1(3), 142 - 9
The application of monoclonal antibodies to the characterization and diagnosis of lymphoid neoplasms: a review of recent studies; Knowles DM 2nd et al.; Twenty-three T-cell neoplasms were divided, according to their reactivity with the OKT monoclonal antibodies as follows: Fourteen neoplasms of diverse histopathology expressed the OKT3+ OKT4+ phenotype, commonly associated with the helper T-cell subset; seven histologically similar lymphoblastic neoplasms expressed diverse phenotypes consistent with various stages of intrathymic differentiation and two neoplasms expressed the uncommon OKT3+ OKT10+ phenotype . Thus, T-cell neoplasms are divisible into phenotypes that correspond to normal stages of T-cell differentiation and functionally distinct T-cell subsets . Benign and malignant lymphoid cells were investigated in cell suspension and in cryostat tissue sections for their reactivity with OKB1, OKB2, OKB4, and OKB7, monoclonal antibodies known to detect distinctive B lymphocyte antigens . Fetal liver pre-B cells, cases of pre-B acute lymphoblastic leukemia, and common-type acute lymphoblastic leukemia were OKB2+ but unreactive with the other OKB antibodies . All mature lymphoid tissue B cells and 47/47 surface immunoglobulin (SIg)-positive B-cell neoplasms were OKB2+ . Interfollicular, follicular center, and many, but not all, mantle zone B cells were OKB1+ OKB7+ . Follicular center B cells were OKB4+ but mantle zone and interfollicular B cells were OKB4- . 45/47 SIg+ B-cell neoplasms were OKB1+ OKB4+ . 45/46 SIg+ B-cell neoplasms were OKB7+ . Benign and myeloma plasma cells were OKB- . T-cell neoplasms were OKB2- OKB4- but were occassionally OKB1+ and OKB7+ . The OKB antibodies appear to detect distinctive antigens that may be expressed at different stages of B-cell differentiation.

Acta Physiol Scand Suppl, 1983, 522, 19 - 28
Intracerebral grafting of neuronal cell suspensions . III . Activity of intrastriatal nigral suspension implants as assessed by measurements of dopamine synthesis and metabolism; Schmidt RH et al.; The activity of intrastriatal grafts of nigral cell suspensions has been monitored biochemically, using radioenzymatic assays of dopamine, its major acidic metabolite, DOPAC, and DOPA accumulation after DOPA-decarboxylase inhibition . Implants of 4-9 microliter of nigral cell suspension restored striatal DA levels by an average of 13-18%, with the highest individual values reaching about 50% of control . DOPAC was restored from about 5% in the lesioned controls to about 20% of normal in the grafted animals . The DOPAC: DA ratios and the DOPA accumulation measures indicated that the grafted DA neurons were spontaneously active and that the transmitter turnover rate was on the average some 50-100% higher than the intact intrinsic nigrostriatal DA neurones . These results thus provide evidence that the intrastriatal nigral suspension grafts are capable of restoring dopaminergic neurotransmission in the previously denervated striatum.

Breast Cancer Res Treat, 1983, 3(1), 15 - 22
A comparison of methods for the production of monodispersed cell suspensions from human primary breast carcinomas; Besch GJ et al.; Production of monodispersed cell suspensions from primary human breast tumors is difficult due to the predominant stromal composition of most breast tumors . Our studies were designed to optimize dispersion of breast tumors of known stromal content and histopathology . In a first series of experiments three enzymatic protocols were compared to disperse minced tissue: (A) treatment with collagenase (2 mg/ml) in the presence of 5% serum for 24 hours; (B) treatment with collagenase (6 mg/ml) and DNase (0.002%) in 10% serum for 3 hours; (C) treatment with collagenase (2 mg/ml) for 3 hours followed by pronase (0.075%) for 1 hour . Protocol A produced better cell yields than B or C for all tumors tested . The monodispersed cells were suspended in a 0.3% semi-solid agar with alpha modified Eagles medium (alpha MEM), 10% serum, and selected hormones, then layered over similarly enriched 0.5% semi-solid agar . The cells prepared by protocol A had a higher plating efficiency and larger average colony size than B or C . In a second series of experiments, protocol A was repeated and compared to two additional procedures: (D) treatment with collagenase (2 mg/ml) and hyaluronidase (1 mg/ml) in the presence of 5% serum for 24 hours; and (E) mechanical disaggregation . Protocol D exhibited a small but significant negative difference from A, while E was the least efficient in producing viable monodispersed cells from the tumors . All enzymatically monodispersed cells produced clonal growth in our agar system . However, mechanically dispersed cells gave growth in only 4 of 7 tumors . Protocol A, in addition to yielding the highest number of viable cells per gram of tissue, gave the highest plating efficiency of all protocols tested.

Cell Tissue Res, 1983, 231(2), 457 - 61
Scanning electron microscopy of pure Kurloff cell suspensions . Observation of an extrusion process; Landemore G et al.; Scanning electron microscopy has been used to examine suspensions of Kurloff cells, which are present only in guinea pig . The cells are round with a surface characterized by uniformly distributed short stubby microvilli and some ridges . Some cells show one or more pores surrounded by a smooth margin, others a smooth globular expansion about 10 microns in diameter which may correspond to a Kurloff body undergoing exocytosis . The Kurloff body is released into the medium where it can exist freely . The surface of the free body and of the expansion display the same smooth appearance . Although the nature of the Kurloff cell remains unknown, this study provides further information concerning its morphology and describes an extrusion process not observed previously.

Avian Dis, 1983 Jan-Mar, 27(1), 271 - 82
Studies of reticuloendotheliosis virus-induced lymphomagenesis in chickens; Fadly AM et al.; Seventy-three percent of chickens inoculated with the chick syncytial strain of reticuloendotheliosis virus (REV) at hatching developed lymphomas by 39 weeks of age . Neonatal treatment with cyclophosphamide or surgical bursectomy at 2, 4, 8, or 12 weeks of age significantly (P less than 0.01) reduced lymphoma development . In a further experiment, surgical bursectomy of REV-infected chickens followed by intravenous inoculation of the chickens with a single cell suspension of their own bursa cells at 2, 4, 9, or 13 weeks of age resulted in lymphoid tumors in chickens treated at 9 or 13 weeks but not in chickens treated at 2 or 4 weeks of age . Furthermore, this treatment did not shorten the incubation period for lymphoma development . These findings argue very strongly that transforming target cells are primarily in the bursa of Fabricius . The data also suggest that a minimum residency of 4 weeks in the bursa is required for infected bursa cells to become transformed . Therefore, lymphomagenesis induced by REV in chickens appears similar to that induced by the avian leukosis virus group.

Gen Comp Endocrinol, 1983 Jan, 49(1), 81 - 9
Preparation of enriched populations of corticotrophs from goldfish rostral pars distalis; Portanova R et al.; A mammalian isolated adrenal cell system was validated as a bioassay for goldfish ACTH; the log dose-response curve for the goldfish hormone is parallel to that for synthetic mammalian ACTH1-24, and the two ACTHs induce the same maximum rate of corticosterone production . Using this assay, it was observed that (1) there is a marked and consistent biphasic change in pituitary ACTH content as related to the length of time the fish are held in laboratory aquaria, and (2) the absolute and relative (to tissue wt) ACTH content of the rostral pars distalis is considerably greater than that of the proximal portion of the gland . Using a trypsin technique, isolated pituitary cells were prepared from both (rostral and proximal) portions of the gland; bioassay data indicate that cell suspensions prepared from the rostral pars distalis are enriched with respect to corticotrophs . The implications of these findings with regard to formulating an advantageous in vitro system for studying ACTH secretion are discussed.

Am J Physiol, 1983 Jan, 244(1), H109 - 14
Maximum fluid concentrations of materials released from platelets at a surface; Adams GA et al.; We examine the estimation of local concentrations of materials that are released from the dense and alpha-granules of platelets during accumulation of platelets upon collagen-coated glass . Platelet/red blood cell suspensions were perfused through a 1.3-mm-ID tube . Empirical data were used in a calculation procedure, based on diffusion and convection, designed to yield an upper bound on the interfacial fluid concentration (IFC) for each substance considered . The necessary empirical data are the rate of platelet accumulation and the maximum amount of material in the platelet capable of secretion . It was found that the IFC is dependent on the shear rate at the surface (G) and is proportional to G0.27 . This means that an eightfold increase in flow rate would increase the IFCs approximately twofold . Serotonin, pyrophosphate, adenosine 5'-monophosphate (AMP), and adenosine 5'-triphosphate (ATP) were found not to be present in sufficient quantities to produce IFCs that could influence platelet aggregation if used alone at the IFC . A second set of materials, fibrinogen, fibronectin von Willebrand factor, and calcium, had IFCs less than their concentrations normally found in plasma . A third category, containing adenosine 5'-diphosphate (ADP) alone, had an IFC close to those known to affect platelet aggregation . The role of metabolites of arachidonic acid, which may promote or inhibit platelet aggregation, awaits further description.

Acta Biol Hung, 1983, 34(4), 407 - 14
Modification of the cell cycle of limb bud mesenchyme during in vitro cartilage differentiation; Hadhazy C et al.; A consistent chondrogenesis takes place in micro high-density cultures derived from limb mesenchymal cells of chick embryos of stages 23-24 . Flow-cytometric measurements of DNA content showed that cells in the phase of G1 or G0 made up 51% of the dispersed cell suspensions . The proportion of these cells increased to 71% by the onset of cartilage differentiation in day-2 cultures . This ratio was 84% when the voluminous matrix formation began on the 4th day of culturing . Thereafter, it increased to 90% by the 6th day, and to 93% by the 14th day . The results suggest that cartilage differentiates from G0 mesenchymal cells of the limb . In our measurements, however, the G0 phase includes all non-proliferative cell population which have identical DNA content with G1 cells . Therefore, the G0 phase contains also an increasing number of chondroblasts and chondrocytes as the chondrogenesis proceeds.

J Neurooncol, 1983, 1(2), 87 - 94
Depressed T lymphocyte function in brain tumor patients: monocytes as suppressor cells; Wood GW et al.; Depressed cellular immune function has been demonstrated in patients with a variety of lymphoreticular and nonlymphoreticular neoplasms, including patients harboring brain tumors . In the present study, peripheral T lymphocytes from more than fifty percent of patients with central nervous system tumors, both primary and metastatic, exhibited depressed incorporation of 3H-thymidine in response to phytohemagglutinin (PHA) when tests were performed in the absence of autologous serum . Increased numbers of monocytes were present within mononuclear cell suspensions from brain tumor patients, and most of the cell populations containing elevated monocytes also exhibited depressed responses to PHA . A role for monocytes as suppressor cells was suggested by the finding that partial reconstitution of T cell function could frequently be effected by adherent cell depletion . However, total reversal of the defect was rare and there was no relationship between monocytes and T cell dysfunction in some patients . The results suggest that while monocytes may be involved in the immune depression seen in some patients with a brain tumor, the complete explanation is as yet unknown.

J Bacteriol, 1983 Jan, 153(1), 310 - 5
Carbonyl cyanide-m-chlorophenyl hydrazone-resistant Escherichia coli mutant that exhibits a temperature-sensitive unc phenotype; Ito M et al.; Two spontaneous Escherichia coli mutant strains which are resistant to an oxidative phosphorylation uncoupler, carbonyl cyanide-m-chlorophenyl hydrazone, were isolated . Strain CM22 (ccr-2) was resistant to another uncoupler, pentachlorophenol, and to the inhibitors of proton-translocating ATPase, namely tributyltin and sodium azide . Carbonyl cyanide-m-chlorophenyl hydrazone or pentachlorophenol administered to cell suspensions of strain CM22 did not cause a pH change induced by H+ influx, and a similar result was obtained with everted particles . The respiratory rate of strain CM22 with succinate was twice that of wild-type strain KH434 . When carbonyl cyanide-m-chlorophenyl hydrazone was administered, a stimulation of O2 uptake was observed in wild-type strain KH434 but not in the mutant strain CM22 . Strain CM22 did not grow on succinate at 42 degrees C . Isolation of a true revertant at a frequency of 10(-8) demonstrated that the pleiotropic phenotype was induced by a single mutation . P1 transduction indicated that the mutant allele, ccr-2, was cotransduced with the ilv genes at a frequency of about 55%.

Cancer Invest, 1983, 1(1), 5 - 13
Suppression of the immune response in tumor-bearing mice . III . Induction of functionally suppressive antigen-driven macrophages; Bluestone JA et al.; A functionally suppressive antigen-driven subpopulation of macrophages has been demonstrated in mice-bearing tumors produced by MSV-transformed BALB/3T3 cells . The suppressor macrophages depressed markedly both allogeneic and syngeneic mixed leukocyte culture responses and functioned even when assayed at low concentrations, suggesting that high concentrations of normal macrophages were not responsible for the suppression . Studies also showed that suppressor macrophages acted to suppress the proliferation of hyperreactive Lyt 1+,2- cells found in the splenic cell suspensions of tumor-bearing mice after the removal of the adherent suppressor cells . Finally, the suppressive activity of macrophages induced in tumorous mice could be augmented by KiMSV or MMSV but not by MuLV antigen . These results suggest that suppression might be a direct response to transformation-associated determinants expressed on the tumor cells as well as on the MSV virions or the result of cellular interactions between suppressor macrophages and other antigen-specific spleen cell populations.

Acta Derm Venereol, 1983, 63(4), 290 - 5
Induction of leukocyte adherence at the basement membrane zone with subsequent activation of their metabolic pathway by pemphigoid antibodies and complement; Iwatsuki K et al.; We observed in our in vitro studies that human leukocytes were attached by their complement receptors to the basement membrane zone (BMZ) of the skin specimens pretreated with pemphigoid antibodies and complement . The resultant histologic feature was very similar to that of an early stage of the inflammatory reactions observable in the lesions of bullous pemphigoid . Eosinophils were also attracted to the BMZ in proportion to their number in cell suspensions, but the adhesion of eosinophils occurred neither selectively nor excessively . The nitroblue tetrazolium (NBT) test, which was carried out after the leukocyte attachment test, demonstrated that only those leukocytes bound specifically to the BMZ incorporated NBT dye complexes vigorously to form formazan crystals in their cytoplasms . The present studies showed that complement-fixing anti-BMZ antibodies induce enhancement of leukocyte activities as well as immune adherence to the BMZ, indicating that the enhanced leukocyte function plays a role in the separation between epidermis and dermis in bullous pemphigoid.

Teratog Carcinog Mutagen, 1983, 3(4), 321 - 34
New approaches to mutagenicity studies in animals for carcinogenic and mutagenic agents . II . Clastogenic effects determined in transplacentally treated mouse embryos; Adler ID; Cell suspensions from whole embryos were obtained on the 12th day of gestation from treated pregnant female mice . For the present experiments five model mutagens-BaP, bleomycin, mitomycin C, procarbazine, and TEM--were chosen for their different modes of action in inducing chromosomal aberrations . Time-response and dose-response were studied for chromosomal aberrations induced by transplacental treatment of mouse embryos . All five known mutagens gave a positive response in the present system . The maximum time of response varied from compound to compound and was found as early as 3h after treatment with the G2-clastogen bleomycin or as late as 18 h after treatment with the bifunctional alkylating agent TEM . Chromatid breaks and exchanges increased with dose in all instances . The correlations between clastogenic, carcinogenic, and teratogenic effects are discussed . It is concluded that the transplacental cytogenetic test may be applicable to identify chemicals that exhibit all three properties.

Int Arch Allergy Appl Immunol, 1983, 72(3), 273 - 8
Granular and globular acid alpha-naphthyl-acetate-esterase staining pattern of resting and mitogen-activated murine lymphocytes; Treves S et al.; Available methods for the identification of T-lymphocytes in tissue sections and in single cell suspensions are often tedious and require a lot of manipulation . Here we confirm previous findings that nonspecific alpha-naphthyl-acetate esterase (ANAE) is a reliable method for the identification of resting thymus-derived lymphocytes but not for concanavalin-A-stimulated lymphocytes, as a small proportion of lipopolysaccharide-stimulated lymphoblasts exhibit ANAE activity as well . We have also shown that there is a relationship between the globular staining pattern and the degree of cell activation . Finally, we noted that prolonged incubation of lymphocytes at 37 degrees C reduces the number of ANAE-positive cells.

Folia Haematol Int Mag Klin Morphol Blutforsch, 1983, 110(1), 36 - 41
The effects of incubation of marrow in tissue fragments in vitro on the growth of adherent cells from this marrow; Ptasznik A et al.; Femoral marrow was either cultured as a single cell suspension immediately following collection from the donor mouse or following 4 day incubation in vitro of the whole marrow shaft . Several parameters of growth of adherent, i.e . composed of fibroblastoid cells and macrophages, colonies were determined following 14 day culture in Dulbecco medium at 37 degrees C . These included: number and diameter of macroscopic colonies, number of macrophages per fibroblastoid cell inside the colonies and per eyefield in intercolony spaces, number of cells in supernatant from the culture . The 4 day incubation of marrow fragments in vitro (Dulbecco medium, 37 degrees C) doubled the number of adherent colonies grown from this marrow and, moreover, the colonies formed were increased in size . Other parameters of cell growth in these cultures were unchanged . These data suggest that under conditions of in vitro incubation of marrow shaft (close cell-to-cell contact) marrow fibroblastoid colony forming units (MF-CFU) are stimulated to self-renewal.

Acta Derm Venereol, 1983, 63(2), 115 - 21
Pemphigus serum-induced loss of microvilli from human epidermal cells; Hietanen J et al.; Scanning electron microscopic examination of human epidermal cells prepared from suction blister roofs by treatment with trypsin and dithioerythritol and incubated (37 degrees C, 18 hours) with pemphigus sera revealed a smooth or wrinkled surface texture in 83 to 96% and surface microvilli in only 17 to 4% . The disappearance of microvilli was not affected by the presence of complement . In the control specimen 54% of the cells retained their microvilli . The difference between cells incubated with pemphigus and those incubated with control sera was highly significant (p less than 0.001) . Epidermal cell suspensions were also incubated with pemphigus vulgaris sera plus a protease inhibitor (ovomucoid) . The loss of microvilli from human epidermal cells induced by pemphigus vulgaris serum was highly significantly inhibited by ovomucoid (p less than 0.001) . The results suggest that the loss of microvilli induced by pemphigus sera, irrespective of the presence of complement, may be an important factor in acantholysis of pemphigus . Proteolytic enzymes may be responsible for the loss of microvilli and smoothing of the cell surface of human epidermal cells in vitro.

Scand J Immunol, 1983 Jan, 17(1), 13 - 8
Measurement of IgE on rat mast cells: relation to serum IgE and allergen-induced histamine release; van Toorenenbergen AW et al.; IgE antibodies against trinitrophenylated ovalbumin (TNP-OVA) were raised in Brown Norway rats by a highly reproducible immunization procedure . With these rats, the kinetics of the IgE response was studied, both for free circulating and cell-bound IgE . Cell-bound IgE was dissociated from peritoneal cell suspensions at acid pH; TNP-specific and total IgE in serum and in eluates of peritoneal cells were determined by radioimmunoassay . Both for total and TNP-specific IgE, a significant correlation was found between the serum IgE level and the amount of IgE molecules per mast cell . One day before TNP hapten could induce histamine release from mast cells, TNP-specific IgE appeared in the circulation and on the cells . These results suggest that also in active sensitization, there is a lag time for sensitization of mast cells.

Gerontology, 1983, 29(1), 49 - 63
Age-dependent change in a soluble factor responsible for Th-2 type of help in the indirect plaque-forming antibody response; Matsuzawa T et al.; A carrier-independent type of helper activity for the indirect plaque-forming response was detected in the supernatant of spleen cell suspensions from primed mice . The potency of the factor increased during the adult life of the spleen cell donor . The factor(s) acted antigen nonspecifically and across the major histocompatibility complex . The ability of spleen cells to generate this activity was lost after treatment of primed cells with anti-Thy 1.2, anti-Lyt 1.2, or anti-Ly 7.2, in the presence of complement . The capacity to generate the activity depended on the presence of cells which adhered specifically to surfaces coated with the sensitizing antigen . The production of factor(s) involved cooperation of nonadherent T cells from primed old animals and adherent cells from unprimed donors; the age-dependent change in factor-production did not depend on the age of the donor of adherent cells.

Oncodev Biol Med, 1983, 4(6), C71 - 8
Evidence for the presence of a precursor form to gamma-glutamyl transpeptidase in cultured yolk sac tumor cells; Yokosawa N et al.; The biosynthesis of gamma-glutamyl transpeptidase was studied in a yolk sac tumor cell line . Cell suspensions were labeled with {14C}leucine; immunoprecipitates were obtained with anti-gamma-glutamyl transpeptidase antibody and analyzed on SDS-polyacrylamide gel electrophoresis++ . A radioactive 80 000 dalton peak and peaks identical to heavy and light subunits were detected in the immunoprecipitates from the extract which was labeled for 15 min . After 45 min labeling, the 80 000 dalton species disappeared, although the two subunits were still observed . Tunicamycin-pretreated cells were labeled with {3H}glucosamine and {14C}leucine for 3.5 h . The 80 000 dalton species was glycosylated, and accumulated to a great degree compared to non-treated cells.

Virchows Arch B Cell Pathol Incl Mol Pathol, 1983, 43(2), 199 - 212
Interaction of three human malignant cell lines with chick hypoblast in culture; Van Peteghem MC et al.; The hypoblast (lower layer) was dissected from young chick blastoderms and explanted in vitro, where it formed an epitheloid sheet . Cells from the following malignant lines were explanted on top of the sheet both as aggregates and as cell suspensions: Hu456 human bladder carcinoma, SAOS-2 human osteosarcoma, LICR(LOND)-HN-4 laryngeal carcinoma . The interaction of the malignant cells with the hypoblast was studied by time lapse cinephotography, light microscopy, and transmission electron microscopy . All malignant cells penetrated through the hypoblast, so that a gradually enlarging hole formed in it . Apart from this common pattern of behaviour, the three types of malignant cells differed in their interactions with the hypoblast in the following ways . 1) Both the Hu456 and to a lesser extent the SAOS-2 cells brought about an initial retraction of the hypoblast so that a temporary cell-free space was formed . No such retraction occurred in response to the LICR-(LOND)-HN-4 cells . 2) Each of the three types of malignant cells migrated for some distance beneath the hypoblast, and in this area of underlap, there were differences in the amount and disposition of extracellular material . Thus, there was more extracellular material between the hypoblast and underlying SAOS-2 cells than between the hypoblast and underlying Hu456 cells, whilst there was no extracellular material between the hypoblast and underlying LICR(LOND)-HN-4 cells . Indeed, the hypoblast and LICR(LOND)-HN-4 cells often shared desmosomes . 3) When explanted as aggregates on hypoblast Hu456 and SAOS-2 cells left the corona and migrated as solitary cells underneath the hypoblast in contrast with control aggregates explanted on plastic . These cells which had migrated beneath the hypoblast were flatter than their corresponding control cells which had spread on the plastic substrate . The flatter cells appeared to have been using the extracellular materials as a substrate, rather than the plastic . Such differences in the migratory behaviour between experimental and control cultures were not observed with LICR(LOND)-HN-4 cells.

Virchows Arch B Cell Pathol Incl Mol Pathol, 1983, 43(1), 43 - 54
Immune suppression and histophysiology of the immune response . I . Cortisone acetate and lymphoid cell migration; Van den Broek AA et al.; Seven daily intramuscular (im) injections of cortisone acetate (25 mg/Kg b.w.) given to rats or rabbits produced, (i) a pronounced reduction in the numbers of small lymphocytes in thymus-independent areas, (ii) atrophy of the thymic cortex, (iii) atrophy of germinal centres and (iv) a consequent depressed production of germinal centre-derived cells . Lymphocyte depletion was not caused by cell lysis . Moreover cell traffic between peripheral lymphoid organs did not seem to be altered . A revival of the depressed germinal centres in cortisone-treated (inbred) rats could be achieved by a transfer of bone-marrow cell suspensions from normal, cortisone-treated or T-cell-deprived animals . It was concluded that cortisone acetate arrests the migration of B-lymphocytes from the bone marrow to germinal centres in peripheral lymphoid organs, and that the accumulations of lymphoid cells in the bone marrow of cortison-treated animals might be composed of immature or mature T- and B-lymphocytes.

Virchows Arch B Cell Pathol Incl Mol Pathol, 1983, 42(2), 161 - 72
Human dendritic reticulum cells of lymphoid follicles: their antigenic profile and their identification as multinucleated giant cells; Gerdes J et al.; The B-dependent areas of human lymphoid tissue contain non-lymphoid, non-phagocytic cells known as dendritic reticulum cells (DRC) . These cells can be detected only very occasionally in routinely stained histologic sections . Recently we were able to overcome this limitation by preparing a monoclonal antibody, termed R 4/23, that reacts selectively with DRC . Thus by using an optimized immunoperoxidase method applied to frozen sections, it is possible to detect DRC in situ . To determine the antigenic profile of DRC, serial frozen sections of human tonsils were immunostained with R 4/23 and a large panel of other monoclonal antibodies or conventional antisera . In addition, touch imprints of tonsils and cytocentrifuge slides of cell suspensions with increased concentrations of DRC were immunostained with these reagents . DRC proved to be positive for mu, gamma, alpha, kappa and lambda chains, complement component C3b, C3b receptors, C3d receptors, HLA-A,B,C antigens, human Ia-like antigens, common ALL antigen (cALLa), and antigens that are characteristic of the monocyte/macrophage lineages . DRC did not express delta chains, T cell antigens, or antigens that are expressed on interdigitating reticulum cells (IDC) and Langerhans cells . DRC in touch imprints and suspensions prepared from hyperplastic tonsils were found to be giant cells often with 10 or more nuclei . In certain cases of follicular hyperplasia and of centroblastic-centrocytic lymphoma, DRC with several nuclei were also detectable in situ . These results show that (1) the phenotype of DRC differs from that of all other cell types in lymphoid tissue, (2) this phenotype most nearly resembles that of cells of the monocyte/macrophage series, thus suggesting that DRC are related to these cell lineages, and (3) DRC are multinucleated giant cells.

Physiol Bohemoslov, 1983, 32(1), 1 - 9
Gamma-glutamyl transpeptidase activity in isolated cells and in various regions of the mouse brain during early development and aging; Lisy V et al.; Differences in the distribution of gamma-glutamyl transpeptidase (GGT) activity during early and late postnatal development of the mouse brain were studied at the cellular and regional level . From the 7th to the 25th day of brain development, GGT activity rose evenly in the neuronal perikarya, glial cells and in the neuropil . At 25 days and 5 weeks of age, a 1.5-fold enrichment was observed in nerve cell bodies and in neuroglia as compared with the neuropil and the original cell suspension . During the same period, an abrupt increase in GGT activity was observed in brain capillaries, and between the 10th day and the 5th week, this was 1.5-5 times higher than the enzyme activity in cellular elements and in the neuropil . This increase in enzyme activity in brain capillaries continued up to the age of 12 months, when it was succeeded by a decrease in GGT activity to the level found at 3 months . GGT activity in the choroid plexi rose significantly (more than double) between the age of 5 weeks and 18 months and was a whole order higher than GGT activity in 10 other regions of the mouse brain, whose development displayed no pronounced changes in the activity of the enzyme . A single dose of dexamethasone, or adrenalectomy, were used to demonstrate any participation by steroid hormones in the regulation of GGT activity in the brain tissue . Of the two techniques, only adrenalectomy led to a statistically significant decrease in activity in brain and liver tissue, while the apparent increase in GGT activity after a single dose of dexamethasone was not statistically significant.

Virchows Arch B Cell Pathol Incl Mol Pathol, 1983, 42(1), 25 - 32
Proliferation of antibody-forming cells within the lymphoid system; Geldof AA et al.; Mice were given subcutaneous priming injections of horse radish peroxidase (HRP) in the hind leg foot pads, and ten weeks later intravenous booster injections . Cell suspensions of popliteal lymph nodes, spleen and bone marrow were obtained at various time intervals after boosting and incubated in vitro for 1 h in the presence of 3H-thymidine (3H-TdR) . In each suspension the number of cells showing simultaneously anti-HRP antibody and incorporation of 3H-TdR was determined . Whereas in spleen and bone marrow only very few 3H-TdR labelled antibody-forming cells were observed, a clear proliferative response was noticed in the popliteal lymph nodes . Already on the first day after booster injection proliferation of cells showing incipient antibody synthesis was demonstrable in lymph nodes . It is strongly suggested that most - if not all - antibody-forming cells in these animals are derived from proliferating cells in the local draining lymph nodes . A relationship between the route of primary immunization and nature and location of the secondary proliferative response is discussed.

J Neurooncol, 1983, 1(2), 149 - 66
Chemosensitivity of malignant human brain tumors . Preliminary results; Bogdahn U; Tumor cell proliferation and morphological changes in tumor cells under the influence of different cytostatic agents were measured in an in vitro assay to determine chemosensitivity of human malignant brain tumors . Aliquots of 2 to 5 x 10(4) cells of a tumor cell suspension (prepared from biopsy specimen by mechanical and enzymatic disintegration) were incubated for 72 hr in the presence of different cytostatic agents . After another nine days of incubation in fresh medium, cell proliferation was calculated by incorporation of 14C-leucine and 3H-uridine and measurement in a liquid scintillation counter . Morphological changes in tumor cells were evaluated in Labtek tissue culture slides cultured under identical conditions . All drugs were tested in pharmacologically achievable concentrations . In vitro BCNU-sensitivity of 7/17 patients correlated with a postoperative recurrence free interval of 15.1 months, in vitro BCNU-resistance of 10/17 patients correlated with a postoperative recurrence free interval of 6.38 months (p less than 0.001) . Tumors of lower grades of malignancy (astrocytoma III, malignant ependymoma, medulloblastoma) in our series have a statistically significant higher median BCNU-sensitivity (63.5%, p less than 0.05) and are sensitive to more drugs (3.6 drugs, median out of six selected drugs, p less than 0.01) than tumors of higher grades of malignancy (astrocytoma IV, glioblastoma multiforme; 29.54% BCNU-sensitivity, 1.4 effective drugs) . In our series differences of BCNU-tumor sensitivity and number of effective drugs were not statistically significant if related to age and sex of tumor bearing patients . We think this assay provides the opportunity to test a large number of drugs in a very short period of time . Results may indicate alternative drug regimen for those patients resistant to conventional chemotherapy.

J Exp Zool, 1982 Dec 30, 224(3), 331 - 44
Isolated osteoclasts and their presumed progenitor cells, the monocyte, in culture; Osdoby P et al.; Osteoclasts were isolated from the endosteal surface of day 19 embryonic chick tibias by mild trypsinization . Osteoclast enrichment was achieved by passing cell suspensions through Nitex screening of selective sizes, including the eventual selective retention of osteoclasts on 12 micrometers polycarbonate filters or by sequential sieving through Nitex screens and fractionation on Percoll gradients . The enrichment procedures produced osteoclast populations of 50-75% based on morphological criteria with the latter isolation method providing populations with less matrix debris . The results of light microscopy, transmission and scanning electron microscopic observations indicate that osteoclasts can be maintained in culture for up to 10 days with retention of osteoclast morphology . This morphology includes a specialized ruffled plasma membrane, large numbers of mitochondria, lysosomes, as well as a multinucleated cytoplasm . Furthermore, acid phosphatase and butyrate esterase histochemical measurements support these morphological observations . In addition, chick hatchling circulating monocytes were isolated and purified by Ficoll-hypaque gradient centrifugation with subsequent adhesion to glass petri dishes . With time in culture, these cells form multinucleated cells, but lack the ultrastructural complexity of the isolated osteoclasts . This report describes a unique culture system to study osteoclast function and illustrates the similarities and differences of this system to the monocyte-to-giant cell culture system.

J Immunol Methods, 1982 Dec 30, 55(3), 327 - 36
Deletion of active human suppressor T lymphocytes from peripheral blood by Sephadex G-10 filtration; Hoffmann MK et al.; A method for antigen-specific generation of antibody-forming B cells in cultures of human peripheral blood mononuclear (PBM) cells based on the Mishell-Dutton system has recently been established in this laboratory . Comparing PBM cell cultures from healthy donors and from patients with advanced cancer we found the latter to be unresponsive in our assay . Passage of PBM cell suspension over Sephadex G-10 columns restored the response of patient PBM cells to normal levels . The cell population trapped on the column can be recovered and its inhibitory potential demonstrated by its graded addition to cells eluted from the column . The cell responsible for inhibition is sensitive to treatment with OKT8 antibody and complement, indicating its T cell nature . Passage of PBM cells from healthy individuals did not alter antibody responses substantially but made the activation requirements less stringent.

J Immunol Methods, 1982 Dec 17, 55(2), 221 - 9
Rapid isolation of mononuclear cells from buffy coats prepared by a new blood cell separator; Figdor CG et al.; A newly developed cell separator for the preparation and fractionation of buffy coat cells from human peripheral blood is described . In this cell separator buffy coats (BC-1) as routinely obtained from blood banks were used for the preparation of a second buffy coat (BC-2) with a volume of only 5-6 ml . A special fractionation device allowed sterile isolation of almost pure platelets and mononuclear cells with recoveries of 75 +/- 10% and 89 +/- 4% respectively . The white blood cell contamination of the platelet suspension never exceeded 20 X 10(6) leukocytes (i.e., less than 1 leukocyte per 5000 platelets) . Furthermore, the mononuclear cell suspensions were shown to be contaminated with only 3 +/- 2% granulocytes, whereas the white blood cell/red blood cell ratio was 2.6 +/- 1.6, so that they could therefore be directly used for further separation by means of centrifugal elutriation . These results indicate that this cell separator provides a rapid (+/- 1 h) isolation of both platelets and mononuclear cells without exposing the buffy coat cells to foreign substances like Ficoll or Percoll.

J Biol Chem, 1982 Dec 10, 257(23), 14187 - 91
Translational control of ferritin synthesis by iron in embryonic reticulocytes of the bullfrog; Shull GE et al.; The regulation of ferritin synthesis by iron was examined in the reticulocytes of bullfrog tadpoles where the induction was 40- to 50-fold, increasing from 0.17 +/- 0.05% of total protein synthesis ({3H}leucine incorporation in cell suspension) to 7.4 +/- 1.6% following intraperitoneal injection of ferric ammonium citrate . No significant difference was observed between the levels of ferritin mRNA in control or iron-induced cells, determined by translation of isolated mRNA in a wheat germ system, demonstrating that ferritin induction by iron occurs by a post-transcriptional mechanism . Total protein synthesis in the wheat germ system was half-saturating at 10 micrograms of mRNA/ml whereas ferritin synthesis increased linearly up to 40 micrograms of mRNA/ml, demonstrating that the ferritin mRNA is translated with high efficiency relative to the total proteins synthesized . Studies with the cap analogue 7-methylguanosine-5'-monophosphate, suggest that cap binding is not directly involved in the high translational efficiency of the ferritin mRNA in the wheat germ system . The results indicate that iron-modulated changes in the availability of ferritin mRNA for translation, coupled with the high translational efficiency of the ferritin message, can account for the induction of ferritin synthesis by iron in embryonic erythroid cells.

Biochim Biophys Acta, 1982 Dec 8, 693(1), 262 - 4
Decreased deformability in aging erythrocytes; Bartosz G et al.; Deformability of bovine erythrocytes separated according to density (and age) was estimated by a modified Teitel's filterability test, the centrifugational test of Sirs, and viscosity measurements of cell suspensions . Both youngest and oldest erythrocytes were found to be less deformable than middle-aged cells, a result speaking against any chief role for deformability in the recognition of senescent erythrocytes and their removal from the circulation.

Am J Pathol, 1982 Dec, 109(3), 343 - 58
Ultrastructural localization of lactoferrin and iron-binding protein in human neutrophils and rabbit heterophils; Parmley RT et al.; Lactoferrin in marrow and blood granulocytes from rabbits and humans was stained with an immunoferritin method . Iron-binding protein(s) was localized by the staining of granulocytes with acid ferrocyanide after saturation of the iron-binding protein with iron . The latter was most readily accomplished by treatment of the glutaraldehyde-fixed cell suspension with 1% saponin, followed by treatment with an iron-nitrilotriacetate (Fe-NTA 3mM:4mM) solution, adjusted to pH 7.0 with NaHCO3 . The affinity of purified lactoferrin and transferrin for radioiron after such treatment was minimally diminished . Both immunoferritin and iron-binding methods heavily stained osmiophiliuc (phospholipid-containing) mature primary granules in late promyelocytes, myelocytes, and polymorphonuclear cells . Early promyelocytes containing abundant immature primary granules lacked immunoferritin or iron staining . Trypsin digestion of rabbit marrow cells considerably diminished the cytochemically demonstrable iron-binding capability of the mature primary granules . Specimens sequentially stained for peroxidase and immunostained for lactoferrin or cytochemically stained for iron-binding protein confirmed that lactoferrin and iron-binding protein were in peroxidase-positive primary granules . Some peroxidase positive granules appeared to lack staining for lactoferrin and iron-binding proteining protein, and all secondary granules uniformly lacked staining . Treatment of human neutrophils with phorbol myristate acetate demonstrated early release of granules containing iron-binding protein with subsequent agglutination of neutrophils and attachment of iron-binding protein to the cell surface . In summary, this study distinguishes at least two subpopulations of primary granules and identifies lactoferrin and an iron-binding protein(s) in a subpopulation of peroxidase-positive primary granules in rabbit heterophils and human neutrophils.

Anat Embryol (Berl), 1982 Dec, 165(3), 405 - 13
Isolated osteoclasts in primary culture: first observations on structure and survival in culture media; Zambonin Zallone A et al.; Osteoclasts were isolated mechanically from the medullary bone of laying hens kept 7 days on a low calcium diet . Osteoclast enrichment was achieved with 3-4 sedimentations of the cell suspension in test-tubes prepared by layering on the bottom with BSA 10% in MEM-HEPES or PBS, above which the cells were suspended in MEM-HEPES or PBS . The final suspension of osteoclasts was cultivated in MEM with 10% FCS for 3 weeks . The cultures were observed by phase-contrast and scanning electron microscopy (SEM) . By the third day, the osteoclasts were completely spread onto the plastic dishes and a variety of morphologies and of intercellular contacts was established . Osteoclasts in culture do not lose their morphology; they survive for long periods and can be used in many experimental systems.

Invest Ophthalmol Vis Sci, 1982 Dec, 23(6), 787 - 95
Biosynthesis of procollagens and collagens by tissue explants and matrix-free cells from embryonic chick cornea; Kao WW et al.; Tendon and cornea of 17-day-old chick embryos primarily synthesize type I collagen as the major collagenous component nd type V collagen as a minor collagenous component . Matrix-free cells isolated from cornea and tendon synthesize and secrete mainly type I procollagen . A short lag period for the secretion of collagenous polypeptides into the medium of cell suspensions is observed . There is no significant difference in the synthesis and secretion of type I procollagen between matrix-free cells from tendon or cornea, except that cornea cells contain more free pro alpha 2(I) chains . The pro alpha 2(I) chain does not contain interchain disulfide linkages and can be separated from type I procollagen by ion exchange chromatography . However, no free pro alpha 2(I) chain can be detected in the medium of cell suspensions, suggesting that free pro alpha 2(I) is probably not secreted normally and may be rapidly degraded intracellularly . These results suggest that proper chain composition of type I procollagen is not regulated at the transcriptional or translational levels . Rather it is determined by the primary structure of the pro alpha chains and/or a posttranslational factor.

Cancer Res, 1982 Dec, 42(12), 5147 - 51
Drug and hormone sensitivity of estrogen receptor-positive and -negative human breast cancer cells in vitro; Goldenberg GJ et al.; A clonogenic assay of long-term breast cancer cell cultures in vitro has been developed to provide a highly reproducible method with which to quantitate tumor cell killing by hormones and/or cytotoxic chemotherapeutic agents . Monolayer cultures of estrogen receptor-positive MCF-7 human breast cancer cells and of estrogen receptor-negative Evsa T cells are harvested by treatment with 0.01% trypsin:0.02% EDTA in Hanks' balanced salt solution . Cell suspensions are treated with drug or hormone in serum-free medium for 1 hr at 37 degrees; treated cells are washed, plated, and cultured for approximately 14 days; and colonies consisting of greater than or equal to 30 cells are counted . Compared to estrogen receptor-positive cells, estrogen receptor-negative cells were 2-fold more sensitive to melphalan but were conversely 1.9-fold more resistant to Adriamycin; these differences were statistically significant (p less than 0.001) . Thus, response to cytotoxic chemotherapeutic agents appeared to be independent of estrogen receptor status . For cells treated with diethylstilbestrol, the dose of drug or hormone reducing the surviving cell fraction to 1/e (DO) for estrogen receptor-positive cells was 2.27 nmol/ml, and that for estrogen receptor-negative cells was 2.80 nmol/ml; this difference was not statistically significant . However, with tamoxifen therapy, the DO for estrogen receptor-positive cells was 0.601 nmol/ml, and that for estrogen receptor-negative cells was 3.64 nmol/ml; this 6-fold greater degree of resistance to tamoxifen of estrogen receptor-negative cells was highly significant (p less than 0.001) . Treatment of cells for 24 hr with 17 beta-estradiol stimulated proliferation not only of estrogen receptor-positive cells but also of estrogen receptor-negative cells . However, estradiol at concentrations up to 200 microM had no apparent cytocidal activity, as measured by the clonogenic assay . Furthermore, treatment of MCF-7 cells simultaneously with estradiol and either diethylstilbestrol or tamoxifen failed to reverse the cytocidal activity of those two agents . These findings suggest that, in the clonogenic assay described herein, diethylstilbestrol and tamoxifen may kill human breast cancer cells by an independent mechanism of action and that the cytocidal activity of diethylstilbestrol and the proliferative effect of 17 beta-estradiol appear to be independent of estrogen receptor status.

Aust J Exp Biol Med Sci, 1982 Dec, 60(6), 707 - 16
A comparison of counterflow centrifugation and unit gravity sedimentation in the separation of mouse bone marrow progenitor cells; Attwood PA et al.; Both Unit Gravity Sedimentation and Counterflow Centrifugation separate cells primarily on the basis of size . However, when performed concurrently on the same normal mouse bone marrow cell suspensions, Unit Gravity Sedimentation was found to be far superior in terms of the recovery of total nucleated cell load and the recovery and enrichment of two classes of functional progenitors: (1) those responsive to the combined stimulus of pregnant mouse uterus extract (PMUE) plus human spleen conditioned medium (HUSPCM), and (2), those responsive to PMUE alone . Neither technique was able to resolve PMUE from PMUE + HUSPCM progenitors . The major limitation of Counterflow Centrifugation for the separation of sub-populations of marrow cells is related to the complexity and range of cell sizes in marrow cell suspensions which lead to significant pellet formation within the elutriator chamber . Furthermore, cells undergoing Counterflow Centrifugation are subjected to forces not encountered in Unit Gravity Sedimentation . Therefore, the basis of the separations is not identical and this is reflected in cell volume distributions of comparable fractions separated by each method.

J Dent Res, 1982 Dec, 61(12), 1408 - 12
Isolation and characterization of inflammatory cells from the human periapical granuloma; Stern MH et al.; Twelve histologically-confirmed periapical granulomas were evaluated by conventional immunologic rosette assays for the presence of T-lymphocytes and complement receptor-bearing lymphocytes . A technique for dispersing the granuloma cells into suspensions was adopted to facilitate performance of the assays which were not applicable to tissue sections . Differential cell counts by an acridine orange vital dye method disclosed that the cell suspensions contained 30% macrophages, 44% lymphocytes, 15% plasma cells, and 12% neutrophils . Complement receptor-bearing cells comprised 17.9%, and T cells comprised 34.5% of the unseparated inflammatory cells . This study provides the first direct evidence of a predominance of thymic-derived lymphocytes in the lymphocyte compartment of the periapical granuloma . Analysis of the data shows that cell-mediated immunity most likely plays a role in the pathogenesis of the periapical granuloma.

J Immunol, 1982 Dec, 129(6), 2698 - 707
Surface phenotype of Peyer's patch germinal center cells: implications for the role of germinal centers in B cell differentiation; Butcher EC et al.; The surface phenotype of Peyer's patch germinal center lymphoid cells in the mouse is described . It is confirmed that most germinal center lymphocytes bind high levels of peanut agglutination (PNA), a lectin with specificity for terminal galactosyl residues . It is shown that germinal center lymphocytes can be identified in cell suspensions as a discrete PNAhi population distinct from other B cells, plasma cells, and most T cells, which bind only low levels of PNA . Using fluorescence-labeled PNA as a marker in dual fluorescence studies, we found that the majority of Peyer's patch germinal center cells are B lymphocytes: PNAhi Peyer's patch cells express B220, the B lineage-specific form of the T200 family of molecules, as well as low levels of surface Ig . They do not express the T cell-lineage antigens Thy-1, Lyt-1, or Lyt-2 (only 1 to 3% positive) . They bear lower levels of H2-K than PNAlo B cells, but two to three times the level of surface I-A-encoded determinants . A discrete but variable subpopulation of PNAhi Peyer's patch cells bear ThB in AKR/c mice, but BALB/c PNAhi lymphocytes are ThB- . About 10 to 30% bear surface IgM or IgG, but in contrast to essentially all PNAlo B lymphocytes in this site, they express no detectable surface IgD . The majority of Peyer's patch germinal center cells bear surface IgA, and this IgA is allelically excluded in F1 mice, indicating it is synthesized by the germinal center cells themselves . In fact, germinal centers contain most of the IgA-bearing cells in Peyer's patches (70 to 85%) . These findings lend considerable support to the concept that germinal centers in Peyer's patches are the site of generation of precursors of the IgA-secreting plasma cells that characterize mucosal immune responses, and also suggest that germinal centers may play an important role in the process of heavy chain class switching.

J Immunol, 1982 Dec, 129(6), 2529 - 31
In vitro inhibition of pancreatic B cell function by lymphocytes from diabetics with associated autoimmune diseases: a T cell phenomenon; Boitard C et al.; Lymphocytes from insulin-dependent diabetic patients were previously shown to suppress insulin release from mouse islet cells in vitro . Glucagon release was not suppressed . In order to further analyze this phenomenon, lymphocytes from three insulin-dependent diabetic patients with associated autoimmune diseases were treated by using the panning method for cell separation before testing on islet cell suspensions . The OKT3+, OKT3-, and OKT4- cell subsets were obtained . Insulin release was suppressed by the OKT3+ (T lymphocyte-enriched) subset, but not by the OKT3- (T lymphocyte-depleted) subset . These results suggest that T lymphocytes are directly involved in the suppression of insulin release in this model . Furthermore, the OKT4- (T helper-depleted) subset also suppressed insulin release.

Can J Physiol Pharmacol, 1982 Dec, 60(12), 1450 - 8
Some functional and morphological characteristics of an acutely dispersed purified cell suspension of rat lactotrophs prepared with Percoll; Milligan JV et al.; A cell suspension containing more than 90% lactotrophs can be prepared from enzymically dispersed adenohypophyses obtained from male rats pretreated with estradiol . The lactotrophs are separated from the mixed cell population by centrifugation on a discontinuous density gradient prepared from a commercial preparation of colloidal silica (Percoll, Pharmacia) . The method allows isopycnic separation of these delicate cells under very mild conditions; normal ionic strength and normal pH were maintained throughout the gradient, centrifugal acceleration did not exceed 1600 X g, and all procedures were done at room temperature . Histological verification that at least 90% of the cells were lactotrophs was done using specific immunoperoxidase staining . The functional capability of the lactotrophs was established by measuring the dose--response to the dopamine agonist bromocriptine and to thyrotropin-releasing hormone (TRH) . Bromocriptine decreased spontaneous release in a dose-related way over the concentration range of 10(-10) to 10(-8) M . TRH, which causes an in vivo release of prolactin (PRL) in estrogen-primed rats, produced a dose-related increase in the release of PRL over the concentration range of 3 X 10(-10) to 3 X 10(-8) M after the high spontaneous release had been previously reduced by bromocriptine (3 X 10(-8) M).

Biochim Biophys Acta, 1982 Nov 22, 692(3), 431 - 40
Uniform ionophore A23187 distribution and cytoplasmic calcium buffering in intact human red cells; Simonsen LO et al.; The divalent cation-selective ionophore A23187 has been used to characterize cytoplasmic Ca and Mg buffering, Ca2+-pump parameters and the properties of a Ca2+-activated K+-channel in intact red cells . A critical assumption in these studies has been that the ionophore causes a uniform increase in divalent cation-permeability in all the cells . This has now been tested directly in ATP-depleted human red cells by analysing the kinetics of ionophore-induced 45Ca-tracer and net Ca2+ fluxes . The experimental curves were all adequately fitted by single-exponentials at all ionophore concentrations tested . Moreover, statistical analysis of 61 individual tracer influx curves and of pooled data showed no trend towards fast second exponential components . These results demonstrate uniformity of ionophore distribution, ionophore-induced Ca2+-permeability, and cytoplasmic Ca-buffering among all the cells . Experiments involving mixing of cell suspensions with high and low original ionophore content, and involving ionophore extraction by albumin, demonstrate a rapid redistribution of ionophore among the cells, indicating that homogeneity of ionophoric effects is achieved through dynamic ionophore redistribution.

Biochem J, 1982 Nov 15, 208(2), 479 - 86
The use of N-methylprotoporphyrin dimethyl ester to inhibit ferrochelatase in Rhodopseudomonas sphaeroides and its effect in promoting biosynthesis of magnesium tetrapyrroles; Houghton JD et al.; N-Methylprotoporphyrin dimethyl ester inhibits ferrochelatase in isolated membranes of Rhodopseudomonas sphaeroides at low concentrations (around 10 nm) . Full inhibition developed after a short lag phase . The inhibition was non-competitive with porphyrin substrate . Addition of inhibitor to growing cultures of Rps . sphaeroides caused a decrease (near 40%) in cytochrome content and a severe inhibition of ferrochelatase; the excretion of haem into the medium by cell suspensions was also severely inhibited . The addition of N-methylprotoporphyrin dimethyl ester to suspensions of photosynthetically competent Rps . sphaeroides Ga caused excretion of Mg-protoporphyrin monomethyl ester . When added to mutants V3 and O1, magnesium divinylphaeoporphyrin a5 monomethyl ester and 2-devinyl-2-hydroxyethylphaeophorbide a were excreted, with maximum effect at around 3 microM-inhibitor in the medium . The results are interpreted to suggest that the inhibitor decreases concentration of intracellular haem, which normally controls the activity of 5-aminolaevulinate synthetase . Unregulated activity of this enzyme leads to overproduction of protoporphyrin, which is diverted to the bacteriochlorophyll pathway . Further control operates at magnesium protoporphyrin ester conversion in normal cells.

J Immunol Methods, 1982 Nov 12, 54(3), 331 - 41
Cryopreservation of antibody-coupled red cells for use in immunoassays; Pegg DE et al.; Red cells, trypsin-treated to render them more agglutinable and coupled with antiglobulin reagents, may be preserved by droplet freezing in liquid nitrogen . A 2% cell suspension in 0.45% w/v sodium chloride, 5% w/v sucrose and 10% w/v dextran 40 was used . After thawing the frozen pellets in phosphate-buffered saline at 40 degrees C, more than 80% cell recovery was obtained . Sheep and ox red cells preserved in this way were as satisfactory in antiglobulin and in reverse passive haemagglutination tests as unfrozen indicator red cells . Trypsin-treated human red cells coupled with anti-IgE could likewise be frozen and on reconstitution used to assay IgE in human serum . Reconstituted ox red cells were slightly less efficient in rosetting than cells which had not been frozen.

Nature, 1982 Nov 11, 300(5888), 194 - 7
Kinetics of sickle haemoglobin polymerization in single red cells; Coletta M et al.; Kinetic studies on solutions of purified haemoglobin S indicate that the rate of intracellular polymerization is an important variable in the pathophysiology of sickle cell disease . Until now, however, no experimental technique has been available to measure directly the kinetics of intracellular polymerization . Indirect methods, which use visual determination of cellular shape changes or changes in filterability of red cell suspensions, have given apparently conflicting results . Here we report our initial results on the application of a laser-photolysis, light scattering technique to measure directly the kinetics of haemoglobin S polymerization in single red cells . In our experiment, deoxyhaemoglobin S is rapidly formed by photolysing the carbon monoxide complex with an argon ion laser focused inside the cell, and the change in scattered light is used to detect the appearance of polymer . We find a very wide distribution of delay times, ranging from 1 ms to greater than 100 s, indicating that the polymerization inside red cells proceeds by the same nucleation and growth mechanism as in solutions of purified haemoglobin S.

Biull Eksp Biol Med, 1982 Nov, 94(11), 69 - 72
{Kinetics of oxygen consumption, luminescence of pyridine nucleotides and the cyanine dye 3',3'-diethylthiodicarbocyanine iodide after energizing Ehrlich ascites carcinoma cells with glucose}; Zinchenko VP et al.; It has been shown that in the absence of inhibitors of oxidative phosphorylation, changes in fluorescence of 3',3'-diethylthiodicarbocyanine iodide (dis-C2-/5/) in a cell suspension of Ehrlich's ascites carcinoma reflect those in the transmembrane mitochondrial potential . Addition of glucose to the cells in the presence of respiratory inhibitors similar to rotenon induces oscillations in the membrane mitochondrial potential due to H+-ATPase that uses glycolytic ATP . The described changes in energy metabolism parameters are determined by impairment of the interplay between glycolysis and oxidative phosphorylation . The low rate of cytoplasmic NADH oxidation by tumor cell mitochondria favors the latter's accumulation by the cytoplasm and glycolysis inhibition . Pyruvate has been shown to be responsible for NADH oxidation and to remove glycolysis inhibition.

Am J Physiol, 1982 Nov, 243(5), C247 - 53
Intracellular oxygen supply during hypoxia; Jones DP et al.; The intracellular supply of O2 to mitochondria was studied in single-cell suspensions of rat hepatocytes by measuring the O2 dependence of oxidation of cytochromes . Values were obtained by adding standardized O2-containing solutions to anaerobic cell suspensions and observing absorbance changes at wavelength pairs specific for cytochromes a + a3, c + c1, and b561 + b566 . Half-maximal oxidation, calculated relative to aerobic cells, occurred at 3.5, 6.2, and 4.9 microM O2, respectively . These values are similar to those for changes in cellular ATP/ADP, lactate/pyruvate, and NADH/NAD+ but are much higher than corresponding values for isolated mitochondria . Analysis of these data indicate that the diffusion coefficient in the region of mitochondria in situ is considerably smaller than the extracellular diffusion coefficient and suggests that a significant O2 gradient in the vicinity of mitochondria occurs under hypoxic conditions . These results suggest that the rate of O2 diffusion may be a critical factor for intracellular O2 supply to mitochondria during hypoxia.

Surgery, 1982 Nov, 92(5), 793 - 8
In vitro studies of parathyroid hormone release: effect of cimetidine; Saxe AW et al.; We studied parathyroid hormone (PTH) release from cell suspensions of fresh and cryopreserved human parathyroid tissue . A sensitive PTH assay with midregion specificity and adaptation of "micro" methods of cell plating permitted conservation of tissue (200 microliters aliquots) and high reproducibility (5 to 10 aliquots per experimental condition) . Viability of cells in suspension was confirmed by increased PTH release with increased incubation, increased incorporation of tritiated leucine with increased incubation, and incorporation of tritiated leucine following experimental incubation . Because of conflicting reports regarding the in vivo effect of cimetidine on PTH release, we incubated cells from both adenomatous and hyperplastic glands at 0, 10(-4)M, 10(-5)M, and 10(-6)M cimetidine (10(-5)M is therapeutic) and at 0.25, 1 to 1.2, and 2 mM Ca++ . Despite occasional statistically significant (P = 0.05) differences in PTH release with and without cimetidine, therapeutic levels of cimetidine had no consistent effect on PTH release from pathologic parathyroid tissue in vitro.

Immunol Lett, 1982 Nov, 5(5), 233 - 7
A monoclonal antibody to the HLA-A3 alloantigen; Bourel D et al.; Monoclonal antibody production recognizing the HLA-A3 antigen is described . The XI-23 antibody reacted with all of the 89 cell suspensions carrying the HLA-A3 antigen (100% cytotoxicity) among a total of 191 suspensions tested . No extra-reactivity or cross-reactivity was observed, particularly with that of HLA-A11 . This antibody can thus be considered as a good HLA-typing reagent.

J Natl Cancer Inst, 1982 Nov, 69(5), 1039 - 47
In vitro growth stimulation of human ovarian cancer cells by xenogeneic peritoneal macrophages; Welander CE et al.; For improvement of the in vitro growth rate of human ovarian tumors, an irradiated xenogeneic feeder layer of adherent peritoneal cells was added to such cultures . This adherent peritoneal cell population was obtained from oil-primed adult (C57BL/6J x DBA/2J)F1 mice and contained predominantly macrophages . After irradiation of the feeder layer with 3,000 rad 137Cs, cell suspensions prepared from fresh, solid human ovarian cancers were cocultivated with the feeder cells . Two end points were used to evaluate the growth enhancement added by such a feeder layer: 1) Primary monolayer cultures of ovarian tumors were successfully grown in continuous culture through five or more serial subcultures in 25 of 64 different patient specimens studied . Control cultures without the feeder layers did not continue growing through serial subcultures . 2) With the use of the soft agar clonogenic assay, addition of the feeder layer to the petri dishes increased the number of tumor cell colonies in each of six separate experiments, compared to the number seen in control plates without the feeder layer.

Transfusion, 1982 Nov-Dec, 22(6), 472 - 4
Detection and quantitation of fetal maternal hemorrhage utilizing an enzyme-linked antiglobulin test; Riley JZ et al.; Existing methods to evaluate fetal-maternal hemorrhage depend upon red blood cell agglutination or blood film elution techniques . These tests are insensitive and difficult to quantitate and reproduce . An enzyme-linked antiglobulin test was evaluated to determine its suitability for clinical testing of postpartum candidates for Rh immune globulin administration . Prepared mixtures of Rh positive fetal and Rh negative adult red blood cells approximating fetal maternal hemorrhage ratios of 0-2.0 percent were studied . In 43 assays, the enzyme-linked antiglobulin test consistently detected Rh positive fetal red blood cells in the 0.5 and 0.25 percent mixtures representing a 25 ml and a 12.5 ml hemorrhage, respectively, in a 70-kg woman . The 0.125 percent red blood cell suspension was positive in 85 percent of the assays and the 0.0625 percent suspension was positive in 56 percent of the tests . Agglutination testing by Du variant technique failed to detect 25 percent of the 0.5 percent mixtures . Only 45 percent of tests with the Rh immune globulin crossmatch detected the 0.5 percent mixture . A modified Kleihauer-Betke procedure was as sensitive, but less reproducible than the enzyme-linked antiglobulin test . Forty-seven Rh immune globulin candidates were studied to assess the quantity of fetal maternal hemorrhage . Fourteen patients (29.8%) had detectable Rh positive red blood cells by enzyme-linked antiglobulin tests but all hemorrhages were less than 12 ml; agglutination tests did not detect any fetal red blood cells . We conclude that the enzyme-linked antiglobulin test is a simple, sensitive, and objective procedure for detecting small amounts of Rh positive red blood cells in Rh negative blood and should be applicable to clinical testing of post-partum Rh immune globulin candidates.

Immunology, 1982 Nov, 47(3), 415 - 21
T-cell recirculation in the sheep: migratory properties of cells from lymph nodes; Reynolds J et al.; T lymphocytes derived from different sources in sheep were compared for their ability to recirculate from blood to lymph . Nylon wool columns were used to prepare T-cell-enriched populations from efferent intestinal lymph, efferent prescapular lymph and from cell suspensions of mesenteric lymph nodes and prescapular lymph nodes . With each animal, T cells from two of the above sources were labelled in vitro, one population with fluorescein isothiocyanate the other with rhodamine isothiocyanate; both populations were returned to the animal at the same time by intravenous injection . The intestinal lymph and prescapular lymph were continuously monitored to compare the recirculating properties of the two populations of T cells . This technique led to confirmation of the earlier reports in sheep of a preferential recovery of intestinal lymph T cells and of prescapular lymph T cells in the lymph from which the cells were originally collected . This phenomenon was much less evident with T cells from mesenteric nodes and prescapular nodes and in a number of experiments a random pattern of recirculation occurred . It is concluded that there are differences in the composition of the T-cell population in a node compared with that of the lymph draining the node . The advantages of using fluorescently-labelled cells to study lymphocyte migration are discussed.

J Immunol, 1982 Nov, 129(5), 2202 - 5
Immunologic deficiency during experimental Chagas' disease (Trypanosoma cruzi infection): role of adherent, nonspecific esterase-positive splenic cells; Kierszenbaum F; The involvement of adherent splenic cells in the production of deficient lymphocyte responses during the acute phase of experimental Chagas' disease was investigated . When cultured together, purified adherent splenocytes from mice acutely infected with Trypanosoma cruzi caused a significant reduction in the responses of normal mouse spleen cells to T and B cell-specific mitogens . Similar observations were made when infected mouse adherent splenocytes were co-cultured with normal mouse nonadherent cells . Exchange of adherent cells in infected mouse spleen cells suspensions for adherent cells from uninfected mice resulted in increased responses to stimulation with the T and B cell mitogens tested . Treatment of infected mouse cell suspensions with indomethacin improved the responsiveness of these cells to the mitogens . These results support the concept that the immunosuppression that is characteristic of experimental acute Chagas' disease is at least in part mediated by an adherent cell population and is dependent on a prostaglandin-mediated mechanism.

J Natl Cancer Inst, 1982 Nov, 69(5), 1163 - 74
Exclusive binding of immunoglobulin to Fc gamma receptors on macrophages in 3-methylcholanthrene-induced murine tumors; Lindsay JM et al.; Immunogenic, chemically induced, murine tumors generally are infiltrated with large numbers of Fc gamma receptor (Fc gamma R)-positive cells, most of which are mononuclear phagocytes . This study demonstrates in inbred C3H/HeN mice that such immunogenic tumors contain large amounts of cell-bound immunoglobulin (Ig) and that, within the sensitivity limits of indirect immunofluorescence, the tumor-associated Ig is bound exclusively to host-derived Fc gamma R-positive cells . Depletion of Fc gamma R-positive cells from tumor cell suspensions removed all Ig-bearing cells . Specific competitive elution of the endogenous Ig with heterologous nonimmune IgG established that the endogenous Ig was bound through the Fc gamma R . Tumor-associated Fc gamma R-positive, Ig-bearing cells could be detected with soluble antigen-antibody complexes, and mononuclear phagocytes to which the exogenous complexes were bound expressed Fc gamma R at a level significantly above normal . The role of this macrophage-associated Ig in determining the level of interaction between host cells and the tumor remains to be determined.

J Pathol, 1982 Nov, 138(3), 219 - 33
Comparison of mycobacterial granulomas in guinea-pig lymph nodes; Narayanan RB et al.; A study was made of mycobacterial-induced granulomas in guinea-pig lymph nodes . Live BCG (Pasteur) induced a granuloma containing epithelioid cells while Cobalt irradiated Mycobacterium leprae induced a granuloma comprised of phagocytic macrophages . The granulomas were quantitated by measurement of lymph node weight and the areas of infiltration in histological sections . The time course of granuloma formation induced by Co-irradiated M . leprae was veary different from the time course of the granuloma formation induced by BCG . Collagen synthesis assessed by incorporation of 14C-proline into collagenase sensitive protein was greater in lymph nodes draining the site of injection of Co-irradiated BCG than those draining the site of injection of Co-irradiated M . leprae during the first 10 weeks . Collagen synthesis was delayed in the nodes from animals injected with live BCG for at least 10 weeks . Single cell suspensions of draining lymph nodes containing granulomas consisted of lymphocytes and large cells (epithelioid cells and macrophages) . A high proportion of the large cells were found to be non-adherent in the live BCG-induced epithelioid cell granuloma . In contrast, M . leprae-induced granulomas contained a high percentage of adherent large cells . In both the granulomas, the majority of large cells were esterase positive and showed the presence of fibronectin . Most of the large cells in the granulomas did not carry receptors for the Fc component of IgG or the C3 component of complement and did not exhibit peroxidase activity.

J Invest Dermatol, 1982 Nov, 79(5), 277 - 82
Use of the fluorescence-activated cell sorter to quantitate and enrich for subpopulations of human skin cells; Morhenn VB et al.; A variety of immunologic staining techniques were compared in a quantitative study of antigen expression by human epidermal cells . Virtually all nucleated epidermal cells express beta 2-microglobulin, which is associated with HLA-A, -B, and -C antigens, whereas only about 4% expressed T6, an antigen expressed by Langerhans cells but not other cells in the skin . With the fluorescence-activated cell sorter (FACS), epidermal cell suspensions were selectively enriched 10- to 15-fold for T6-positive Langerhans cells . An average of 6.5% of cells were specifically stained by anti-HLA-DR antibody . When dispersed cells stained with anti-DR plus peroxidase were examined with the technique of immunoelectron microscopy, only mononuclear leukocytes (probably Langerhans cells) were stained . After separating HLA-DR positive skin cells with the FACS, the DR-positive population but not the DR-negative population stimulated proliferation of allogeneic responder lymphocytes, indicating that sorted cells are metabolically active . We conclude that HLA-DR antigen is not expressed by keratinocytes in normal human skin cell suspensions and that the FACS can be used to selectively enrich or deplete skin cell suspensions of antigenically distinct subpopulations such as Langerhans cells.

Clin Exp Immunol, 1982 Nov, 50(2), 426 - 33
In vitro mixed skin cell lymphocyte culture reaction (MSLR) in man: analysis of the epidermal cell and T cell subpopulations; Czernielewski J et al.; The nature of normal human epidermal cells (EC) and peripheral blood (PB) cells that react in vitro in allogeneic mixed skin cell lymphocyte culture reaction (MSLR) was investigated using monoclonal antibodies (MCAB) specific for cell subpopulations . T cells and helper-inducer and suppressor-cytotoxic T cell subsets were defined by OKT3, OKT4, OKT8 MCAB, respectively, whereas, among EC, Langerhans cells were characterized by reactivity with OKT6 or anti-HLA-DR MCAB . MSLR were conducted with untreated cell suspensions as controls and cells suspensions depleted of various functionally active cell subset(s) . Two approaches were used for cell depletion: (1) complement (C')-mediated lysis by MCAB of T cells, T cell subsets, HLA-DR or OKT6 positive cells; (2) panning of PB cells or EC after pre-incubation with the appropriate MCAB to deplete or enrich (OKT6) cell suspensions with the respective cell subset . Responses in MSLR were abolished after treatment of PB cells with OKT3 + C' or OKT4 + C', significantly reduced with OKT8 + C'; they were abolished after incubation of EC with anti-HLA-DR + C' and significantly reduced with OKT6 + C' . After panning, OKT3 and OKT4 depleted populations did not proliferate in MSLR while OKT8 depleted populations respond as controls . OKT6 depleted EC were not able to stimulate PB cells, yet proliferation rates were increased after stimulation by OKT6 enriched EC . Data show that helper-inducer T cells (OKT3+; OKT4+) play the major role in MSLR and that the presence of Langerhans cells is necessary for the stimulation of PB cells . They also suggest that co-operation between helper and suppressor cells is necessary for an optimal response . Differences in results using either OKT6 or anti-HLA-Dr-C'-mediated treatment of EC may be related to differences in the cellular expression of these markers by EC.

Cytometry, 1982 Nov, 3(3), 177 - 81
Flow sorting of mouse pancreatic B cells by forward and orthogonal light scattering; Nielsen O et al.; Mouse islet cell suspensions were separated into subpopulations based on forward low angle and orthogonal scattered light . Separation fractions were analyzed for hormone content by radioimmunoassays . The pancreatic B-cells had the highest orthogonal light scattering, while they overlapped in forward low angle light scattering with other endocrine (glucagon, pancreatic polypeptide and somatostatin containing cells) and nonendocrine cells . Living and paraformaldehyde fixed islet cell suspensions showed similar distributions of combined light scattering.

Biochim Biophys Acta, 1982 Oct 29, 699(1), 49 - 52
Inhibition of DNA synthesis does not influence the H1 histone synthesis in Tetrahymena pyriformis; Klenow S; In the protozoan Tetrahymena pyriformis the DNA synthesis is stopped immediately and completely after addition of one of the two DNA synthesis inhibitors methotrexate + uridine and hydroxyurea to a cell suspension . However, the present experiments show, that the accumulation of labeled H1 histone in the inhibited cells is almost totally unaffected for more than two-thirds of a cell cycle after addition of either inhibitor.

Klin Wochenschr, 1982 Oct 15, 60(20), 1279 - 87
{Bone marrow transplantation in patients with leukemia}; Wilms K et al.; Between October 1979 and March 1982, bone marrow transplantations were performed by the Tubingen Group for BMT on 19 patients with acute leukemia in remission and on one patient with chronic myelocytic leukemia in chronic phase . The conditioning regimen consisted of 2 x 60 mg cyclophosphamide/kg and 10 Gy whole-body irradiation with the linear accelerator . The lung dose was limited by shielding to 8 Gy . In 15 patients, the bone marrow cell suspension of the donor was preincubated with antihuman T-cell globulin (AHTCG) for prophylaxis of graft-versus-host disease (GVHD) . All patients showed prompt engraftment of donor cells with good hemopoietic function and complete chimerism . Under reverse isolation in sterile units, no severe bacterial or fungal infections were seen in the phase of bone marrow aplasia . Twelve in twenty patients survived between 25 and 900 days . A severe GVHD was seen only in two patients - one after preincubation with AHTCG . One patient died from relapse of his leukemia, another patient had a testicular relapse which was treated with local radiotherapy . Major problems were seen with chronic GVHD (six patients) and infectious complications, most importantly interstitial pneumonia, in the late post-transplant period.

C R Seances Acad Sci III, 1982 Oct 11, 295(5), 355 - 8
{Use of a new rheometer for the study of the filtrability of a suspension of sickled red cells as a function of PO2}; Martin-Caburi J et al.; The filtration time of a small volume (0.1 ml) of red cell suspension from normal (AA), heterozygous (AS) and homozygous (SS) subjects for sickle cell disease was investigated as a function of PO2 The curve of filtration time of AS and SS red cell suspensions was biphasic . At high values of PO2, the progressive reduction of filtrability of sickle cell suspensions with decreasing PO2 occurred without new change in morphology of most of the cells . In contrast, at lower PO2 the apparent filtrability was improved and the cells were sickled." However the red blood cells were retained by the filter and the "solvent" filtrability was improved because rigid and highly deformed sickled cells did not clogged completely the pores of the filter . This study allowed to distinguish a new concept of apparent filtrability for red blood cells in sickle cell disease.

J Clin Lab Immunol, 1982 Oct, 9(1), 7 - 11
Immune transfer using spleen fragments; Gagnon RF et al.; The adequacy of spleen slices for the purpose of adoptive immune transfer was evaluated in rodents . Transfers using spleen cell suspensions and spleen slices were conducted in parallel from hyperimmune donors to non-immune intact and irradiated syngeneic recipients . It was demonstrated that the level of immune responsiveness attained with transfer of spleen slices was at least equal to and most often greater than that achieved with transfer of spleen cell suspensions.

Br J Exp Pathol, 1982 Oct, 63(5), 463 - 71
Some characteristics of the participation of lymphocytes in non-immune inflammation; Bersani Amado CA et al.; Leucopenic rats were injected i.v . with either allogeneic or syngeneic lymphocyte suspensions and their capacity to respond to carrageenin tested at various intervals after the injection of the cells . The carrageenin-induced inflammatory response, which is characteristically attenuated in leucopenic animals, was partially but none the less definitely restored when allogeneic cells were transfused 20 min before the irritant, and less so when they were given 1 or 2 h before . Syngeneic cells were effective in restoring this response up to 2 h after injection . Only a small fraction of the exogenous cells, either allogeneic or syngeneic, remained in the blood, and this for short intervals . One hour after administration of cell suspensions blood leucocyte counts again attained previous values . The same rate of disappearance of the cells from blood was observed in splenectomized rats . Removal of the cells from circulation was thought to reflect "homing" into lymphoid tissues and other tissues, but not selectively in the spleen . Lymphocyte constituents, obtained by cell disintegration and injected i.v . into leucopenic rats, did not cause any change in blood leucocyte counts and yet restored the previously inhibited inflammatory response to carrageenin, as much as the viable cells . It is suggested that inflammation-producing materials, either deposited in tissues or formed or transformed at a site of injury, might gain access to the lymphatics during the early stages of a non-immune, inflammatory response, and "stimulate" lymphoid cells in adjacent lymphoid tissues either to release pro-inflammatory factors or to migrate to the site of injury, where they would exert pro-inflammatory activities . Apparently, only a small number of these cells in involved in the response.

J Clin Invest, 1982 Oct, 70(4), 747 - 51
Generation of leukotrienes by purified human lung mast cells; MacGlashan DW Jr et al.; Although mediator release from mast cells and basophils plays a central role in the pathogenesis of human allergic disease, biochemical studies have been restricted to rat peritoneal mast cells and basophilic leukemia cells because they could be easily purified . We have used two new techniques of cell separation to purify human lung mast cells to 98% homogeneity . Lung cell suspensions were obtained by dispersion of chopped lung tissue with proteolytic enzymes . Mast cells were then purified from the suspensions by countercurrent centrifugal elutriation and affinity chromatography . The purified mast cells released both histamine and slow-reacting substance of anaphylaxis (SRS-A) (leukotriene C and D) during stimulation with goat anti-human IgE antibody . Moreover, these preparations were able to generate significant quantities of SRS-A (32 +/- 7 x 10(-17) LTD mole-equivalents/mast cell) at all stages of purification, indicating that a secondary cell is not necessary for the antigen-induced release of SRS.

J Immunol, 1982 Oct, 129(4), 1642 - 8
Prepubertal orchidectomy induces thymic abnormalities in aging (NZB X SJL)F1 male mice; Dumont F et al.; Female but not male (NZB X SJL)F1 (NS) mice develop abnormalities of their intrathymic lymphocyte population in the course of aging . To determine the role played by androgens in this sex-related difference, we monitored the evolution of the cellular composition of the thymus in NS males deprived of androgens by prepubertal orchidectomy . Although in young mice this operation resulted in a twofold enlargement of the thymus, there was no histologic alteration or major change in the surface phenotype and mitogenic reactivities of the thymocytes, which suggests that all thymocyte subsets were increased to the same extent . In 12-mo-old control (BALB/c X SJL)F1 mice, prepubertal orchidectomy also produced an equal expansion (1.4-fold increase) of all thymocyte subsets . In contrast, in 12-mo-old orchidectomized NS males, there was a marked depletion of the thymic cortex and a hyperplasia of the medullary lymphoid tissue reflecting the selective expansion of a subset of phenotypically mature T cells (dull Thy-1+, Lyt-1+2+/-, dull PNA+) together with the emergence of intrathymic surface immunoglobulin-bearing cells . These latter cells probably represented B cells because there was a concomitant augmentation of the mitogenic responsiveness in vitro of thymic cell suspensions to lipopolysaccharide . Such thymic abnormalities induced by prepubertal orchidectomy in old NS males resemble those occurring spontaneously in the NS females . This suggests that the absence of thymic disease in intact NS males is primarily due to a suppressive effect of androgens.

Cancer Res, 1982 Oct, 42(10), 4086 - 9
Heterogeneity in growth pattern and drug sensitivity of primary tumor and metastases in the human tumor colony-forming assay; Schlag P et al.; The human tumor colony-forming assay was used to compare primary tumors with their metastases . Cell suspensions were prepared from 38 primary tumors and from metastases taken simultaneously from the same patient . Considerable differences were observed . Cloning efficiency was significantly higher in the cell suspensions taken from metastases than from material from the primary tumor . In 10 paired samples which allowed in vitro drug sensitivity testing, the data showed no satisfactory correlation in resistance of sensitivity to cytostatic drugs between primary tumor and metastases . The regression coefficient calculated for the different cytostatic agents (Adriamycin, bis(chloro)ethylnitrosourea, 5-fluorouracil, 4-hydroxyperoxycyclophosphamide) varied between 0.02 and 0.1 . The results indicate that experiments designed to analyze chemosensitivity of tumor cells in the tumor colony-forming assay should be interpreted with caution . In particular, in vitro sensitivity data obtained from the primary tumor may have severe limitations in planning treatment of metastatic disease.

J Immunol, 1982 Oct, 129(4), 1384 - 90
Formation of IgE-binding factors by rat T lymphocytes . V . Effect of adjuvant for the priming immunization on the nature of IgE binding factors formed by antigenic stimulation; Uede T et al.; Rats were immunized with keyhole limpet hemocyanin (KLH) included in complete Freund's adjuvant (CFA) or with the same antigen absorbed to aluminum hydroxide gel . Incubation of their spleen and MLN cells with homologous antigen resulted in the formation of IgE-binding factors . The nature of IgE-binding factors formed by antigenic stimulation was different depending on the adjuvant employed for priming and the period after immunization . If the lymphoid tissues were obtained 2 to 4 wk after antigen priming, IgE-binding factors derived from lymphoid tissues of KLH-alum-primed animals selectively potentiated the IgE response, whereas those produced by the lymphoid tissues of KLH-CFA-primed animals suppressed the IgE response . Analysis of purified IgE-binding factors from KLH-primed spleen cells revealed that the factors were heterogeneous with respect to their m.w., and that IgE-binding factors of a m.w . of 13,000 were responsible for the biologic activities . Thus, the 13,000-dalton factors obtained from KLH-alum-primed spleen cells selectively potentiated the IgE response, and those from KLH-CFA-primed spleen cells suppressed the IgE response . The selective formation of either IgE-potentiating factors or IgE-suppressive factors by KLH-primed spleen cells was observed only when the cells were stimulated by specific antigen . When the same KLH-primed spleen cells were incubated with IgE, IgE-binding factors formed in the cultures neither potentiated nor suppressed the IgE response . It was also found that stimulation of a mixture of BCG-primed spleen cells and KLH-alum-primed spleen cells with PPD resulted in the formation of IgE-suppressive factors; the stimulation of the same mixed cell suspension with KLH resulted in the formation of IgE-potentiating factors . The results collectively indicate that the nature of IgE-binding factors is decided by antigen-primed cells.

J Clin Invest, 1982 Oct, 70(4), 798 - 805
Mesenchyme-epithelial interactions in human endometrium . Prostaglandin synthesis in separated cell types; Gal D et al.; Glandular epithelium and stromal cells of human endometrium were separated and maintained in monolayer culture . At the time the cells became confluent, cell suspensions were prepared and incubated with {14C}arachidonic acid . Radiolabeled prostaglandin E2 and, to a lesser extent, prostaglandin F2 alpha and metabolites of these prostaglandins, were formed principally in stromal cells . There was considerably less prostaglandin formation in endometrial glands either after maintenance in monolayer culture or in freshly separated glands . In stromal cells of endometrium prostaglandin formation was linear with time of incubation for 2.5 min and with {14C}arachidonic acid concentrations up to 8 microM . When stromal cells and epithelial cells were combined, all prostaglandin formation could be accounted for by that produced in stromal cells . Little or no prostaglandin formation was detected in stromal cells from human adipose tissue or in fibroblasts from human genital or abdominal skin or human fallopian tube.

Pathol Biol (Paris), 1982 Oct, 30(8), 681 - 7
{Monoclonal antibodies (BL7 and BL9) having crossed reactivity with cells of the human epidermis}; Schmitt D et al.; The relationships between epithelial cells and immunocompetent cells could be approached by studying of the common antigens expressed by these two cell types . This paper reports the study of two monoclonal antibodies (BL7 and BL9) which react with epidermal cells . BL7 was obtained after immunization of mice with human thymic cell suspensions and BL9, after immunization with Raji cells . BL7 stained the epithelial network of the thymus and the basal cell layer of the epidermis . BL7 also reacted with the endothelial cells of the vessels of the dermis . This reactivity against the basal cell layer of the epidermis was observed in man, mice and rabbit . BL9 showed a reactivity against thymic epithelial cells and stained the membrane of the keratinocytes of the human epidermis . The antigenic expression revealed by BL9 decreased during the epidermal cell differentiation and disappeared in the horny layer . BL9 showed no reactivity with the epidermis of mice and rabbit . These two monoclonal antibodies are new tools in cutaneous immunopathology: BL7 is the first monoclonal experimental marker which identifies the basal cells of the epidermis, BL9 identifies an antigen related to the human epidermal cell differentiation.

Am J Physiol, 1982 Oct, 243(4), E319 - 24
Affinity alteration of insulin receptor induced by a phorbol ester; Grunberger G et al.; The effect of a phorbol ester tumor promoter, 12-O-tetradecanoyl phorbol-13-acetate (TPA) on 125I-insulin binding to human cells was examined . TPA markedly inhibits insulin binding to cultured human lymphocytes and macrophages but has a minimal effect on human fibroblasts . This inhibition is temperature, time, and concentration dependent . The inhibition of insulin binding to the cells at 37 degrees C occurs within minutes and diminishes by 6 h of incubation . Insulin binding is decreased by TPA whether the phorbol ester is added before, after, or simultaneously with 125I-insulin to the cell suspension . Scatchard analysis of binding to IM-9 lymphocytes indicates that TPA affects the affinity rather than the number of insulin receptors . The phorbol ester has only a small effect on 125I-human growth hormone binding in cultured human lymphocytes . TPA perturbs the insulin receptor of cultured human lymphocytes in a fashion similar to its effect on the epidermal growth factor receptor of several other cell types . The specific mechanism of TPA action that affects the receptor of these two potent growth factors (i.e., insulin and epidermal growth factor), however, is unknown.

Endocrinology, 1982 Oct, 111(4), 1045 - 50
Angiotensin II receptors and prolactin release in pituitary lactotrophs; Aguilera G et al.; Logical properties of angiotensin II receptors in the rat adenohypophysis were analyzed in cultured rat pituitary cells incubated with angiotensin II and known stimuli of pituitary hormone secretion . PRL release during incubation for 3 h with 3 nM angiotensin II was consistently increased by 68 +/- 5%, comparable with that elicited by TRH (63.1 +/- 4%) . The ED50 of 0.5 nM for PRL release by angiotensin II was significantly lower than that of TRH (2.9 nM) in the same cell cultures . The antagonist analog {Sar1,Ala8}angiotensin II prevented the angiotensin-induced rise in PRL production but not that evoked by TRH, whereas dopamine and SRIF inhibited basal, angiotensin, and TRH-stimulated PRL release . Angiotensin II also caused a small increase in ACTH release but had no effect on the release of LH, TSH, and GH . Angiotensin II binding and PRL release were measured in partially purified lactotrophs prepared by elutriation, by which the initial cell suspension was separated into seven fractions . Most of the lactotrophs were present in the two fractions eluted at flow rates of 15.7 and 19.8 ml/min, as indicated by their immunoreactive PRL content . The 2.5- to 3.2-fold enrichment of lactotrophs was accompanied by a 2- to 3.5-fold increase in angiotensin II receptor concentration, with no change in binding affinity (Ka = 3.5 x 10(9) M-1) . In the same fractions, angiotensin II-induced PRL release was similarly increased by 1.6- to 3.5-fold above basal, compared with values of less than 1 in the initial cell suspension and other fractions . The preferential location of angiotensin II receptors in the lactotroph-containing fractions and the close correlation between angiotensin II binding sites and stimulation of PRL release indicate the functional importance of the pituitary angiotensin II receptor sites . These findings also suggest that angiotensin II could contribute to the physiological regulation of PRL secretion.

Am J Reprod Immunol, 1982 Oct, 2(5), 260 - 4
Suppression of maternal lymphocyte mitogenic responses by supernatants from short-term placental cell cultures; Rubinstein A et al.; Human placental cells were freed from lymphocyte contamination on discontinuous BSA gradients . Supernatants from short-term cultures of these purified placental cell suspensions were predominantly suppressive for maternal lymphocyte blast transformation to phytohemagglutinin and in the mixed lymphocyte culture . The suppressor factor was nondialyzable and did not contain IgG or human chorionic gonadotropin.

Eur J Biochem, 1982 Oct, 127(3), 559 - 66
Purification and properties of the coupling-factor ATPases F1 from Rhodopseudomonas palustris and Rhodopseudomonas sphaeroides; Muller H et al.; The coupling-factor ATPases from photosynthetically grown Rhodopseudomonas palustris and Rhodopseudomonas sphaeroides were purified by the same procedure to homogeneity . Gel chromatography on Sephacryl S-300 Superfine shortened the process of purification and improved its yield . Solubilization of the ATPase from both bacteria was found to be dependent on a specific sonication treatment of the cell suspensions, indicating a very weakly bound F1-ATPase in R . palustris . Depleted chromatophores could be restored in photophosphorylation and membrane-bound ATPase activities by adding the solubilized ATPase protein . The purified enzymes did not show a markedly trypsin-stimulated or dithiothreitol-stimulated activity . Isoelectric focusing and chromatofocusing revealed isoelectric points of 5.0 for both F1-ATPases . The molecular weights were determined by gel chromatography plus high-performance liquid chromatography . Hence, we calculated a molecular weight of 350000 for both F1-ATPases . Sodium dodecylsulfate/polyacrylamide gel electrophoresis revealed five subunits for both enzymes . Kinetic parameters, regarding substrate specificity, the effect of divalent cations, Km and Ki values for the membrane-bound and solubilized ATPases were determined.

Pflugers Arch, 1982 Oct 1, 394(4), 279 - 86
Functional impairments of human red cells, induced by dehydroepiandrosterone sulfate; Kon K et al.; A study has been made on the incorporation of dehydroepiandrosterone sulfate (DHAS), one of the most abundant adrenal C-19 steroids, into human red cells, and of the resulting effects on red cell functions . 1 . DHAS was incorporated into red cell membrane mainly by a partition mechanism: The apparent partition constant was small ({DHAS}cell/{DHAS}free = 1.34), indicating that DHAS in red cells would be easily removed by dilution . 2 . At least part of the DHAS taken up was apparently bound to band 3 protein and thereby was able to inhibit the exchange of intracellular and extracellular SO4(2-) (Ki = 70 micro M) . 3 . Using a fatty acid spin label, it was established that the presence of DHAS in lipid bilayer of the membrane increased the acyl chain motion in the middle portion of the membrane . 4 . DHAS induced echinocytosis of red cells . It is suggested that the increase in the viscosity of red cell suspension, the decreased deformability and the decrease in the deoxygenation rate of hemoglobin in the presence of DHAS probably reflect the presence of echinocytes . 5 . In the presence of plasma proteins, the incorporation of DHAs into red cells was remarkably suppressed.

J Immunol, 1982 Oct, 129(4), 1458 - 64
A monoclonal antibody against a surface antigen shared by human large granular lymphocytes and granulocytes; Rumpold H et al.; A monoclonal IgM antibody VEP13 was raised against a cell membrane antigen by immunizing BALB/c mice with a cell preparation enriched for human large granular lymphocytes (LGL) . These cells are known to include the effector cells responsible for natural killer (NK) and killer (K) cell activity . By indirect immunofluorescence, VEP13 antibody reacted with 21.4 +/- 7.9% of peripheral blood lymphocytes (PBL) and more than 95% granulocytes but not with B lymphocytes, monocytes, erythrocytes, thrombocytes, thymocytes, and immature bone marrow cells . Cell suspensions enriched for T cells by means of E-rosetting using neuraminidase-treated sheep red blood cells contained 14.7 +/- 5.5% of VEP13+ cells . When PBL were treated with VEP13 antibody and complement, NK activity against a variety of target cells was greatly reduced and K cell activity was also strongly affected . Enrichment experiments of VEP13+ cells by means of the fluorescence-activated cell sorter revealed that all the NK activity resides in the VEP13+ fraction . This cell fraction was a homogeneous population of LGL . Studying the coexpression of various other antigens as defined by monoclonal antibodies, 84.6 +/- 4.6% of VEP13+ cells also expressed M1, 81.6 +/- 5.5% T10, 47.9 +/- 14.4% Leu 2a, 51.6 +/- 17.0% human Lyt-3, and 6.0 +/- 4.0% HLA-DR antigens . Leu 1, Leu 3a, and T6 antigens were found to be absent from VEP13+ cells . We conclude that VEP13 antibody should be helpful in determining LGL in peripheral blood as well as in tissue sections of tumor patients.

Boll Soc Ital Biol Sper, 1982 Sep 30, 58(18), 1158 - 63
{Isolation and characterization of epithelial cells of colon mucosa: preliminary results}; Paolicchi A et al.; A procedure for the isolation of epithelial cells from rat and human large bowel is described; the minced tissue was carefully washed with saline and incubated for 45 min in a collagenase-jaluronidase solution; the dispersion of the epithelial cells was achieved by a subsequent treatment with the calcium chelator EDTA . The isolated cells were characterized by cytological and histochemical (PAS, alkaline phosphatase, N-acetyl-D-glucosaminidase) procedures; viability index was assessed by the trypan blue exclusion test; the ability to grow in culture on both liquid and semisolid media was also tested . This method can be successfully applied either to normal or neoplastic colonic mucosa, the resulting cell suspension being suitable for further characterization.

Life Sci, 1982 Sep 13, 31(11), 1103 - 10
Influence of cholecystokinin on hypothalamic-stalk median-eminence-extract stimulation of ACTH output from isolated pituitary cells; Sander LD et al.; The influence of cholecystokinin 33 (CCK33) on CRF-like stimulation of ACTH output was tested in vitro using isolated pituitary cells . ACTH was assayed using isolated adrenal cell preparations . The CRF-like material was contained in a crude acetic-acid extract of hypothalamic stalk median eminence (HSME) . CCK33, in doses of 1 U, 10(-3) U, and 10(-6) U/ml cell suspension had no influence on basal or ACTH-stimulated corticosterone output from isolated adrenal cells . Isolated pituitary cells responded in a dose-related fashion of HSME extract, however, the absolute response to a given dose of HSME extract varied according to the basal (non-stimulated) output of a particular cell preparation . CCK33, in the dose range tested, had no influence on basal ACTH output . In contrast, 10(-3) U/ml oc CCK33, which corresponds to a concentration of 8 X 10(-11) M, significantly inhibited the output of ACTH from isolated pituitary cells stimulated by 0.2 equivalents of HSME . Higher concentrations of CCK33 had a variable effect . We conclude that cholecystokinin may have a role in the regulation of HSME-stimulated ACTH output from the pituitary.

Clin Reprod Fertil, 1982 Sep, 1(3), 229 - 33
The rosette inhibition test in early pregnancy diagnosis; Koh LY et al.; PIP: The rosette inhibition test (RIT) was assessed for its ability to diagnose pregnancy around the time of implantation . A blind study was conducted on 20 sera consisting of 10 samples from pregnant women and 10 samples from nonpregnant women and males . Sera from pregnant women were collected 7 days postovulation from 10 women who attended the Flinders Medical Centre (Bedford, Australia) Fertility Clinic for artificial insemination with donor sperm and who subsequently were shown to have had a conceptual cycle . In the nonpregnant group, 6 of the samples wre from males and 4 were from women who were inseminated but who did not conceive during the treatment cycle and subsequently expierenced normal menstruation . Blood was collected from a male donor into sodium heparin tubes and the lymphocytes isolated using the Ficol-Hypaque method . 3 New Zealand white rabbits were injected weekly with 1 ml of cell suspension intravenously (into the ear vein) for 5 weeks . The rabbits were bled for antilymphocyte serum (ALS) 1 week after the 4th and 5th injections and the sera separated and stored at -70 degrees . The ALS was inactivated at 56 degrees Centigrade for 30 minutes before use . 6 male guinea pigs were bled monthly for complement . The RIT was carried out using the technique of Morton et al . (1977) with modifications . The rosette inhibition titre was taken to be reciprocal of the highest diluation of ALs in which there was 25% inhibition (or greater) of rosette formation compared to that observed in the control . 8 of the 10 samples from pregnant women had titres of 8 x 100,000 and the remaining 2 had tires of less than 4 x 100,000 . It was possible to confirm 8 very early pregnancies in the 10 sera from women subsequently proved to be clinically pregnant . In the nonpregnant group, 1 serum sample from a female who exhibited no clinical evidence of pregnancy gave a titre of more than 8 x 100,000 . Like Morton et al . (1977, the presence of an early pregnancy factor (EPF) was detected in the sera of a high proportion of women at a very early stage of pregnancy . The sera were obtained 7 days after ovulation and therefore presumptively around the time of implantation . The incidence of false positiveresults for the diagnosis of very early pregnancy appears to be quite low with this assay .

Gan To Kagaku Ryoho, 1982 Sep, 9(9), 1628 - 33
{Correlation between ara-C sensitivity and intracellular ara-C concentration in a cultured human lymphocyte model}; Abe I et al.; Sensitivity to cytosine arabinoside (ara-C), i.e.; the relative viable cell count on day 5 after incubation of the cell suspension (3 X 10(5) cells/ml) with 0.1 microM (2.4 ng/ml) ara-C, could be correlated with the plateau level of ara-CTP in eight out of 10 human lymphoblastic cell lines tested in vitro (correlation coefficient; r = -0.76, p less than 0.05) . The plateau ara-CTP level was estimated by the incubation of the cell suspension (5 X 10(6) cells/ml) with 0.1 microM 3H-ara-C for 4 hr . The ara-CTP level was certified to be determined by the capacity for not only ara-C phosphorylation but also the ara-CTP degradation in the B-cell lines, and, in addition, by the ara-C deamination in the T-cell lines.

Tsitologiia, 1982 Sep, 24(9), 1080 - 7
{Colony-forming capacity of precursor cells of granulomonopoiesis in the liver and spleen at different stages of human embryo- and fetogenesis}; Petrov NS et al.; The claster- and colony-forming capacities (CFF) of cells-precursors of granulomonopoiesis (CPGM) in the livers and spleens of 37 human embryos and 9 foetuses were studied in the agar system . The CFC of CPGM in the liver and spleen of 6-8 week old embryos were higher than those during the later embryogenesis . In haemopoietic cell suspension, separated from liver cells, the CFC of CPGM were lower than in the total cell suspension from the liver of 6-8 week old embryos to constant within 6-16 weeks of embryogenesis . The discussion involves stem cell heterogeneity in human embryogenesis, a possible role of the embryonal liver in regulating the number of granulopoietic cells-precursors and their functional activity.

Cell, 1982 Sep, 30(2), 427 - 37
Heat-shock-induced alterations of ribosomal protein phosphorylation in plant cell cultures; Scharf KD et al.; Heat shock of cell suspension cultures of tomato (Lycopersicon peruvianum) results in a rapid and reversible decline of the phosphorylation level of a single basic ribosomal protein of the small subunit (tentatively identified as ribosomal protein S6) . Simultaneously, phosphate labeling of several acidic ribosomal proteins of the large subunit is enhanced . Data on the temperature-dependent distribution of S6 subspecies and on the kinetics and reversibility of S6 phosphorylation are given . The decreased phosphorylation of S6 at temperatures higher than 35 degrees C coincides with the onset of heat shock protein synthesis and precedes a decline of the mitotic index . Recovery from heat shock is characterized by S6 rephosphorylation and, subsequently, leads to an abnormally high mitotic index.

J Gen Virol, 1982 Sep, 62 (Pt 1), 127 - 36
Vaccinia virus-induced changes in {Na+} and {K+} in HeLa cells; Norrie DH et al.; A flame photometric technique is described for determining average values for intracellular {Na+} and {K+} in HeLa cells . Ion measurements were made on unwashed cells disrupted ultrasonically in the presence of residual medium; corrections for the latter were made by measurement of extracellular volume in cell plus medium preparations using 125I-labelled polyvinylpyrrolidone (PVP) as the marker in an isotopic dilution technique . Accurate measurement of the volume occupied by the cells was critical and required a concentration step . This was achieved by concentrating cell suspensions in a microhaematocrit centrifuge using calibrated capillary tubes . Most reliable values were obtained in our system using HeLa S3 (suspension) cells grown as monolayers, which were removed by EDTA and held in suspension for a minimum of 2 h . Uninfected HeLa cells had values of 20 to 30 and 110 to 120 mM for Na+ and K+ respectively . At 13 h after inoculation with vaccinia virus, a dramatic reversal in {Na+} and {K+} occurred, but throughout the infection cycle the total {Na+ + K+} varied little . The significance of these data is discussed in relation to theories of virus-induced modulation of protein synthesis in infected cells and in cell-free systems.

Inflammation, 1982 Sep, 6(3), 305 - 18
A rapid quantitative assay for activated neutrophils; Bullock WW et al.; We present here a rapid, sensitive, and convenient assay for activated human PMNs based on detecting the decreased optical density (OD) of aggregated cell suspensions . This quantitative assay uses an ELIZA machine to measure OD changes, with time, of activated cells (5 X 10(5) cells/well) in microtiter plates . The assay is sensitive, detecting aggregation induced by as little as 0.0001 microgram/ml of LPS, or lymphokines in Con-A-activated supernatant diluted 1/500 . The assay permits analysis of 400 separate PMN suspensions on the same day starting with less than 80 ml of blood.

Cancer Res, 1982 Sep, 42(9), 3724 - 8
Unique cell surface glycoprotein expression in hairy cell leukemia: effect of phorbol ester tumor promoters on surface glycoproteins, cell morphology, and adherence; Lockney MW et al.; Hairy cell leukemia cells from eight different patients exhibited a characteristic cell surface glycoprotein pattern when labeled by the neuraminidase-galactose oxidase-NaB3H4 procedure . The diffuse high-molecular-weight glycoprotein band (Mr 230,000 to 300,000) was not seen in other leukemic cell types and may represent a specific hairy cell leukemia antigen . Hairy cell leukemia cells can be maintained as cell suspension cultures, but treatment with a variety of tumor-promoting phorbol esters caused the cells to adhere to plates, assume a fibroblastic elongated shape, and extend long processes . This dramatic morphological change was not associated with any change in surface glycoproteins.

Thymus, 1982 Sep, 4(5), 273 - 8
Enhancement of the in vitro colony-forming capacity of human T lymphocytes induced by levamisole; Pistoia V et al.; The colony-forming capacity of normal peripheral blood mononuclear cells and of TG-depleted cell suspensions induced by PHA was investigated in the presence of different concentrations of levamisole . The results obtained demonstrate that: (1) the drug enhances significantly the number of colonies formed by both mononuclear cells and TG-depleted cells at concentrations ranging from 5 x 10(-6) to 5 x 10(-8 M; (2) this stimulatory activity results from a direct effect on T cells; and (3) adherent accessory cells are unaffected by the in vitro treatment with levamisole.

J Reticuloendothel Soc, 1982 Sep, 32(3), 167 - 78
Characterization of nonlymphoid cells in rat spleen, with special reference to strongly Ia-positive branched cells in T-cell areas; Dijkstra CD; By use of a monoclonal antibody against Ia antigen in an immunoperoxidase method, strongly Ia-positive branched cells are found in the T-cell areas of the splenic white pulp of the rat . In order to further characterize these cells, enzyme histochemical characteristics, phagocytic capacity, and irradiation sensitivity have been studied . Evidence is presented that these strongly Ia-positive branched cells represent interdigitating cells . The influence of whole-body irradiation on interdigitating cells is discussed . Comparison with data from the literature on the in vitro dendritic cell isolated from spleen cell suspensions reveals many similarities between the described interdigitating cell in vivo and the dendritic cell in vitro.

J Natl Cancer Inst, 1982 Sep, 69(3), 709 - 13
In vitro infection of lymphocytes with Marek's disease virus; Calnek BW et al.; Suspension cultures of splenic lymphocytes, incubated at 41 degrees C, became infected with Marek's disease virus (MDV) following exposure to a) other infected lymphocytes, b) infected chicken kidney monolayer cultures, or c) cell-free MDV . Both viral antigen expression and virus isolation could be demonstrated after more than 40 passages made by the addition of fresh spleen cells at 2- to 3-day intervals . Susceptibility of spleen cells from bursectomized chickens was markedly lower than that of cells from intact birds . Furthermore, when spleen cell suspensions were depleted of cells having characteristics of bursa-derived cells, e.g., those with surface IgM, Fc receptors, or ability to adhere to nylon wool, the susceptibility of the cell suspension was diminished . Enrichment of the suspension with cells having those features enhanced overall susceptibility . The target cells for virus infection in vitro also were shown to be nonphagocytic, to be of low or medium density, and to bear Ia-like antigen . In vitro susceptibility to infection of spleen cells did not correlate with the genetic susceptibility of the donor to Marek's disease.

Immunology, 1982 Sep, 47(1), 157 - 63
In vitro effect of (3-amino-1-hydroxypropylidene)-1,1-bisphosphonic acid (APD) on the function of mononuclear phagocytes in lymphocyte proliferation; de Vries E et al.; The effect of (3-amino-1-hydroxypropylidene)-1,1-bisphosphonic acid (APD) on pokeweed mitogen-stimulated and non-stimulated cultures of peripheral blood mononuclear cells was studied in vitro . It is shown that APD can inhibit partially lymphocyte proliferation when added to a suspension of mononuclear cells before stimulation, but that lymphocyte proliferation can continue when the drug is withdrawn . In contrast; when APD is added to the cell suspension together with the mitogen, lymphocyte proliferation remains low even when the drug is withdrawn . Addition of different concentration of mononuclear phagocytes (MNP) to non-adherent cells, followed by stimulation in the presence of APD, indicates that APD acts on MNP function preferentially and does not affect lymphocyte proliferation.

J Histochem Cytochem, 1982 Sep, 30(9), 967 - 72
A stable propidium iodide staining procedure for flow cytometry; Deitch AD et al.; A propidium iodide (PI) staining procedure is described in which 50 micrograms/ml PI in 10(-2) M Tris, pH 7.0, with 5 mM MgCl2 is used to stain murine erythroleukemia cells (MELC) grown in suspension culture as well as single cell suspensions derived from rat kidney adenocarcinoma and human prostatic carcinoma . Specificity of staining of nuclear DNA is achieved by enzymatic removal of RNA using RNAse in the staining solution . Virtually identical histograms, with the same G1 peak height and closely similar coefficients of variation (CVs), are obtained using a wide range of RNAse concentrations on replicate samples of MELC if the incubation times are sufficiently prolonged when employing the lower enzyme concentrations . For 1 mg/ml RNAse on logarithmically growing MELC, 30 min incubation at 37 degrees C is needed to obtain a maximum G1 peak height and optimal CV and there is no significant change in the histogram if the incubation is prolonged to 4 hr . For every 4-fold decrease in RNAse concentration, the incubation time at 37 degrees C must be doubled to obtain the same maximal G1 peak height and optimal CV . Unfixed cell preparations, whether derived from suspension or monolayer cultures or from solid tumors, are stable for 2 or more weeks if stored at 4 degrees C between flow cytometric analyses and histograms are usually only minimally altered if the stained cell samples are stored for 1-2 months at 4 degrees C . Sample decay is associated with bacterial contamination . If sterile preparative techniques are used initially, subsequent contamination of the stained preparations may be minimized by adding sodium azide to the stained samples at 0.1% without influencing fluorescence intensity . Glycerine may be added to 10% and the samples slowly frozen for storage without altering DNA histogram shapes . The simplicity of sample preparation and the stability of the resulting stained cell samples makes this procedure suitable for repetitive comparative sampling of tissue and cell populations over prolonged time spans.

Transfusion, 1982 Sep-Oct, 22(5), 379 - 83
A study of granulocyte cytotoxins and detection of granulocyte allospecific antigens; Korinkova P et al.; Granulocytes were isolated b overlaying leukocyte-rich plasma discontinuous density gradients . The cell suspensions were 95 percent granulocytes . Sera of 383 polytransfused patients and 521 multiparous women were examined for lymphocytotoxins, granulocytotoxins, and granulocytoagglutinins . On the basis of these tests, sera were identified which contained only granulocytotoxins . These were further identified with more precision as to their titers and antibody absorption; reactivity with granulocytes, B, and T lymphocytes; and K 562 cell line . On the basis of calculated correlation coefficients, nine sera were selected which determined three granulocyte specificities having genotype frequencies consistent with the Hardy-Weinberg equilibrium.

Acta Virol, 1982 Sep, 26(5), 353 - 61
Comparison of the anti-influenza activities of antibodies and immune system cells administered intranasally to syngeneic mice; Maiorova LP et al.; A novel experimental approach was employed to investigate the protective role of cellular and humoral factors in antiviral immunity under conditions of their prophylactic administration into the tracheobronchial cavities of normal mice-recipients which were challenged 24 hr later with influenza virus A/PR/8/34 (H0N1) . The highest protective effect was afforded by intranasal administration of immune serum (91.3%) as compared with splenocytes (65.2%) . The addition of immune serum to cell suspensions or pooled lymphocyte and macrophage fractions previously isolated from the spleen increased the protective effect of these materials by 97--100% thus surpassing the protective effect of immune serum alone . The protective effect of passively administered cells from inflammatory exudates of peritoneal or tracheobronchial cavities was higher than that of a more concentrated splenocyte suspension; the results were comparable only if the number of splenocytes was 4- to 5-fold that of the exudate cells . The degree of the protective effect was strictly proportional to the concentration of immune system cells administered to the concentration of immune system cells administered to the syngeneic recipients.

J Immunol Methods, 1982 Aug 27, 53(1), 109 - 22
Albumin magnetic microspheres: a novel carrier for myelin basic protein; Ovadia H et al.; Myelin basic protein (MBP) of guinea pig origin was incorporated into magnetically responsive albumin microspheres . Protein-protein bonding and stabilization of the GPMBP microspheres by heating at 120 degrees C did not adversely influence their capacity to bind anti-MBP antibodies or demonstrably alter the encephalitogenic activity of the incorporated GPMBP . The magnetic properties of the particles and the fact that immunodeterminants of some of the incorporated MBP fortuitously were distributed on the exterior surfaces of the microspheres allowed a number of experiments to be carried out in Lewis rats for the first time: (a) selective capture and deletion of that particular subpopulation of lymphoid cells responsible for transfer of experimental allergic encephalomyelitis (EAE) represented within the lymph node cells (LNC) of donor animals sensitized to neutral antigen, (b) enhancement of in vivo uptake of MBP by macrophages (M phi s) contained in oil-induced peritoneal cell exudates and exposed briefly to MBP microspheres, and (c) preparation of cell suspensions specifically enriched with respect to MBP-containing M phi s.

Int J Cancer, 1982 Aug 15, 30(2), 197 - 202
Demonstration of the tumorigenicity of transformed rat kidney cell-lines by intravenous allotransplantation in the neonate; Hard GC; The tumorigenic potential of allogeneic rat kidney cell-lines transformed in vivo by a carcinogenic dose of dimethylnitrosamine (DMN) was investigated by intravenous injection of single cell suspensions into neonatal outbred rats within 24 h of birth . The cell-lines tested included mesenchymal populations obtained from DMN-induced renal mesenchymal tumors (RRMT-2,-8,-9) and transformed mesenchymal (TRKM-5,-7,-8) and epithelial (TRKE-1) cell-lines derived from the kidneys of rats treated only hours previously with the carcinogen . Additionally, spontaneous nephroblastoma-derived embryonal cell-lines (REN-1,-2) were included in the study to extend the potential of the neonatal rat system over a range of differentiation lineages . The transplantation system proved to be a rapid and efficacious assay for demonstrating the malignancy of the mesenchymal and embryonal cell-lines . The major sites for tumor growth were the lungs, heart and eye, but differences in organ predilection were observed for individual cell-lines . The transformed epithelial cell-line (TRKE-1) proved refractory to the single intravenous inoculation but was transplantation-positive when a follow-up subcutaneous dose was administered several days later . The resultant growths produced by the diverse differentiation lineages were histologically characteristic for the tumor tissue affiliate of each cell-line . The results demonstrate the utility of this transplantation system for testing the malignant potential of morphologically transformed cells across the allogeneic barrier as well as proving that DMN is capable of inducing malignant transformation in both mesenchymal and epithelial cell types of the rat kidney after a very short period of exposure in vivo.

Biochem J, 1982 Aug 15, 206(2), 367 - 72
Cytochrome a620 in Tetrahymena pyriformis . Reactions with carbon monoxide and oxygen at subzero temperatures and photochemical action spectra; Lloyd D et al.; 1 . Mitochondria-enriched fractions of the ciliate protozoan Tetrahymena pyriformis ST contained CO-reacting cytochromes b560 and a620 . 2 . A non-photodissociable oxygen-containing compound of cytochrome a620 was formed in whole cell suspensions at -114 degrees C after photolysis of CO in the presence of 200 microM-O2 . 3 . Electron transport, indicated by the oxidation of cytochrome a620 and cytochrome c, occurred at temperatures higher than -72 degrees C . 4 . Photochemical action spectra for the relief of respiratory inhibition of whole cells by CO obtained by using a liquid dye laser indicate that the only CO-reacting terminal oxidase detectable was cytochrome a620 . 5 . It is concluded that the alternative electron transport chains in this organism utilize non-cytochrome terminal oxidases.

J Biol Chem, 1982 Aug 10, 257(15), 8623 - 6
Siderophore iron transport followed by electron paramagnetic resonance spectroscopy; Ecker DJ et al.; Siderophore iron transport was followed in Ustilago sphaerogena using isotope transport assays coupled with EPR spectroscopy . EPR spectroscopy was used as a quantitative tool to follow the rate of reduction of siderophore iron(III) to iron(II) in the cell suspension by following the disappearance of the signal at g = 4.3 . This rate was compared with the rate of iron transport, measured by the disappearance of radioactively labeled iron from the medium . The transport of three iron chelates was examined: the ferric siderophores ferrichrome and ferichrome A, and iron(III) chelated to excess citrate . For the transport of ferrichrome, an iron(III) ionophore, the rate of reduction of iron(III) to iron(II) was significantly lower than the rate of uptake of isotope from the medium supernatant, which is consistent with the established mechanism of uptake of the entire complex followed by intracellular reduction to remove the iron from the ligand . However, the rate of reduction of ferrichrome A, a non-ionophore, was identical with the rate of transport of iron into the cell . Iron(III) citrate was reduced at a rate slightly lower than the rate of transport . These data suggest that reduction of iron(III) is involved in the transport of iron from ferichrome A and possibly from iron(III) citrate.

Can J Microbiol, 1982 Aug, 28(8), 982 - 6
Sensitivity of methanogenic bacteria to dicyclohexylcarbodiimide; Sprott GD et al.; Methanospirillum hungatei strains GP1 and JF1 when exposed to the adenosinetriphosphatase inhibitor N,N'-dicyclohexylcarbodiimide experienced a marked decline in growth rates, methane synthesis activities, and intracellular ATP concentrations . Although growth was inhibited, the intracellular ATP concentrations of all other methanogens tested were little affected by high concentrations of dicyclohexylcarbodiimide (500 microM) . Thus, for studies of ATP synthesis, or for ATP depletion in whole cell suspensions of methanogens, the use of dicyclohexylcarbodiimide appears limited to M . hungatei.

Exp Hematol, 1982 Aug, 10(7), 628 - 36
Electron microscopy of human fetal erythroid cells before and after cryopreservation; Orlic D et al.; Fetal erythropoiesis was studied in human livers at 10 to 12 weeks of gestation . The most primitive blood cells were often observed in large indentations of the surface of hepatocytes and the plasma membranes of the two cell types were adherent at sites of attachment . Erythroid cell maturation occurred predominantly in the lumen of the sinusoids . Cell suspensions obtained from fetal livers were centrifuged, frozen at -196 degrees C, thawed and studied by electron microscopy . The primitive cells were morphologically altered by these procedures . Changes included damage to mitochondria and cell membranes and vacuole formation . Erythroblasts, by comparison, were virtually intact and even displayed some indications of reestablished functions within 10 minutes after thawing.

Biull Eksp Biol Med, 1982 Aug, 94(8), 97 - 100
{Importance of cell contacts for the differentiation of the precursor cells of hematopoietic stroma in long-term bone marrow cultures}; Gurevich OA et al.; The composite adherent cell layer is produced in the culture of bone marrow fragments . On the contrary, in the culture of single cell suspension of bone marrow, only of monolayer of fibroblast-like cells and macrophages is created . After implantation of the adherent cell layer from the Dexter type culture but not from "fibroblast" culture beneath the renal capsule of syngeneic recipients the ectopic hemopoietic sites are formed . Regeneration of the adherent cell layer may occur only in the course of multilayer formation, i.e . one week after bone marrow transplantation but not three weeks after the formation of the adherent cell layer.

J Microsc, 1982 Aug, 127 (Pt 2), 227 - 31
An improved specimen table for the Balzers freeze etching system BAF 400; Wisse DM et al.; A time saving, simple and inexpensive adaptation of a Balzers specimen table is described . This specimen table is used in combination with the small specimen holders originally developed for the Denton Freeze-etching module . They are in particular suited for cell suspensions, and are easy to handle for freezing procedures . Ten replicas are produced routinely per freeze fracture run, with the aid of this combined system.

J Cell Physiol, 1982 Aug, 112(2), 298 - 300
The use of fluorescent dyes to measure membrane potentials: a critique; Johnstone RM et al.; Under controlled conditions, fluorescent cyanine dyes can be used to measure membrane potentials of cell suspensions . Similar changes in membrane potential can be followed both with fluorescent dyes and electrophysiological probes in response to changes in the ion composition of the medium . Recent reports that attempt to abrogate the use of the cyanine dyes in measurements of the membrane potential are misleading.

J Cell Biol, 1982 Aug, 94(2), 472 - 7
Specific pancreatic beta-cell surface antigens recognized by a xenogenic antiserum; Dyrberg T et al.; An antiserum (R4) from a rabbit immunized with suspensions of C57BL/61 ob/ob mouse islet cells contains antibodies which in a 125I-protein A radioligand assay can be demonstrated to bind to single cell suspensions of normal Naval Medical Research Institute (NMRI) mouse islet cells . The binding of 125I-protein A to islet cells was about four times that of normal rabbit serum (NRS) after incubation at a 1/600 dilution of R4 antiserum quantitatively absorbed to mouse spleen lymphocytes (R4A antiserum) and hepatocytes . Subsequent absorption of the R4A antiserum to islet cells significantly reduced the binding of 125I-protein A to islet cells incubated with the doubly absorbed serum . Immunoprecipitation of radiolabeled islet cell lysates followed by SDS polyacrylamide gel electrophoresis and autoradiography suggested that the R4A antiserum recognized a Mr 40,000 glycoprotein . This glycoprotein was not detected in spleen lymphocytes . Electron microscope detection of gold-protein A complexes suggested that the binding of islet cell surface antibodies was cell specific . islet cell suspensions incubated with R4A antiserum and gold-protein A showed that 86 +/- 3 gold particles were bound per 100 beta-cells (mean +/- SE for six experiments) . In contrast, the number of gold particles per 100 endocrine non-beta-cells was 8 +/- 1 which was similar to the number achieved with NRS (3 +/- 1) on all endocrine islet cells . Our observations suggest that the pancreatic islet cells, in particular the beta-cells, express a specific antigen.

J Natl Cancer Inst, 1982 Aug, 69(2), 415 - 23
Role of anchorage in the expression of tumorigenicity of untransformed mouse cell lines; Wells RS et al.; Cultured mouse cells were tested for tumorigenicity in nude mice with both a conventional assay (injection of cell suspensions) and a new test involving implantation of cells grown on gelatin sponges . Sublines of Balb/3T3 cells, obtained from different sources, varied in their tumorigenic potential with either assay . One subline (A) formed distinctive precancerous nodules only in the sponge assay; these nodules often became progressive after a latent period of 3-4 months . However, suspensions of cells of this subline also caused tumors after a similar latent period, but no nodular phase preceded tumor formation . Another subline of Balb/3T3 (M) has failed to form tumors in either assay . The Balb/3T3 sublines did not differ in vitro properties, such as low saturation density, failure to grow in methylcellulose, and monolayer morphology . A second experimental approach involved tests on nude BALB/c mouse-embryo fibroblasts at various passage levels . The cells were passaged from primary culture, through crisis, to heteroploid, established cell lines . Tumorigenicity was demonstrable earlier in the sponge assay, at which time in vitro parameters putatively associated with malignant behavior were unchanged . Possible relationships with the in vivo phenomenon of solid-surface sarcomagenesis are discussed.

Diabetes, 1982 Aug, 31 Suppl 4, 1 - 10
The surface concentration of H-2 antigens on mouse pancreatic beta-cells and islet cell transplantation; Parr EL et al.; Measurement of the concentration of H-2 complex antigens in the surface membrane of mouse pancreatic beta-cells by a quantitative immunoferritin-labeling technique revealed that the antigens were present in very small amounts on the beta-cells of five strains . A comparison of the labeling densities in the five strains suggested that beta-cells from C57BL/10Sn congenic strains have about half the H-2 antigen density of BALB/c and C3H/HeJ cells . In C57BL/10Sn mice the antigen density on beta-cells was slightly greater than that on erythrocytes, about 20% of that on thymocytes, and about 2.5% of that present on peritoneal macrophages . Intraperitoneal allografts of c57BL/10Sn islets were uniformly rejected by B10.BR/SgSn diabetic recipients only when the islets were accompanied by 10(7) peritoneal lymphoid cells . When transplanted without peritoneal cells, C57BL/10Sn islets were only marginally rejectable . In a group of nine such allografts, three diabetic recipients were permanently cured and three others showed rejection times of about 30 days . Sensitization of the three mice showing permanent cures, using 10(7) allogeneic peritoneal cells at about 40 days after the transplant, did not cause rejection of the allografts . Isogeneic transplantation of cell suspensions from dissociated islets of Langerhans was markedly less effective in controlling diabetes than intact islets, and dissociation did not obviously improve the rate of allograft survival . However, 5/19 diabetic mice receiving allografts of dissociated islet cells showed long-term reversals of diabetes that were unaffected by administration of 10(7) peritoneal cells at about 100 days after the transplant . Recipient mice whose diabetes was reversed by islet allografts and unaffected by specific sensitization had pancreatic insulin concentrations characteristic of diabetic mice . Our reversals of diabetes with untreated islet allografts may be due to the cleanliness of islet preparations obtained with a modified isolation technique, and to the very low density of H-2 complex antigens on C57BL/10Sn beta-cells.

J Exp Med, 1982 Aug 1, 156(2), 567 - 84
A natural model of immunologic tolerance . Tolerance to murine C5 is mediated by T cells, and antigen is required to maintain unresponsiveness; Harris DE et al.; A unique experimental model is described, where natural immunologic tolerance to a well-defined soluble native antigen (murine C5) is examined in congenic strains of mice that differ only by the presence or the absence of C5 . A highly sensitive hemolytic assay was developed to detect nanogram amounts of C5 as well as an assay of anti-C5 inhibition of C5 hemolytic activity . The latter was more sensitive than immunodiffusion . Two reciprocal approaches were used to study the cellular basis of tolerance in irradiated hosts of either strain . In the first, lymphoid cells from either strain were transferred to irradiated B10.D2OSN hosts that were lacking C5 and so would not hinder detection of anti-C5 antibody upon challenge with murine C5 . Second, lymphoid cells from either strain were transferred to irradiated B10.D2NSN hosts, whose native C5 provided the antigenic stimulus . The immune response of whole nonadherent spleen cell suspension as well as mixtures of T and B cells (separated on the basis of surface immunoglobulin) from either strain were studied . In addition, the duration of tolerance and the antigen requirement to maintain it in irradiated C5-deficient hosts repopulated with C5-sufficient spleen cells was examined . The positive control of irradiated C5-deficient hosts repopulated with syngeneic spleen cells showed a primary and secondary response to immunization . In contrast, C5-sufficient spleen cells failed to respond both in the primary and the secondary response . Because the unresponsiveness was not caused by antigen carryover and was not antigen specific, it represents central tolerance . In C5-sufficient irradiated hosts (where immunization was not required and antigen was present in natural form and physiological concentration), transfer of C5-deficient cells mediated a drop in C5 levels to 10-20% of that noted in unreconstituted controls . T and B cell mixing experiments from the two strains into deficient or sufficient hosts demonstrated that tolerance is T cell dependent and that C5-sufficient or -deficient B cells could cooperate with nontolerant C5-sufficient T cells to produce significant anti-C5 antibody or mediate a significant drop in C5 levels . In addition, the presence of antigen was necessary to maintain tolerance . In conclusion, these results show that (a) natural tolerance to C5 is an active process that is T cell dependent and requires the presence of antigen; (b) in this natural model, clonal abortion does not seem to occur; and (c) both tolerant and nontolerant B cells retain the capacity to produce autoantibody.

Infect Immun, 1982 Aug, 37(2), 800 - 4
Chemiluminescence response of peritoneal macrophages to parasitized erythrocytes and lysed erythrocytes from Plasmodium berghei-infected mice; Makimura S et al.; The chemiluminescence response of normal mouse peritoneal macrophages to parasitized erythrocytes isolated from mice 3 weeks after infection with Plasmodium berghei was examined . Only 4 of 12 animals showed positive responses, whereas 8 showed negative responses . Photomicrographs revealed that only in chemiluminescence-positive animals were parasitized erythrocytes attached to or phagocytized by macrophages . When lysed parasitized-erythrocyte cell suspensions were added to the peritoneal macrophages, chemiluminescence could be induced in all cases . The response was enhanced remarkably by the addition of very small amounts of immune serum . Normal macrophages activated in vitro by supernatant from antigen-stimulated spleen cells from immune mice showed much higher parasite-induced chemiluminescence responses than did nonactivated macrophages, especially in the presence of immune serum.

Biokhimiia, 1982 Aug, 47(8), 1370 - 4
{Peculiarities of specific binding of serotonin by blood leukocytes, peritoneal cells and synaptosomes of mice}; Stefanovich LE et al.; The properties of serotonin-active sites were studied on peritoneal cells, blood leukocytes and synaptosomes of mice (CBA line) . Treatment of cell suspensions with EDTA, ouabain, strophanthin, 2,4-dinitrophenol, dithiothreitol and trypsin demonstrated that serotonin binding by peritoneal cells and leukocytes depends on bivalent ions and K+, Na+-pump operation, requires energy and intact disulfide bonds and is determined by a protein structure . ATP and ADP were found to inhibit amine adsorption by peritoneal cells . These cells specifically bind ATP 10 times more intensively than leukocytes . The data obtained are suggestive of differences in the composition of serotonin-active structures of blood leukocytes, peritoneal cells and synaptosomes.

J Gen Virol, 1982 Aug, 61 (Pt 2), 271 - 5
Virus particles and glycoprotein excreted from cultured cells infected with varicella-zoster virus (VZV); Shiraki K et al.; Virus particles and virus proteins excreted from cultured human embryonic lung (HEL) cells infected with varicella-zoster virus (VZV) were examined by electron microscopy and affinity column chromatography using an antibody to VZV followed by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) . Approximately 1 X 10(9) to 2 X 10(9) virus particles/ml with no detectable infectivity, of which 30 to 80% were enveloped, were observed in the culture fluid 48 to 72 h after infection, when cytopathic effect (c.p.e.) appeared . In the sonicated infected cell suspension, 1 X 10(9) to 2 X 10(9) virus particles/ml, of which 30 to 50% were enveloped, were observed and the virus particle/infectivity ratio was approx . 10(6):1 . The culture fluid of infected HEL cells labelled with {35S}methionine or {3H}glucosamine was centrifuged at 1000000 g for 2 h to remove virus particles and the supernatant was examined for excreted virus proteins . Affinity column chromatography of the supernatant using immobilized human zoster convalescent serum, led to the isolation of virus antigens which were analyzed by SDS-PAGE . Polypeptides with mol . wt . of approx . 115K and 45K, both of which were glycosylated, were detected, suggesting that these VZV glycoproteins were excreted from the infected cells.

J Cell Physiol, 1982 Aug, 112(2), 302 - 5
The use of fluorescent dyes to measure membrane potentials: a response; Smith TC; The use of fluorescent cyanine dyes to estimate membrane potential in cell suspensions has been considered . Several problems related tot he application of the dyes have been reviewed . These problems include: 1) alteration of the membrane potential (Em) and factors involved in establishing Em by the dyes themselves, 2) the effects of altered energy metabolism on the fluorescent response of the dyes and on Em, and 3) calibration of dye fluorescence . Recent reports that advocate the use of the fluorescent dyes are misleading.

Clin Exp Immunol, 1982 Aug, 49(2), 355 - 63
Cytoplasmic as opposed to surface Ia antigens expressed on human peripheral blood lymphocytes and monocytes; Hofman FM et al.; Peripheral blood leucocytes were examined for the presence of HLA-Dr or Ia-like antigen on the cell surface and in the cytoplasm . Surface staining of viable cell suspensions, using monoclonal anti-Ia antibody in both immunofluorescence and immunoperoxidase methods gave similar results . Using a newly developed immunoperoxidase double staining procedure, which stains the cells when either viable or fixed, both the surface and cytoplasm of the individual cells were labelled and examined . Twenty to thirty percent of the mononuclear cells were positive for Ia staining on the cell surface and in the cytoplasm . Morphologically these positive cells were identified as both lymphocytes and monocytes . A small percentage (5-9%) of these monocytes and lymphocytes consistently labelled only cytoplasmic Ia determinants . In contrast, only surface binding was observed with monoclonal antibodies recognizing T cell subpopulations . The ability to detect intracellular Ia antigenic determinants is of value in distinguishing stages of lymphocyte and monocyte development.

Nature, 1982 Jul 22, 298(5872), 377 - 9
Germinal centre B cells: antigen specificity and changes in heavy chain class expression; Kraal G et al.; Germinal centres are histologically defined aggregates of blast cells that occur in B-cell areas of lymphoid tissues after antigenic stimulation . They are believed to be associated with the development of B-cell memory and plasma cell (especially secondary, IgG and IgA) responses . Recent studies of murine lymphoid tissues have defined cell-surface markers that distinguish germinal centre B cells from other mature B cells, permitting their identification and characterization in cell suspensions . Here we have used these markers to define and study germinal centre cells in lympho nodes, and have found that they constitute a unique population of B cells which (1) arises in response to antigenic stimulation, (2) contains nearly all of the demonstrably antigen-specific B cells in the stimulated organ, (3) bears surface IgM after primary stimulation and (4) as a population, demonstrates isotype switching to a predominant population, demonstrates isotype switching to a predominant surface IgG phenotype after secondary stimulation with specific surface IgG phenotype after secondary stimulation with specific antigen . These findings demonstrate that germinal centres are a major site of proliferation and differentiation of antigen-specific B cells in vivo, and suggest that the germinal centre microenvironment may have an important role in heavy chain class switching during B-cell responses.

Chem Biol Interact, 1982 Jul 15, 41(1), 15 - 24
Effects of potassium dichromate on ATP content of mammalian cells cultured in vitro; Debetto P et al.; In order to elucidate the mechanism of the cytotoxic activity of hexavalent chromium (Cr(VI)), the alterations of intracellular ATP levels induced by potassium dichromate in cultured hamster fibroblasts (BHK line) have been studied . Two kinds of treatment procedures were adopted: (1) BHK cell suspensions were exposed to 0.05--1.00 mM K2Cr2O7 in Hanks' balanced salt solution (BSS) for up to 180 min and ATP concentrations were determined immediately after the exposure to Cr(VI) . A decrease of ATP content was observed with 0.25--12.00 mM K2Cr2O7 but only in the case of the highest dose was it related in a linear fashion to the duration of the treatment . (2) Cells were preincubated in BSS for 30 min with 0.05--1.00 mM dichromate . They were then reincubated in Eagle's minimal essential medium (MEM) for up to 180 min and ATP was measured at different time points . Immediately after the exposure to chromium all the treated cultures showed a depletion of ATP content . However while the cells treated with 0.25--0.25 mM dichromate rapidly resumed ATP levels very similar to that of the control, no recovery was detected in cells treated with 0.50 and 1.0 mM K2Cr2O7, even after 180 min . The observed effects have been attributed to the oxidizing activity of Cr(VI), which subtracts electrons from electron donors involved in metabolic pathways producing ATP, and to the ability of Cr(III), deriving from Cr(VI) reduction, to form stable coordination complexes with ATP precursors and enzymes involved in ATP synthesis.

Biochem J, 1982 Jul 15, 206(1), 121 - 9
On the mechanism by which hormones induce the release of Ca2+ from mitochondria in the liver cell; Whiting JA et al.; 1 . The abilities of dinitrophenol, NaCl, Ruthenium Red and the Ca2+-selective ionophore A23187 to release 45 Ca2+ from isolated hepatocytes and liver mitochondria (incubated at 37 degrees C in the presence of 0.1 microM-free Ca2+, Mg2+, ATP and phosphate ions) were compared with the action of adrenaline on 45Ca2+ release from isolated hepatocytes . The effects of adrenaline were most closely described by those of the ionophore A23187 . 2 . In isolated hepatocytes, a release of 45Ca2+ and stimulation of O2 utilization similar to that induced by adrenaline was observed in the presence of 500 and 20 microM-arachidonic acid respectively . The effect of arachidonic acid on 45Ca2+ release was not specific for this unsaturated fatty acid . 3 . Inhibitors of arachidonic acid metabolism, including indomethacin and eicosa-5,811,14-tetraynoic acid, did not block the effects of adrenaline on 45Ca2+ or glucose release from isolated hepatocytes . 4 . The ability of adrenaline to stimulate 45Ca2+ release from isolated hepatocytes was rapidly reversed after the subsequent addition of phenoxybenzamine to the cell suspension, and was completely blocked by 0.5 mM-dibucaine . 5 . The results are consistent with the action of a Ca2+-selective ionophore in the mechanism by which adrenaline induces the release of Ca2+ from mitochondria in the liver cell and indicate that it is unlikely that arachidonic acid or a metabolite of arachidonic acid is involved in this process.

J Biol Chem, 1982 Jul 10, 257(13), 7666 - 71
Role of exogenous cholesterol in regulation of adrenal steroidogenesis in the rat; Verschoor-Klootwyk AH et al.; Rat steroidogenic tissues take up cholesterol, and it has been suggested that this process plays a regulatory role in steroid hormone synthesis . To provide evidence for this hypothesis, we carried out studies in lipoprotein-deficient rats . Lipoprotein deficiency, achieved by treating male rats with pharmacological amounts of estradiol, led to profound lowering of plasma cholesterol (8 +/- 2 versus 54 +/- 4 mg/dl) and adrenal cholesteryl ester content (113 +/- 57 versus 747 +/- 108 micrograms/organ) . Basal serum corticosterone levels were decreased by 50%, and the response to adrenocorticotropic hormone (ACTH) was totally abolished . Injection of high density lipoprotein (HDL) to estradiol-treated animals restored the response of corticosterone to ACTH . Comparable in vitro studies with adrenal cell suspensions obtained from lipoprotein-deficient rats confirmed the in vivo data . Measurement of {14C}acetate incorporation and uptake of both HDL- and low density lipoprotein (LDL)-cholesterol in these adrenal cells showed a progressive increase with the duration of estradiol treatment, and neither of these two phenomena was altered by ACTH . These results provide in vitro and in vivo evidence for the hypothesis that normal adrenal steroidogenesis depends upon cholesterol delivery from plasma . Furthermore, under the conditions studied, ACTH does not stimulate adrenal de novo cholesterol biosynthesis nor the uptake of either HDL- or LDL-cholesterol.

J Reprod Fertil, 1982 Jul, 65(2), 425 - 9
Estimation of viability of bovine granulosa cells; Metcalf MG; Granulosa cells harvested from non-atretic, antral follicles of cow (and pig and sheep) ovaries were incubated over glass cover slips in medium containing 20% (v/v) donor calf serum . Cell attachment to the cover slips was rapid, being in most cases complete within 3 h at 37 degrees C . There was little further change over the next 20 h . The number of bovine granulosa cells which attached to a cover slip was proportional to the volume of cell suspension added to the medium; and the amount of oestradiol secretion by attached cells in a testosterone-enriched medium increased in parallel with their number . Granulosa cells which did not attach within 3 h produced little oestradiol . There was no clear relationship between the number of nigrosin-impermeable cells in suspensions prepared from different follicles and plating efficiency . It is concluded that the 3-h attachment of granulosa cells in culture is a useful measure of the number of viable cells in a suspension and is to be preferred to less direct techniques based on dye exclusion.

Clin Exp Immunol, 1982 Jul, 49(1), 225 - 9
Isolation of human monocytes by rosetting with antibody coated human erythrocytes and isopycnic gradient centrifugation; Zembala M et al.; Human erythrocytes O,Rh+ (D) coated with hyperimmune human anti-serum formed rosettes primarily with monocytes in peripheral blood mononuclear cell suspensions . The rosetting cells (EAhu-FRC), separated on a Lymphoprep gradient were found in the pellet and were up to 95% pure monocytes as judged by cytochemical criteria . A two step procedure involving depletion of T lymphocytes followed by the isolation of EAhu-RFC yielded monocytes up to 98% pure . The isolated cells did not respond to mitogens in vitro, but they act as accessory cells in mitogen-induced T lymphocyte proliferation . This simple method may be useful in studying monocyte function in human disease as it does not require a large amount of blood.

Cancer Res, 1982 Jul, 42(7), 2861 - 6
Reactivation of S-adenosylhomocysteine hydrolase activity in cells exposed to 9-beta-D-arabinofuranosyladenine; Helland S et al.; 9-beta-D-Arabinofuranosyladenine (ara-A) inactivates isolated S-adenosyl-L-homocysteine (AdoHcy) hydrolase (EC 3.3.1.1) as well as AdoHcy hydrolase in intact cells . Whereas the inactivation in cell-free systems is an irreversible process, the AdoHcy hydrolase activity in rat hepatocytes exposed to ara-A gradually recovered upon prolonged incubation of the cells in a medium devoid of ara-A . This process, tentatively termed reactivation of the enzyme, was nearly totally dependent on a high level of adenosine deaminase in the extracellular medium, which induced a decrease in intracellular content of adenosine as well as ara-A . Reactivation of intracellular enzyme was inhibited by adenosine deaminase inhibitors {2'-deoxycoformycin and erythro-9-(2-hydroxy-3-nonyl)adenine} and the synthetic substrate for AdoHcy hydrolase, 3-deazaadenosine . An inhibitor of protein synthesis (cycloheximide) was without effect . Homocysteine, which protected the intracellular AdoHcy hydrolase against inactivation by ara-A, induced no reactivation of the enzyme . The half-life of the intracellular ara-A-AdoHcy hydrolase complex was about 90 min and was not affected by adenosine deaminase, 3-deazaadenosine, or homocysteine added to the cell suspension . However, the rate of elimination of the complex in the hepatocytes exceeded the rate of reactivation of AdoHcy hydrolase . Thus, the elimination process accounted for the reactivation, but not correlation between these two processes was observed . Reactivation of intracellular AdoHcy hydrolase caused a pronounced fall in cellular content of AdoHcy . The possibility that reduced cellular level of AdoHcy induced the reactivation of AdoHcy hydrolase seemed unlikely . This statement was based on the observation that reactivation was observed also under conditions of high concentrations of AdoHcy (obtained by the addition of homocysteine to the cell suspension) . Reactivation of AdoHcy hydrolase with a concomitant decrease in cellular level of AdoHcy could also be demonstrated with mouse plasmacytoma (MPC-11) cells and mouse fibroblasts (L-929) exposed to ara-A, but the reactivation process was far less pronounced than with hepatocytes.

Gan To Kagaku Ryoho, 1982 Jul, 9(7), 1285 - 92
{Application of the peripheral leukocyte cytofluorogram to diagnosis and treatment of hematopoietic tumors--with special reference to its effectiveness in malignant lymphoma}; Kamimura N et al.; In 1970 Adams and Kamentsky reported on a technique for classifying human peripheral leukocytes into three groups-lymphocytes, monocytes and granulocytes using supravital staining with a solution of acridine orange, and analyzing the resulting cell suspension with the Cytofluorogram . For the purpose of clinical application of flowcytometry, we investigated the clinical significance of this cytofluorogram of peripheral leukocytes . Cytofluorograms showed characteristic pattern in such hematopoietic malignancies as acute leukemias, chronic leukemias and malignant lymphomas, and turned out to be applicable to the diagnosis of these diseases . Especially in malignant lymphoma, cytofluorogram showed either normal or abnormal pattern . An abnormal cytofluorogram indicated poor prognosis, and an abnormal pattern of cytofluorogram became normalized after chemotherapy against lymphoma . Interestingly, those patients who showed in abnormal pattern relapsed in a short period, even when they were though diagnosed in remission . These data suggest that cytofluorograms are useful in diagnosing, predicting relapse and evaluating therapeutic effects for the treatment of malignant lymphoma . We hope that the analysis of cytofluorogram of peripheral leukocytes in lymphoma will provide a new benefit for further investigation of this disease.

Cancer Res, 1982 Jul, 42(7), 2537 - 43
Mode of action of polyethylene glycol 6000 in potentiating the in vitro generation of antitumor cytotoxicity by MOPC-315 tumor bearer spleen cells; Mokyr MB et al.; Some of the possible mechanisms by which polyethylene glycol (PEG) augments the ability of MOPC-315 tumor bearer spleen cells to mediate in vitro antitumor cytotoxicity were evaluated . The level of antitumor cytotoxicity obtained in 5-day cultures of tumor bearer spleen cell suspensions correlated inversely with the percentage of Trinitrophenol (TNP)-rosettable cells (presumably metastatic tumor cells) present in the spleen . The kinetics of decrease in the percentage of TNP-rosettable cells coincided with the appearance of antitumor cytotoxicity . In addition, PEG was shown to interfere with the ability of viable tumor cells to suppress the in vitro generation of antitumor cytotoxicity in normal spleen cells cultured with mitomycin C-treated tumor cells . However, the decrease in the content of TNP-rosettable cells and the concurrent increase in the level of antitumor cytotoxicity were not due to direct cytotoxic and/or cytostatic effects of PEG on tumor cells . Spleen cells cultured in the presence of PEG had an increased rate of {3H}thymidine incorporation and proliferation compared to spleen cells cultured in the absence of PEG . However, the PEG-induced decrease in the percentage of TNP-rosettable cells either preceded or occurred at the same time that the PEG-induced increase in spleen cell number was observed . Therefore, spleen cell proliferation can at best explain only partially the PEG-induced decrease in the content of TNP-rosettable cells, and other mechanisms for the decrease must be considered.

Horm Res, 1982 Jul-Aug, 16(4), 257 - 67
Glucagon and insulin secretion by dispersed islet cells: possible paracrine relationships; Dunbar JC et al.; The ability of dispersed islet cells in a perifusion system to secret glucagon and insulin in response to physiologic stimuli was investigated . Normal hamster islets were isolated by collagenase digestion and the cells dispersed by sequential digestion with collagenase and trypsin . Following a 50-min period of equilibrium in buffer with high glucose concentrations (5.0 mg/ml), glucagon secretion was stimulated by glucopenia and subsequently, inhibited by increasing the concentration of glucose . The responsiveness to glucose inhibition was significantly less in dispersed islet cells than in intact islets . However, the dispersed islet cells showed significantly greater response to arginine . Glucagon secretion by dispersed islet cells was stimulated to tolbutamide and epinephrine but somatostatin had no effect . Dispersed islet cell preparations did not augment insulin secretion in response to glucose but did secrete more insulin in response to arginine . Intact islets secreted insulin in response to glucose but not arginine . We conclude that A cells in cell suspension do not need direct contact or an intact intra-islet environment in order to respond to glucose, arginine, epinephrine, or tolbutamide but the extent of response may be influenced by paracrine effects . However, paracrine relationships may be important in determining the response of B cells to secretagogues.

J Cell Biol, 1982 Jul, 94(1), 193 - 200
Reexpression of blood group ABH antigens on the surface of human thyroid cells in culture; Khoury EL; Using indirect immunofluorescence (IFL) on viable human thyroid cultures, it has been shown that, although adult follicular cells do not express blood group ABH antigens in vivo, they invariably reexpress the corresponding antigens on the cell surface when cultured in monolayers, even for very short periods . The absence of blood group antigens on noncultured thyroid cells was confirmed by negative IFL on cell suspensions obtained after enzymatic digestion of the glands, whereas these antigens were readily demonstrable on cell suspensions obtained by trypsinization of established monolayers . The quantitative expression of ABH antigens on individual thyroid cells was variable and the cell-surface IFL pattern due to binding of blood group isoantibodies was different from that given by organ-specific thyroid autoantibodies on viable cultures . Reexpression of blood group antigens by cultured thyroid cells could not be related to the secretor status of the donors, the presence of a particular source of serum in the culture medium or cell division in vitro . After 2-3 wk in culture, thyroid cells became morphologically dedifferentiated and no longer displayed blood group antigens, though they still expressed cell-surface beta 2-microglobulin . Fibroblasts present in the primary thyroid cultures were invariably negative for ABH antigens . These results demonstrate that the surface antigenic repertoire of cultured human cells is not necessarily identical to that present on the same cells in vivo . Furthermore, the possibility that blood group natural isoantibodies bind to the cell surface must be taken into account in experiments in which cultured thyroid cells are exposed to human sera.

Int J Radiat Oncol Biol Phys, 1982 Jul, 8(7), 1193 - 201
Monoclonal antibodies: potential role in radiation therapy and oncology; Order SE; Specificity, which is a hallmark of the immune system, will be used in radiation oncology in both diagnosis and therapy through the application of radiolabelled monoclonal and polyclonal antibodies . Antigenic specificities, antibody preparations, and the tumor as a target for radiolabelled antibody is reviewed . Several clinical situations, i.e . single tumor cell suspensions, intraperitoneal single cells and masses, and solid tumors are reviewed in regard to both immune antibody targeting and specific differences between tumors in these regions . The concentration of tumor associated antigens is introductory to radiolabelled antibodies in diagnosis . In the radiation therapy of solid tumors, data regarding tumor dose, tumor effective half-life, varied antibody preparations, and the use of radiolabelled antibody as a method of tumor implantation is discussed using antiferritin 131I-IgG as a model as a model in hepatoma . The theoretical applications of monoclonal antibody integrated in cancer therapy are then presented as a new goal for future development.

J Steroid Biochem, 1982 Jul, 17(1), 11 - 6
Effect of corticosteroid binding proteins on the steroidogenic activity of bovine adrenocortical cell suspensions; Basset M et al.; The possible role of steroid binding proteins in the hormonal secretion process of a steroidogenic tissue was examined using bovine adrenocortical cell suspensions, either under basal conditions or in the presence of half-maximally active concentration (1 x 10(-9) M) of synthetic adrenocorticotropic hormone (ACTH) . Three types of plasma cortisol binding proteins were used, namely bovine serum albumine (BSA), purified transcortin (CBG) and purified anticortisol immunoglobulins (IgG) . When added to the incubation medium, CBG (at 1 x 10(-10) to 2 x 10(-9) M cortisol binding sites) and anticortisol IgG (at 4.8 x 10(-10) to 3 x 10(-9) M cortisol binding sites) did not influence either the basal nor the ACTH-stimulated net cortisol production of the cell preparations . Whereas crystallized and delipidated BSA showed also no effect, crude commercial BSA preparation (Cohn fraction V) exhibited an ACTH-like cofactor effect which resulted in a marked increase in the net cortisol production by stimulated cells . These observations might be explained by the presence in crude BSA of lipoprotein-cholesterol complexes, possibly acting as an extracellular source of cholesterol available for corticosteroidogenesis . It may be concluded that specific high affinity cortisol binding systems present outside adrenocortical steroidogenic cells do not influence their secretory activity under short term in vitro condition . In addition, it can be stressed that use of ill defined protein preparations (e.g . crude BSA) may lead to artifactual observations in the study of the differentiated functions of isolated steroidogenic cells.

Am J Clin Pathol, 1982 Jul, 78(1), 48 - 53
Spurious rapid infections mononucleosis test results in non-infectious mononucleosis sera . The role of high-titer horse agglutinins; Horwitz CA et al.; The present report-describes serologic data from five individuals without infectious mononucleosis (IM) who had very high levels of horse red blood cell agglutinins (1:896-1:7168) that were unusual in that they were minimally absorbed by either guinea pig kidney or beef red blood cell suspensions . Agglutination of sheep red blood cells was negative, and Epstein-Barr virus (EBV)-specific serodiagnostic tests revealed evidence for long past primary infections . In one of the five cases, serologic findings suggested a more than usually active EBV carrier state, as evident from persistently high anti-VCA (IgG) levels of 1:1280 and the continual presence of anti-R at 1:40 . Sera from all cases predictably interfered with several commercially available IM-specific rapid slide agglutination tests . The incidence of such findings was estimated to be no more than one per one thousand non-IM cases . In three patients adequately studied, the poorly absorbable horse agglutinins persisted in serum samples over periods of five to nine years . The significance of these findings is discussed.

J Exp Med, 1982 Jul 1, 156(1), 283 - 8
Idiotype-like determinants on human T lymphocytes alloactivated in mixed lymphocyte culture; Suciu-Foca N et al.; T cells alloactivated in 5-d MLC with an HLA-DR-different stimulator acquire the capacity of stimulating the autologous mixed lymphocyte response (AMLR) . We have demonstrated that activation of AMLR by allosensitized T cells is determined by the expression of the idiotype receptor for the stimulating HLA-DR alloantigen . This has been shown in experiments in which purified, OKT-3-positive T cell suspensions were first primed for 9 d with AMLR-activated T lymphoblasts, then tested in secondary AMLR with autologous lymphoblasts sensitized to various HLA-DR alloantigens . Accelerated memory responses were induced only by autologous lymphoblasts that had been sensitized against the same HLA-DR specificity as the primary AMLR stimulators . This response was not inhibited by a mouse monoclonal antibody recognizing Ia-like determinants, and was not triggered by human allogeneic resting peripheral blood lymphocytes . Thus, recognition of alloactivated T lymphoblasts in secondary AMLR seems to be specific for the idiotype-like determinants expressed by the autologous stimulators.

J Invest Dermatol, 1982 Jul, 79(1), 23 - 9
Pemphigoid, pemphigus and Pr antigens in adult human keratinocytes grown on nonviable substrates; Woodley D et al.; Isolated adult human keratinocytes were grown either on plastic coverslips or a nonviable basement membrane surface containing intact laminin, type IV and V collagens, and heparan sulfate proteoglycan and examined by indirect immunofluorescence for the expression of bullous pemphigoid, pemphigus and Prlh antigens . Initial cell suspensions had a mean of 23% and 30%, respectively, of bullous pemphigoid and Prlh positive staining cells, while those stained with pemphigus serum were usually negative (19 of 22 series) . Pemphigus antigen was expressed as intercellular staining between keratinocytes within 24 hr in both cultures on plastic and basement membrane . Likewise, Prlh antigen was expressed within 24 hr as a homogeneous cytoplasmic fluorescence leaving the basement membrane zone unstained . In contrast, pemphigoid antigen was expressed as a linear fluorescent band at the basement membrane zone between days 3 and 4 of culture . Systematic cell counts of bullous pemphigoid antigen positive cells from trypsin disrupted primary cultures made on plastic over time showed a nadir (8%) of positive cells in early cultures after which the percentage rapidly rose to a peak of 58% between days 14 and 21 of culture . In subcultures repeatedly disrupted at short intervals, the percentage of bullous pemphigoid positive cells remained low when compared to those interrupted and passaged over longer intervals . The percentage of bullous pemphigoid antigen bearing cells in culture over time is similar, but not identical, to the percentage of basal cells and is related to the age and known growth kinetics of the cultures system . Bullous pemphigoid, pemphigus and Prlh antigens are synthesized by the epidermal cell whether cultured on basement membrane or plastic.

Endocrinology, 1982 Jul, 111(1), 86 - 94
The possible importance of contact between pancreatic islet cells for the control of insulin release; Halban PA et al.; Insulin secretion was studied using adult rat intact islets, dissociated isolated islet cells, and isolated islet cells that had been allowed to reaggregate . The three preparations were maintained for 2 h in tisue culture medium at 37 C in order to allow for equilibration after the isolation procedures and to permit restoration of contacts between the reaggregated cells . The isolated cells appeared well preserved at the ultrastructural level . Evidence for intimate contacts between the reaggregated cells was demonstrated by the reappearance of gap junctions between adjacent cells . Basal insulin release (2.8 mM glucose) during 30 min was elevated in the isolated cell suspension but was restored to the level found in intact islets when isolated cells from the same population were reaggregated . The elevated basal release did not appear to be due to cell damage since there was a commensurate increase in the rate of insulin biosynthesis relative to intact islets . Although there was a marked stimulation of insulin release from the intact islets at 16.7 mM glucose, the response was smaller in the reaggregated cells and in the isolated cell suspension . All three preparations responded to elevated cAMP levels evoked by glucagon in the presence of 16.7 mM glucose . Similar results were obtained when insulin release was studied in various preparations derived from newborn rat pancreatic endocrine cells . Thus, basal insulin release was elevated in isolated cells, whereas cultures with cell contacts displayed lower basal insulin release . Taken together, the restoration of lower basal insulin release and the parallel appearance of contacts between the reaggregated cells suggest that these two phenomena are interrelated . Thus, cell contacts may be important in regulating basal insulin release . Whether such regulation is a consequence merely of cell association or of direct communication between cells remains to be established.

Biochim Biophys Acta, 1982 Jun 14, 688(2), 495 - 504
Comparative lipid analysis of purified plasma membranes and shed extracellular membrane vesicles from normal murine thymocytes and leukemic GRSL cells; Van Blitterswijk WJ et al.; The lipid fluidity in purified plasma membranes (PM) of murine leukemic GRSL cells, as measured by fluorescence polarization, is much higher than in PM of normal thymocytes . This was found to be due to relatively low contents of cholesterol and sphingomyelin and a high amount of unsaturated fatty acyl chains, especially linoleic acid, in the phospholipids . PM from GRSL cells contain markedly more phosphatidylethanolamine than those from thymocytes . For both GRSL cells and thymocytes the detailed lipid composition of isolated PM was compared with that of the corresponding shed extracellular membranes (ECM), which were isolated from the ascites fluid and from thymus cell suspensions, respectively . The somewhat decreased lipid fluidity of thymocyte ECM as compared to their PM, can be ascribed to the increased cholesterol/phospholipid molar ratio (0.88 vs . 0.74) . No other major differences were found between the lipid composition of these membranes . In contrast, significant differences were found between PM and ECM from GRSL cells . In this system a much lower lipid fluidity of the shed ECM was found, due to the much increased cholesterol/phospholipid molar ratio (3.5-fold) and sphingomyelin (9-fold) content, as compared to the PM . Further, the ECM contain relatively more lysophosphatidylethanolamine and less phosphatidylcholine and -inositol . ECM contain a higher amount of polyunsaturated fatty acids, especially in the phosphatidylethanolamine and lysophosphatidylethanolamine classes . On the other hand, the fatty acids of phosphatidylcholine and lysophosphatidylcholine are more saturated than in PM . In particular, ECM of GRSL cells contain less oleic and linoleic acid residues and more arachidonic acid and 22:polyunsaturated fatty acid residues than PM . The possible relevance of these differences with respect to the mechanism of shedding of vesicles from the cell surface, is discussed.

Biochim Biophys Acta, 1982 Jun 8, 720(3), 250 - 8
Carbon-13 nuclear magnetic resonance studies of acetate metabolism in intact cells of rhodopseudomonas sphaeroides; Nicolay K et al.; 13C-nuclear magnetic resonance was used to study the metabolism of {2-(13)C}acetate in suspensions of Rhodopseudomonas sphaeroides . In the dark, in logarithmic-phase cells the 13C label appeared first in butyrate C-2 and C-4 and subsequently in glutamate C-4 and succinate C-2 and C-3 . In the light, synthesis of poly(beta-hydroxybutyrate) (PHB) takes place . Butyrate synthesis seems to be independent of PHB synthesis or degradation activity . Starved, logarithmic-phase cells also show massive synthesis of PHB in the dark . Stationary-phase cells incorporate 13C predominantly into glutamate and succinate . No significant butyrate biosynthesis can be detected in the dark or during illumination . The incorporation of label in PHB is very slow in these cells and most probably originates from exchange of 12C for 13C into PHB . This might indicate slow turnover without net synthesis of the polymer occurring under these conditions . The results are discussed in relation to the redox state and the availability of metabolic energy for biosynthetic reactions in the dark and during illumination of cell suspensions of Rps . sphaeroides.

Immunology, 1982 Jun, 46(2), 321 - 8
Subcapsular thymic lymphoblasts expose receptors for soy bean lectin; Raedler A et al.; In the course of analysing thymic cellular constituents concerning the expression of lectin receptors and defined lymphocyte differentiation antigens, respectively, a subpopulation of thymic lymphocytes was detected exposing both peanut lectin (PNL) and soy bean lectin (SBL) receptors on their surfaces, while the majority of cortical thymic lymphocytes were found to be PNL positive, but SBL negative . This subset of SBL+PNL+thymic lymphocytes in the adult populates the area beneath the thymic capsule, forming a narrow rim only, while in perinatal thymus a corresponding subcapsular cell layer comprises almost one third of the thymic tissue and in early ontogenesis the vast majority of thymic lymphoid cells are PNL+SBL+ . By ultrastructural analysis of double labelled cell suspensions as well as of cells separated by affinity chromatography, it could be shown that PNL/SBL+ cells frequently exhibit morphological features of lymphoblastic cells . Thus this subcapsularly located subset seems to represent a compartment of proliferating immature cells, descended from bone marrow and foetal liver prethymic cells, respectively, and giving rise to PNL/SBL- cortical thymic lymphocytes.

J Anat, 1982 Jun, 134 (Pt 4), 729 - 40
Phagocytic lymphoid cells and transitional cells in the peritoneal cavity; Yaffe P et al.; In 71 mice, a study was made of the non-granular phagocytic cells in the peritoneal cavity, both after the intraperitoneal injection of India ink, and by incubation of the non-adherent cells with ink in vitro . Ink was taken up by a number of lymphoid cells, as well as by macrophages and monocytes, referred to jointly as macrophages . In addition, the presence of DNA-synthesizing cells was investigated by radio-autography after incubation of cell suspensions for 1 hour with tritiated thymidine . A small number of labelled macrophages was usually found in the normal peritoneal cavity, and also about 1% of labelled cells with the typical morphological features of the transitional cells seen in bone marrow . During 14 days after the intraperitoneal injection of ink, the combined population of lymphoid cells and macrophages showed an increase in the percentage of lymphoid cells, and a fall in the percentage of macrophages . The phagocytic lymphoid cells did not appear to develop into macrophages . It seems reasonable to assume that the haemopoietic stem cells known to be present in the peritoneal cavity are to be found among the transitional cells, some of which may also be macrophage or lymphocyte precursors.

Tsitologiia, 1982 Jun, 24(6), 710 - 5
{Separation of a rat thymocyte population in a ficoll-paque gradient . III . The optical characteristics of the fractions}; Sungurov AIu et al.; Values of the own ultraviolet fluorescence (UVF) intensity, of the relative UVF intensity (i/R2 proportion, where i--UVF-intensity, R--cell radius), of optical densities of cell suspensions beyond their light absorption band as well as of rates of light scattering by cells at the 90 degrees angle to the beam direction were determined for rat thymocyte fractions obtained by sedimentation of the total thymocyte population on the Ficoll-Paque layer (rho=1.077) . It has been shown that the upper (less dense) thymic fractions have the greatest values of parameters assayed when compared with the bottom cells . Interrelations of the data obtained with those on the cell sizes, cytoprotein contents and internal cellular structure are discussed.

Tsitologiia, 1982 Jun, 24(6), 699 - 703
{Separation of a rat thymocyte population in a ficoll-paque gradient . I . Relation of cell buoyant density to biological parameters}; Zherbin EA et al.; Rat thymic cells were fractioned in the one-step Ficoll-Paque gradient . The three fractions obtained differed from one another with respect to average cell sizes, protein and DNA contents per cell . The light fraction cells revealed the maximum incorporation of labeled precursors into their nucleic acids, a relatively low hydrocortisone sensitivity and a diminished adhesion capacity . Following preincubation of thymus cell suspension and its gamma-irradiation, the number of the heavy fraction cells increased . The enrichment of the heavy fraction of cells with cortical thymocytes, including terminal ones, is supposed.

In Vitro, 1982 Jun, 18(6), 510 - 4
Culture substrate dependence of mouse fibroblasts survival at 4 degrees C; Matsumura T et al.; When L-929 mouse fibroblasts grown in Eagle's medium (MEM) supplemented with 10% fetal bovine serum (FBS) were stored in a monodisperse suspension at 4 degrees C, the viability decreased rapidly from the beginning of storage . The viability in this study was determined by counting electronically the number of cells with the capacity to attach to glass substrate and with the membrane boundary resistant to a proteolytic digestion . When, however, the dissociated cells were preincubated briefly at 37 degrees C, and subsequently stored at 4 degrees C as they were attaching on a glass substrate, the rapid loss of viability could be reduced effectively . A biphasic survival profile consisting of an initial phase of slowly decreasing viability and the subsequent phase of rapidly decreasing viability were than observed . The rapid viability loss occurred not only when the cell suspension was prepared by mechanical dislodging but also after trypsinization or dispase treatment . Such viability loss was also observed when the dissociated cells were not stored at 4 degrees C directly but preincubated in a monodisperse suspension at 37 degrees C in a siliconized plate and then stored at 4 degrees C . The above results show that the rapid loss of viability is associated closely with the fact that the cells were not attached to the substrate but in suspension.

Hoppe Seylers Z Physiol Chem, 1982 Jun, 363(6), 551 - 62
The participation of the Shemin and C5 pathways in 5-aminolaevulinate and chlorophyll formation in higher plants and facultative photosynthetic bacteria; Klein O et al.; Chlorophyll and bacteriochlorophyll formation from 14C-labelled precursors was studied during the illumination of etiolated maize leaves excised from dark-grown seedlings and in cell suspensions of respiring, dark-, aerobically-grown Rhodopseudomonas spheroides adapting to the photosynthetic state in the light under anaerobic conditions . It was found that 1-14C-labelled glutamate and 2-oxoglutarate were incorporated into the tetrapyrrole moieties of chlorophyll and bacteriochlorophyll . This suggests that the C5 pathway of tetrapyrrole biosynthesis operates in both Zea mays and R . spheroides since in the alternative Shemin pathway the label would have been lost as 14CO2 during the formation of succinyl-CoA prior to the condensation with glycine to form 5-aminolaevulinate . It was also found that 5(-14)C-labelled glutamate and 2-oxoglutarate were incorporated into these chlorophylls which is consistent with the operation of both the C5 and Shemin pathways . That the Shemin pathway is also involved was confirmed by the incorporation of {2(-14)C}glycine into both chlorophylls . None of these substrates were incorporated into the phytol moieties of either plant or bacterial chlorophyll or into the carotenoids . However, when {1(-14)C}acetate was added to greening maize leaves not only the tetrapyrrole and phytol moieties were labelled but also the carotenoids: the labelling of these lipids is consistent with their formation from acetate via the isopentenyl pyrophosphate pathway . By comparing the incorporation of {1(-14)C}2-oxoglutarate with that of {5(-14)C}2-oxoglutarate the approximate relative contribution of each pathway to chlorophyll biosynthesis was determined . In maize leaves both pathways contributed almost equally but in R . spheroides the contribution by the Shemin and C5 pathways was 90 and 10%, respectively.

Cancer, 1982 Jun 1, 49(11), 2300 - 4
Effects of antibodies to choriogonadotropin in malignant growth . I . Rat 3230 AC mammary adenocarcinoma; Kellen JA et al.; To determine whether the in vivo production of antibodies against choriogonadotropin (CG) could modify the relationship between host and malignant growth, the effects of preimmunization with a conjugate of the beta-subunit of CG and tetanus toxoid (CG beta-tt) on the development and growth of lung metastasis following I.V . injection of R 3230 rat mammary adenocarcinoma cells into Fischer 344 female rats have been investigated . This neoplasm has been previously demonstrated to synthesize CG-like material . A test group of 70 animals and two matched control groups of 15 animals each were used . One of the control groups was untreated and the other received tetanus toxoid only . All the control animals had multiple lung foci of neoplastic cells ten days after seeding, as has been previously observed, and CG antibodies were undetectable . In contrast, a significant titer of anti-CG was found in all preimmunized animals . At the same time, the pretreated animals rarely had a few small neoplastic nodes in the entire lung sections at the standard 8--10 days post-seeding . The protective effects of preimmunization with CG beta-tt were further demonstrated by the animals living 20 days post-seeding, the absence of lung pathology in the ones killed thereafter, and by six animals that were left alive and have not shown any deterioration for more than six months after the administration of the cell suspension.

Transplantation, 1982 Jun, 33(6), 593 - 8
Migration patterns of lymphocytes in untreated and immunologically manipulated recipients of organ allografts; Kupiec-Weglinski JW et al.; We have investigated the migration patterns of normal LEW rat splenic lymphocytes (SLs) radiolabeled in vitro with {2-3H}adenosine and adoptively transferred i.v . into LEW hosts bearing LBNF1 heterotopic cardiac allografts . Twenty-four hours after cell transfer, the animals were killed and the radioactivity of all lymphoid and nonlymphoid tissues measured in a beta counter . The following experimental groups were studied: group 1, untreated recipients acutely rejecting their grafts at 7 days after transplantation; group 2, actively and passively enhanced recipients bearing long-term surviving grafts, at 7, 14 to 18, and 25 days; group 3, cyclosporin A-treated recipients; and group 4, B rats, each bearing indefinitely surviving grafts, at 7 and 20 to 30 days . In group 1, 28% of recoverable activity was found in spleen and 22% in mesenteric and peripheral lymph nodes . In animals with well functioning grafts of groups 2 and 3, SLs accumulated in both organs equally (25 to 27%) . In animals in group 4, SLs migrated primarily to lymph nodes (30%) and away from spleen (20%) . Sequestration in nonlymphoid tissues of animals experiencing graft rejection was higher than those with prolonged or indefinitely surviving hearts . By using mouse anti-rat monoclonal antibodies, the quantitative relationship between T cell subpopulations in transferred cell suspensions and in lymphoid organs of grafted hosts was also assessed . Lymphocyte migration patterns are influenced dramatically by the immunological status of recipients of vascularized organ allografts.

J Exp Med, 1982 Jun 1, 155(6), 1597 - 609
Formation of Charcot-Leyden crystals by human basophils; Ackerman SJ et al.; Charcot-Leyden crystals (CLC) are currently believed to be unique to the eosinophil and a hallmark of active eosinophilic inflammation or proliferation . The distinctiveness of the CLC to the eosinophil was questioned in 1965 by Archer and Blackwood (9), but their demonstration of CLC formation in basophils was ignored and later dismissed (1) as being the result of eosinophil contamination of basophil-enriched cell suspensions . We reexamined this question and showed that basophils obtained from the peripheral blood of normal individuals form CLC and that basophils contain a protein that is immunochemically indistinguishable from eosinophil CLC protein . These conclusions are based upon the findings that (a) crystal formation in basophils was demonstrated by specific histochemical staining of crystal-containing cells in highly enriched basophil suspensions prepared by fluorescence-activated cell sorter (FACS) purification of surface IgE-positive cells, (b) that enrichment for surface IgE-positive cells (primarily basophils) by the FACS also enriched for cells staining positively by immunofluorescence for eosinophil CLC protein, and (c) that CLC protein was measured by radioimmunoassay in cell extracts prepared from purified basophil suspensions containing 97-99% basophils and absolutely no contaminating eosinophils . These basophil extracts contained a protein immunochemically indistinguishable from eosinophil CLC protein . Based upon these findings, the CLC or the protein comprising the crystal (lysophospholipase) can no longer be considered as distinctive to the eosinophil . We must now consider the possibility that the presence of CLC in tissues, sputum, or stool may also represent basophil involvement in disease processes.

J Immunol, 1982 Jun, 128(6), 2543 - 6
Role of genes in murine major histocompatibility complex (MHC) in the ontogeny of B cells with restricted capacity for cooperation with adherent cells in the production of antibody to thymus-dependent (TD) and thymus-independent type 2 (TI-II) antigens; Gorczynski RM et al.; We have examined the MHC restriction that exists for cooperation between B lymphocytes and antigen-pulsed accessory cells for antibody responses in tissue culture, using B cells prepared from spleen cell suspensions of progeny born to (B10 x B10.BR)F1 female mice mated with B10 males . At early times post-birth (3 to 6 wk) progeny animals types as H2b contained a population of splenic B cells restricted to MHC antigens expressed on B10.BR macrophages; in contrast, B cells from H2b animals born to the incross (B10 x B10) were as expected, restricted to cooperate only with B10 macrophages . This unusual restriction pattern waned with age such that by 20 wk of age considerably fewer of the B10 progeny born in the backcross mating exhibited this behavior . We interpret these data as evidence that the early B cell repertoire is to some degree 'driven' by exposure to those MHC antigens (in the maternal placental circulation) encountered in utero . Post-birth, continued renewal of the mature B cell pool occurs concomitant with a diversification (and selection) of the B cell repertoire as a result of experience with antigens lacking this maternally imparted genetic information.

J Natl Cancer Inst, 1982 Jun, 68(6), 945 - 9
Effect of enzymatic disaggregation on proliferation of human tumor cells in soft agar; Hamburger AW et al.; Cell suspensions were prepared by either mechanical or enzymatic disaggregation methods from biopsy specimens from 54 patients with various tumors . The biologic activities of cells derived from the two suspensions were then examined . Biopsy specimens of solid tumors were minced, and one-half of each specimen was further processed by being teased with needles, whereas the other half was exposed to a combination of collagenase, hyaluronidase, and DNase . The enzymatic disaggregation method yielded fewer cells per gram of tissue than the mechanical method . However, the percentage of dye-excluding cells was increased by the enzymatic procedure in 93% of the cases . Cells obtained by enzymatic means also had higher cloning efficiencies than those obtained by mincing . The histologic types of cells present in the initial cell suspensions were the same for cells obtained by the enzymatic or mechanical disaggregation methods . The number of colonies obtained was linearly related to the number of cells plated in both cases . The tritiated thymidine suicide indices (estimates of the percentage of cells in the S-phase of the cell cycle) were the same for the two cell populations obtained by the two methods . The results indicate that cells obtained from solid tumors by enzymatic dissociation methods did not differ significantly from cells obtained by the more conventional mechanical techniques . However, cell viabilities and cloning efficiencies were significantly improved by the enzymatic technique.

Am J Clin Oncol, 1982 Jun, 5(3), 321 - 8
The modification of melphalan toxicity in tumor bearing mice by s-2-(3-aminopropylamino)- ethylphosphorothioic acid (WR 2721); Millar JL et al.; The toxicity of melphalan in mice was reduced by the injection of S-2-(3-aminopropylamino)-ethylphosphorothioic acid (WR2721) . This was seen in terms of reduced toxicity to the stem cells of the bone marrow and intestinal epithelium as well as improved animal survival . Using human melanoma xenografts and growth delay as an end-point, it was demonstrated that WR2721 did not protect this tumor from melphalan . With radio-labelled WR2721, it was shown that WR2721 was rapidly cleared from the blood and actively accumulated by all normal tissues except the CNS . Intact human tumor xenografts and Lewis lung tumors were less able to accumulate WR2721 than normal tissues, but in vitro studies showed that tissue fragments or single cell suspensions of tumors were as efficient as liver fragments or bone marrow cells in accumulating the drug . The rapid clearance of WR2721 and poor vascularity of the intact tumors were thought to be responsible for the differential uptake and protection of normal tissues by WR2721.

J Invest Dermatol, 1982 Jun, 78(6), 452 - 5
Enrichment of epidermal Langerhans cells: studies with a monolayer technique and flow cytometry sorting; Scheynius A et al.; Langerhans cells were enriched from epidermal cell suspensions by a monolayer technique based on the association of Langerhans cells with solid matrix-bound anti-Ia antibodies or by flow cytometry sorting of fluorescein-isothiocyanate labeled anti-Ia reactive cells . The monolayer technique gave a moderate enrichment (15-37%) whereas considerably higher purity (73-87%) was obtained by flow cytometry sorting . The identity of the enriched anti-Ia reactive cells as Langerhans cells was established by histochemical techniques or electron microscopy . The monolayer-enriched Langerhans cells could function as stimulating cells in the mixed leukocyte culture reaction and as antigen-presenting cells.

Cell Biophys, 1982 Jun-Sep, 4(2-3), 77 - 95
Philothermal and chemotactic locomotion of leukocytes . Method and results; Fu TK et al.; A newly developed technique for quantitating the locomotion of polymorphonuclear leukocyte (PMN) populations in temperature gradients has revealed that PMNs accumulate toward higher temperatures . The experiments yield measurements of the numbers of cells that adhere to glass and migrate from a cell suspension through a liquid/gel meniscus into a glass/agarose interface, and of their spatial distribution at subsequent time intervals . Cell locomotion was investigated as a function of the magnitude, sign, and temporal variation of the temperature gradient, the cell concentration in the source suspension, and the presence or absence of chemoattractant gradients . It was found (1) that a temperature gradient stimulates crossing of the meniscus toward higher temperatures, (2) that only a portion of the cells reverses direction of locomotion in response to reversal of the temperature gradient after the cells have traversed the meniscus, and (3) that the distribution of cells in the migration space depends on cell concentration, suggesting that the dynamics of PMN locomotion depend on cell-cell interactions.

Int J Lepr Other Mycobact Dis, 1982 Jun, 50(2), 183 - 92
A fluorescent staining procedure for determining the viability of mycobacterial cells; Kvach JT et al.; A fluorescent staining procedure has been developed which rapidly, accurately, and economically measures the viability of mycobacterial cells . M . smegmatis and M . phlei have served as prototype organisms to establish conditions which ensure optimal staining . The staining method incorporates the use of the fatty acid ester fluorescein diacetate (FDA) and ethidium bromide (EB) . Non-polar, non-fluorescent FDA enters live cells where it is enzymatically hydrolyzed by acetylesterase to polar, fluorescent fluorescein which rapidly accumulates in the cytoplasm . These cells appear green when viewed under incident ultraviolet illumination . Ethidium bromide enters dead cells and intercalates between the bases of DNA molecules . These cells appear red-orange under UV illumination . Live cells are, therefore, identified on the basis of possessing acetylesterase and their ability to exclude EB; whereas dead cells are identified on the basis of lacking acetylesterase and their inability to exclude EB . The feasibility of applying the staining procedure of M . leprae has been investigated and the results are encouraging . Our findings reveal that armadillo-derived M . leprae possess acetylesterase and, therefore, stain green . M . leprae cell suspensions exposed to adverse physico-chemical conditions give rise to high proportions of red-stained cells as would be expected if the cells are being killed . An alternative means of determining the viability of M . leprae appears to be feasible.

J Neurocytol, 1982 Jun, 11(3), 351 - 62
Immunocytochemical study of the appearance of P2 in developing rat peripheral nerve: comparison with other myelin components; Winter J et al.; Using indirect immunofluorescence on both dissociated cell cultures and frozen sections from rat sciatic nerve, dorsal root ganglion and superior cervical ganglion, we have examined the development and distribution of the peripheral myelin protein P2 . It first appears in development in sciatic nerve and dorsal root ganglion on the first day of birth, at about the same time as P0 and P1 . Like galactocerebroside, P0 and P1, P2 disappears gradually from dissociated Schwann cells in culture . In adult sciatic nerve, and dorsal and ventral roots, it shows an uneven distribution and is absent from some myelinated axons . In electron micrographs the onset of myelination in the sciatic nerve occurs between the day of birth (day 0) and the first day after birth (day 1) . Immunofluorescence studies on freshly dissociated cell suspensions, frozen sections and dissociated cell cultures at these early time points indicate that the myelin glycolipids galactocerebroside and sulphatide are present on the surface of many schwann cells at least one day before myelination starts while the myelin proteins P0, P1 and P2 are not detected until myelination begins . This suggests that the early appearance of galactocerebroside and sulphatide is an important step preceding the formation of compact myelin.

Infect Immun, 1982 Jun, 36(3), 1123 - 7
Effect of cyclosporin A on murine natural killer cells; Gui X et al.; In mice, cyclosporin A decreased the natural killer cell-enhancing effect of two interferon inducers, infective murine cytomegalovirus and nonreplicating Newcastle disease virus . It also inhibited murine cytomegalovirus replication at doses greater than 20 mg/kg, but it did not significantly inhibit interferon induction by Newcastle disease virus . In cell culture, cyclosporin A had no direct effect on the natural killer activity of spleen mononuclear cells derived from normal or murine cytomegalovirus-infected animals . However, at 50 micrograms/ml it significantly reduced the ability of interferon to enhance the natural killer activity of normal spleen cell suspensions . The inhibitory effect of cyclosporin A on natural killer cell activity in infected mice may be partly explained by its ability to block the action of interferon.

J Immunol, 1982 May, 128(5), 1975 - 8
Oscillation of cell surface charge of human peripheral blood lymphocytes after stimulation with concanavalin A; Uzgiris EE et al.; Lymphocytes isolated from human peripheral blood were incubated with concanavalin A (Con A), and relative changes in cell surface charge were monitored by measuring cell electrophoretic mobility . Kinetic studies showed a time lag of 2.5 hr between exposure to Con A and development of a significant change in cell surface charge . The possible involvement of de novo synthesis of a factor was suggested by the following observations: 1) addition of the protein synthesis inhibitor, puromycin, abrogated the change in electrophoretic mobility; 2) supernatants obtained from cells incubated with Con A caused a mobility change in a separate group of lymphocytes not previously incubated with this mitogen; 3) supernatants obtained from cell suspensions treated with puromycin were inactive . The mobility response showed an oscillation of membrane charge ranging from a 1- to 1.5-hr period, which eventually disappeared after 8 hr at 37 degrees C . Cells at the minimum of the mobility oscillation were refractory to addition of active supernatants . This suggests the involvement at the lymphocyte surface of some cyclic metabolic process, or perhaps formation of receptor-factor complexes followed by endocytosis of these complexes with subsequent regeneration of the receptors on the cell surface.

Biochem J, 1982 May 1, 203(2), 493 - 504
Phenolic components of the primary cell wall . Feruloylated disaccharides of D-galactose and L-arabinose from spinach polysaccharide; Fry SC; 1 . Cell walls from rapidly growing cell suspension cultures of Spinacia oleracea L . contained ferulic acid and p-coumaric acid esterified with a water-insoluble polymer . 2 . Prolonged treatment with trypsin did not release may feruloyl esters from dearabinofuranosylated cell walls, and the polymer was also insoluble in phenol/acetic acid/water (2:1:1, w/v/v) . 3 . Treatment of the cell walls with the fungal hydrolase preparation "Driselase' did liberate low-Mr feruloyl esters . The major esters were 4-O-(6-O-feruloyl-beta-D-galactopyranosyl)-D-galactose and 3?-O-feruloyl-alpha-L-arabinopyranosyl)-L-arabinose . These two esters accounted for about 60% of the cell-wall ferulate . 4 . It is concluded that the feruloylation of cell-wall polymers is not a random process, but occurs at very specific sites, probably on the arabinogalactan component of pectin . 5 . The possible role of such phenolic substituents in cell-wall architecture and growth is discussed.

Somatic Cell Genet, 1982 May, 8(3), 329 - 45
Isolation of UV-sensitive mutants of mouse L5178Y cells by a cell suspension spotting method; Shiomi T et al.; We have isolated 56 UV-sensitive mutant clones from a mouse L51 T/t line of L5178Y cells by a cell suspension spotting method . Five mutants have also been isolated from L51 T/t and L5178Y cells by the method reported by Thompson and coworkers (22) . We divided the mutants into two groups, "highly sensitive" and "moderately sensitive" mutants, according to their sensitivity to UV irradiation . Fifty-eight mutants were highly sensitive and three were moderately sensitive to UV . The reconstruction experiments indicate that more than 90% of highly sensitive mutants were recovered by the cell suspension spotting method . Frequencies of recovered mutants highly sensitive to UV increased with increasing dose of mutagens . Recovered mutant frequency reached 10(-2) after treatment with 1.5 micrograms/ml of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) (survival 0.2%) . Eight UV-sensitive mutants were divided into four complementation groups . These mutants were 2-6 times more sensitive to UV than parental L51 T/t cells in terms of D37 (dose required to reduce survival to 37%) . Four representative UV-sensitive mutants which are classified into different complementation groups were examined for their sensitivity to killing by UV, 4-nitroquinoline-1-oxide (4NQO), mitomycin C (MMC), X-rays, and MNNG . All four classes of mutants were found to be cross-sensitive to UV, 4NQO, and MMC, but not sensitive to X-rays and MNNG.

Differentiation, 1982 May, 21(3), 175 - 82
Mechanisms of adhesion among cells of the early chick blastoderm: role of the beta-D-galactoside-binding lectin in the adhesion of extraembryonic endoderm cells; Milos N et al.; Cells from the extraembryonic endoderm of the gastrulating chick embryo contain a beta-D-galactoside-binding lectin inhibited by thiodigalactoside (TDG) . TDG inhibits the aggregation of freshly prepared cells . In these fresh cell suspensions, adhesion is also inhibited when purified lectin is added to the aggregation assay . If these cells are incubated at 22 degrees C their adhesion decreases . Associated with this is an increase in lectin activity in the cell supernatants . In these incubated cells aggregation is stimulated by TDG and desialyzed fetuin . These data suggest that the lectin may have a role to play in cellular adhesion . Under some experimental conditions extraembryonic endoderm cells from rosettes with trypsinized glutaraldehyde-fixed rabbit erythrocytes . This phenomenon is inhibited, to a certain extent, by TDG.

Parasite Immunol, 1982 May, 4(3), 171 - 85
Immunity to coccidia in chickens: adoptive transfer with peripheral blood lymphocytes and spleen cells; Rose ME et al.; Suspensions of cells prepared from the caecal tonsils and spleen, and the peripheral blood lymphocytes of chickens immune to Eimeria maxima, were tested for their ability to transfer resistance to syngeneic recipients . The intravenous injection of approximately 6 x 10(8) spleen cells or peripheral blood lymphocytes caused a significant reduction of oocyst production by the challenged recipients, in comparison with controls which were uninjected or given cells from birds susceptible to E . maxima . Peripheral blood lymphocytes appeared to be most effective when obtained 10-15 days after a primary, or 3-10 days after a secondary inoculation of oocysts . The peripheral blood lymphocytes which participate in the early response to challenge of immune birds were not found to be protective . When given intraperitoneally, greater numbers of spleen cells were required to reduce oocyst production, and small numbers of caecal tonsil cells were ineffective . The ability of the various cell suspensions to transfer antibody and cell-mediated responses was monitored with 'marker' antigens . There was some indication that both types of response were involved in protection.

Exp Hematol, 1982 May, 10(5), 459 - 66
In vivo studies on hemopoietic stem cells and target cells for Friend virus infection in vitro; Seidel HJ et al.; Friend virus replicating target cells have been characterized by in vitro infection of bone marrow cells from DBA/2 mice after multiple injections of hydroxyurea (HU), after bleeding and/or hypertransfusion and after busulfan treatment . The concentration of CFUS, BFUE and CFUE in the cell suspension was correlated with the number of infectious centers (IC) induced after infection . HU treated mice were also infected in vivo . The results demonstrate that there is almost no target cell 4 h after the last of 4 HU injections . The number of IC induced could correlate with the numbers of CFUS in the HU experiment, but this possibility can be rejected based on the busulfan experiments . All results, including those after bleeding and/or hypertransfusion are compatible with a target cell between BFUE and CFUE, as suggested by other in vitro infection methods and also in vivo experiments.

Biull Eksp Biol Med, 1982 May, 93(5), 102 - 4
{Separation of mouse bone marrow cells by preparative electrophoresis}; Kotel'nikov VM et al.; Free-flow electrophoresis was used to separate mouse bone marrow cells . From five to eight fractions with an amount of cells sufficient for morphofunctional analysis could be obtained in different experiments . Granulocyte precursors were found in fractions with low electrophoretic mobility (EPM), whereas erythrocyte precursors in fractions with high EPM . CFUs were detected both in low and in high EPM fractions . In individual fractions the concentration of these cells was 4 times greater than in the initial cell suspension.

Biofizika, 1982 May-Jun, 27(3), 498 - 500
{Fe2+ ion effect on Ehrlich ascites tumor cell conductivity and ion permeability}; Polivoda BI et al.; It is shown that incubation of EAT cells in the presence of Fe2+ ions caused the threshold decrease of active and capacitance constituents of cell suspension impedance and rate of ion efflux from cells to the isotonic sucrose media, and decrease extent rose with an increase of Fe2+ ion concentration and temperature . A discrepancy between the results described and experimental model system data is discussed.

J Exp Med, 1982 May 1, 155(5), 1491 - 500
Novel differentiation antigens on late erythrocytic progenitor cells detected by alloantisera made between mouse strains congenic at the Fv-2 locus; Vaithilingam DS et al.; We have investigated the activities of alloantisera produced in B6 (C57BL/6) and B6.S strain mice reciprocally immunized with unwashed bone marrow and spleen cell suspensions from their respective Fv-2 congenic partner strains, B6.S and B6 . These antisera inhibited the formation of colonies by the late erythrocytic progenitors (CFU-E) in plasma cultures seeded with unwashed bone marrow or spleen cells; washed cells were unaffected . Erythropoietic burst formation by the early progenitors (BFU-E) was not significantly inhibited by the antisera, whether the cells were washed or unwashed . We conclude (a) that the congenic antisera are capable of recognizing alloantigens controlled by alleles of Fv-2 or of a closely linked gene locus on chromosome 9; (b) that these alloantigens are situated on the surface of erythrocytic progenitor cells and can be removed by washing; and (c) that the expression of the alloantigens on these cells is influenced by their stage of differentiation.

Tsitologiia, 1982 May, 24(5), 507 - 21
{Information value of light scattering parameters in cell study (a review of the literature)}; Bezrukova AG et al.; Different methods of light scattering measurements for cell suspension and cells in flow are reviewed . The intensity of light scattered at different angles suggests information about the cell size, the nucleus to cell diameter ratio, the thickness of surface membrane, the changes in the internal cell structure state, and about the heterogeneity of cell population . The efficiency of information obtained from the turbidity measurements depends on the geometry of the spectrophotometer . Measurements made with diaphragms allow us to get information about the mean size of biological particles, their form, refractive index, concentration etc . The main advantages of light scattering methods are their high sensitivity, a possibility to use the same species before and after the experiment, and a compatibility with other methods.

J Clin Pathol, 1982 May, 35(5), 492 - 5
Ratio of blood and marrow-derived cells in bone marrow transplantation; Kay H et al.; The content of the cell suspensions used in bone marrow transplantation can vary by at least twelvefold in the ratio of bone marrow-derived to blood-derived cells . The ratio is somewhat higher in young donors but is similar for male and female donors and is not closely correlated with the incidence of graft v host reactions.

Blood, 1982 May, 59(5), 913 - 22
The immunologic phenotyping of bone marrow biopsies and aspirates: frozen section techniques; Wood GS et al.; Techniques were developed for the preparation of frozen sections from undecalcified bone marrow biopsy cores and bone marrow aspirates . Selected indirect antibody and biotin-avidin detection systems, employing immunoperoxidase or immunofluorescent labels, were studied to determine which were best suited for bone marrow frozen section immunohistopathology . Methods employing murine hybridoma monoclonal antibodies illustrate the immunophenotyping of representative lymphoid neoplasms, involving bone marrow . The results of immunohistologic staining were comparable to those of fluorescence-activated cell sorter (FACS) analysis of cell suspensions . The advantages and limitations of immunohistologic techniques are discussed as they relate to immunophenotypic studies previously feasible only with bone marrow cell suspensions . Frozen section immunohistologic techniques serve as useful adjuncts to the conventional evaluation of bone marrow aspirates and biopsies.

Acta Pathol Jpn, 1982 May, 32(3), 513 - 26
Pagetoid reticulosis (Woringer-Kolopp disease) . An ultrastructural and immunocytological study; Takahashi H et al.; Histopathological, immunocytological and ultrastructural observations are reported in the first case of pagetoid reticulosis (Woringer-Kolopp disease) in Japan . The patient was a 61-year-old woman with multiple skin lesions running a chronic and apparently benign clinical course . Histology of the skin biopsies revealed typical pagetoid appearance of the epidermis due to intraepidermal infiltration of abnormal cells . Ultrastructural investigation showed that the intraepidermal abnormal cells were classified into mycosis fungoides cells, Sezary cells, lymphoblast-like cells, and large blastoid cells and that the mycosis fungoides cells were a major cell population . Intermediate or transitional cells were found between these cells and large blastoid cells were mostly situated in the basal cell layer . By the rosetting assays of the free cell suspensions prepared from the epidermis of the biopsied skin lesions, 93% of the suspended cells were positive for spontaneous rosette formation with sheep erythrocytes . The immunoperoxidase technique demonstrated no cytoplasmic immunoglobulins in almost all the intraepidermal abnormal cells . These results indicate that the intraepidermal abnormal cells are T-lymphocytes . Thus, it is concluded that the present case is a cutaneous T-cell lymphoma of low-grade malignancy showing a prominent epidermotropism . This case is the first description of the disease in Japan.

Exp Hematol, 1982 May, 10(5), 467 - 71
HLA antigens expressed on human pluripotent hemopoietic precursors in vitro (CFUMIX); Ohe Y et al.; The human major histocompatibility antigen HLA, plays an important role in transplantation . To ascertain the expression of the antigens of HLA-A and -B loci on the surface of human pluripotent hemopoietic progenitors (CFUMIX), the cytotoxic effect of specific anti-HLA sera was examined . Anti-HLA sera and complement formed fewer colonies from CFUMIX in a culture medium than the controls which were treated with autologous serum . In our experiments, the HLA antisera killed also T cells that carry the surface antigen . Then the influences of T cells on mixed colony formation were examined . No reduction of colony formation was observed from the cell suspension in which T cells were depleted by treatment of monoclonal antibody OKT3 . These results indicated that CFUMIX expressed HLA antigens on their surface.

J Bacteriol, 1982 May, 150(2), 966 - 8
Obligate methylotrophy: evaluation of dimethyl ether as a C1 compound; Meyers AJ Jr; The suitability of dimethyl ether as a C1 compound was examined with the obligate methylobacterium Methylococcus capsulatus (Texas) . The ether did not support growth and was not formed during growth on methane; it was an inhibitor of growth and oxidation of methane and a poor oxidation substrate for cell suspensions . NADH stimulation of methane, but not dimethyl ether, oxidation occurred in cell extracts.

J Cell Physiol, 1982 May, 111(2), 187 - 94
Changes in phenotypic expression in embryonic and adult cells treated with 5-azacytidine; Taylor SM et al.; We have previously shown that 5-azacytidine (5-Aza-CR) induced the formation of biochemically differentiated myotubes, adipocytes, and chondrocytes in the mouse embryo cell line, C3H/10T1/2CL8 (10T1/2), and that the induction of the muscle phenotype was cell cycle specific . Here we show that the adipocyte phenotype is also induced maximally in cells treated during S phase . During this period, the minimum treatment time required for the subsequent formation of myotubes was 5 min and the number of myotubes formed was dependent on treatment time . The incorporation of 14C-5-Aza-CR into DNA during the cell cycle, however, was not enhanced during early S phase, suggesting that incorporation of 5-Aza-CR into specific DNA sequences synthesized during early S phase may be required for the expression of the new phenotypes . Single cells, obtained by plating cell suspensions into 16 mm wells at limiting dilution, were treated with 5-Aza-CR during S phase . The resulting clones showed a high frequency of phenotypic conversion, indicating that 5-Aza-CR did not act via a selective mechanism, and several of the clones were capable of expressing more than one phenotype . The cells required more than 2 division cycles after treatment with the analog for the expression of the muscle phenotype and the capacity to differentiate was retained for long periods of time in the absence of cell division . The adult mouse line, CVP3SC6, differentiated into functional striated muscle cells following treatment with 5-Aza-RE . The analog also caused oncogenic transformation in the adult line at the same concentration that was effective at inducing myogenic expression.

Arch Ophthalmol, 1982 May, 100(5), 822 - 5
Centrifugal cytology of ocular fluids; Stulting RD et al.; Centrifugal cytology is a new technique for the preparation of ocular fluids for cytologic examination . It differs from conventional cytocentrifuge techniques, since fixation is carried out simultaneously with centrifugation . It avoids air-drying artifacts and can be applied to small (50 microL) or large (200 mL or more) sample volumes and dilute cell suspensions, such as might be obtained by anterior chamber paracentesis, vitreous aspiration, or vitrectomy . The resultant preparation is a permanent, stained, well-preserved, flattened, homogeneous cell dispersion suitable for detailed cytologic evaluation.

Biochemistry, 1982 Apr 27, 21(9), 2146 - 50
Asymmetry of lipid dynamics in human erythrocyte membranes studied with permanent fluorophores; Schachter D et al.; The fluorescence anisotropy and mean excited-state lifetime of 1,6-diphenyl-1,3,5-hexatriene, 12-(9-anthroyloxy)stearate, 2-(9-anthroyloxy)stearate, and pyrenedecanoic acid in the membranes of intact human erythrocytes, lysate suspensions, and ghost membranes were compared . The excited-state lifetime of each lipid fluorophore, estimated by single photon counting, is significantly shorter in the intact erythrocytes as compared to the lysates, owing to nonradiative energy transfer from the lipid fluorophore donors in the membrane to heme acceptors at the endothelial surface of the intact cell . The fluorescence observed in intact cell suspensions is thus weighted in favor of outer leaflet fluorophores, and estimates of the fluorescence anisotropy by steady-state fluorescence polarization indicate that all four fluorescent probes experience greater motional freedom in the outer as compared to the inner membrane leaflet . The results are in accord with prior studies of impermeant pyrene derivatives, which also indicate that the outer leaflet lipids have greater motional freedom.

Neurosci Lett, 1982 Apr 26, 29(3), 297 - 302
Cultivation of immature astrocytes of mouse cerebellum in a serum-free, hormonally defined medium . Appearance of the mature astrocyte phenotype after addition of serum; Fischer G et al.; A hormonally defined culture medium is described which supports survival and proliferation of astroglia from primary cultures of early postnatal mouse cerebellum . This medium consists of bovine serum albumin, insulin, transferrin, selenium, hyaluronic acid, protease inhibitor aprotinin, and epidermal growth factor . Trypsin-dissociated single cerebellar cell suspensions are plated in this medium on poly-l-coated glass coverslips and maintained for two weeks before subcultivation . After subcultivation into defined medium more than 99% of all cells are vimentin-positive and fibronectin- and almost completely glial fibrillary acid (GFA) protein-negative, indicating that these cells are less mature astrocytes . After replacement of defined medium by culture medium containing 10% horse serum, expression of GFA protein is detectable in addition to vimentin by indirect immunofluorescence.

Cancer, 1982 Apr 15, 49(8), 1605 - 12
Immunological characterization of non-Hodgkin's lymphoma: enrichment of neoplastic cells from lymphoid tissues and blood; Bom-Van Noorloos AA et al.; Cell suspensions prepared from lymph nodes, spleen or peripheral blood of patients with non-Hodgkin's lymphoma (NHL) often contain a high percentage of residual nonmalignant cellular elements . By E-rosette sedimentation, it was possible to enrich such suspensions from patients with various types of lymphoma for malignant cells . In patients with a B- or non-B/non-T-cell lymphoma, the neoplastic cells were found in the non-T fraction . The capacity to respond to stimulation by various stimuli was then confined to the T-cell fraction, which contained the residual normal T-cells . In patients with T-cell lymphomas, in which the malignant cells had retained the capacity to form E-rosettes, lymphoma cells were found in the T fraction . These cells usually did not respond to mitogenic stimuli . Using this separation method, small proportions of neoplastic cells could be identified in mixed cell populations . Thus, in the blood from nine out of 23 lymphoma patients without abnormalities in routine blood tests, a population of abnormal cells was detected after cell separation . This included a monoclonal B-cell population in the blood of four patients, a questionably monoclonal B-cell population in the blood of two patients and in increased non-B/non-T cell population in the blood of three patients.

Acta Pharmacol Toxicol (Copenh), 1982 Apr, 50(4), 310 - 5
Uptake of 51Cr-chromate by human erythrocytes-a role of glutathione; Aaseth J et al.; Hexavalent chromium (Cr-VI), as Na2CrO4 in an aqueous solution, was reduced rapidly ot the trivalent form (Cr-III) in the presence of glutathione, GSH (0.3-3.0 mM) . Such GSH-dependent reduction Cr-VI can take place in the cytosolic space of Cr-VI-exposed cells, since GSH is found in reactive concentrations in this compartment . The reduction makes chromium essentially impermeable through the cell membrane, explaining the observation that Cr-VI, when added to red cell suspensions, is bound quantitatively intracellularly after a few hours . Diethylmaleate conjugation of the SH-group of the intracellular GSH preventing the oxidation to GSSG, lowered the chromium-uptake significantly, showing that reduced GSH plays a role for the chromium binding . In healthy red cells chromium is partially bound to haemoglobin and partially to small molecular weight substances, probably in the trivalent form . This intracellular chromium cannot be removed to the extracellular space by addition of chelating agents as long as the cell membrane is intact.

J Invest Dermatol, 1982 Apr, 78(4), 319 - 22
Separation of human skin cells by velocity sedimentation into functionally distinct fractions; Morhenn VB et al.; The epidermis consists of a heterogeneous population of cells including Langerhans cells, Merkel cells, melanocytes, and keratinocytes in various stages of differentiation . The current study was undertaken to determine if skin cell suspensions can be separated into morphologically and/or functionally distinct fractions . Skin cells were suspended by trypsinization and separated into multiple fractions by velocity sedimentation . Certain fractions reproducibly stimulated proliferation of allogeneic lymphocytes in the skin cell lymphocyte reaction, whereas other fractions, containing larger cells, supported growth of keratinocyte colonies in cell cultures . These results indicate that stimulation in the skin cell lymphocyte reaction and growth of keratinocyte colonies are mediated by distinct cells, separable by velocity sedimentation.

Immunology, 1982 Apr, 45(4), 769 - 74
Isolation and functional characteristics of adherent phagocytic cells from mouse Peyer's patches; MacDonald TT et al.; Attempts were made to isolate adherent phagocytic cells (macrophages) from mouse Peyer's patch cell suspensions . Cell suspensions prepared by teasing apart the Peyer's patches contained no adherent phagocytic cells . However, if Peyer's patch fragments were treated with collagenase to disrupt the tissue matrix, cells prepared in this way contained a subpopulation of adherent phagocytic cells . These cells comprised only 0.1-0.2% of the total nucleated cell population of the Peyer's patch . Similar cells could also be isolated from the Peyer's patches of germ-free mice, but as judged by their ability to ingest opsonized erythrocytes, these cells were less activated than cells from the Peyer's patches of normal mice . Adherent cells from the Peyer's patches of normal mice could present antigen (ovalbumin) to T cells, and Peyer's patches cell suspensions containing adherent cells could be stimulated in vitro to produce an anti-sheep red blood cell plaque-forming cell response in the absence of 2-mercaptoethanol . These studies show that although the frequency of phagocytic adherent cells is extremely low in Peyer's patches, these cells have functions consistent with that of adherent cells in other lymphoid tissues.

J Immunol, 1982 Apr, 128(4), 1535 - 40
Macrophages as effector cells of protective immunity in murine schistosomiasis . II . Killing of newly transformed schistosomula in vitro by macrophages activated as a consequence of Schistosoma mansoni infection; James SL et al.; Plastic adherent peritoneal cells from Schistosoma mansoni-infected mice have previously been shown to exhibit nonspecific tumoricidal activity in vitro . In this report we show that these same cell populations kill significant numbers of skin-stage schistosomula in vitro in the absence of added antibody . Larval killing by these activated cells could be enhanced by the use of suspension rather than monolayer cultures and by addition of heat-inactivated immune mouse serum to the cultures . Adherence of cells to schistosomula was also enhanced under the same conditions, suggesting that cell binding to the larvae might be critical in the development or expression of microbicidal activity . In support of this hypothesis, the same level of enhancement of cell binding and larval damage was observed upon substitution of concanavalin A for immune mouse serum . Killing of schistosomula appeared to be mediated solely by activated macrophages in the peritoneal cell suspensions from S . mansoni-infected mice, because partially purified preparations of eosinophils were virtually inactive in these assays . Likewise, inflammatory macrophages from uninfected mice were unable to kill schistosomula under the same conditions, emphasizing the importance of activation in the development of killing capability . The finding that macrophages activated as a consequence of S . mansoni infection are able to kill larval schistosomes in vitro suggests that these cells may play a role in concomitant immunity to schistosomiasis in vivo.

J Histochem Cytochem, 1982 Apr, 30(4), 359 - 63
The kinetics and homogeneity of endocytosis of a receptor-bound ligand in a heterogeneous cell population studied by flow cytofluorometry; Metezeau P et al.; A method is described to study quantitatively and rapidly the kinetics of endocytosis of a receptor-bound ligand by cells of various subclasses within a heterogeneous cell population . The time course of the internalization of the bound ligand is followed by labeling, with a fluorescent antibody, the ligand molecules exposed on the cell surface at various times of the endocytosis process, and by measuring with a flow cytofluorometer the fluorescence of a large number of individual cell within the total cell population . A convenient mathematical treatment of the data is proposed for analyzing the experimental results . This method is applied to the kinetic study of the endocytosis of rabbit antibodies (anti-mouse immunoglobulin) by B-lymphocytes within a mouse spleen cell suspension.

Cancer Res, 1982 Apr, 42(4), 1289 - 95
Initial rate kinetics of the transport of adenosine and 4-amino-7-(beta-D-ribofuranosyl)pyrrolo{2,3-d}pyrimidine (tubercidin) in cultured cells; Harley ER et al.; A procedure is described for determining early time courses of nucleoside uptake by cultured cells in suspension . Replicate samples of cell suspensions were exposed to medium containing 3H-nucleosides for brief intervals (sec) ended by addition of nitrobenzylthioinosine, a potent inhibitor of nucleoside transport that terminated nucleoside uptake virtually instantaneously . Time courses of nucleoside uptake were constructed from the cellular content of nucleoside acquired by the replicate samples during graded intervals of exposure to the labeled permeant . Such time courses were definitive of cellular uptake of nucleosides during the first few sec of exposure to permeant and yielded initial rates of uptake of adenosine and 4-amino-7-(beta-d-ribofuranosyl)pyrrolo{2,3-d}pyrimidine (tubercidin) . Defining initial rates of nucleoside uptake as rates of inward transport, relationships between transport rates and extracellular concentrations of these permeants were evaluated in HeLa cells and in two cultured lines of mouse lymphoma L5178Y cells that differ in their abilities to phosphorylate adenosine and tubercidin . Transport rates for these permeants were similar in the two L5178Y cell types and were saturable in the 3 cell lines with Km values between 14 and 38 microM . Adenosine and tubercidin were mutually competitive permeants in L5178Y cells, indicating that they are substrates for the same transport mechanism.

Exp Hematol, 1982 Apr, 10(4), 352 - 9
In vitro and in vivo specificities of purified thymic factors inhibiting graft-versus-host reaction; Blazsek I et al.; A calf thymic factor has been prepared by extraction in distilled water, followed by selective precipitation with ethanol and ammonium sulfate . The non-dialysable fraction obtained by this method was further purified by a preparative electrofocusing technique which allows the isolation of two fractions which can specifically inhibit, in vitro, without toxicity or species specificity, spontaneous DNA synthesis in hemopoietic cell suspensions . When injected in vivo in mice, 3 days after immunization with SRBC, both these fractions exhibited immunosuppressive properties . In contrast, they were without any effect on a thymus-independent antibody response to TNP-LPS, whatever the day of administration . Still more important, in vitro pretreatment of lymphoid cells by the purified fractions inhibits their ability to elicit a graft-versus-host reaction when injected to histo-incompatible recipients.

Mutat Res, 1982 Apr, 104(1-3), 159 - 63
A method for preparing chromosomes from peripheral blood of the mouse; Barren PR et al.; A modified Triman et al . (1975) procedure for preparing metaphase chromosomes from mouse peripheral blood is described . Modifications include: isolation of white blood cells using a ficoll-hypaque gradient; seeding cultures with a known concentration of leucocytes such that the PHA/cell concentration can be controlled for optimum stimulation through 72 h with no media change; and visual monitoring of cell growth to determine when there should be sufficient numbers of dividing cells for harvest and thus eliminate unnecessary harvest of non-productive cultures . We have found this modified procedure to be highly reproducible with the final 0.5 ml fixed cell suspension yielding 50-75 quality metaphase spreads per drop.

Transplantation, 1982 Apr, 33(4), 400 - 2
Effect of blood transfusions on canine renal allograft survival; van der Linden CJ et al.; In this study significantly prolonged canine renal allograft survival has been demonstrated after transfusion of 100 ml of third-party whole blood given peroperatively . Peroperative transfusions of third-party leukocyte-free blood or pure lymphocyte cell suspensions did not influence graft survival . Furthermore, no improvement in graft survival has been found after a peroperative transfusion of irradiated whole blood (2500 rad) . These data suggest that delayed graft rejection after blood transfusions can only be expected after the administration of whole blood . The role of competent lymphocytes in whole blood is questionable, since a transfusion or irradiated whole blood in combination with nonirradiated lymphocytes did not lead to prolonged graft survival . Immunosuppression of the recipient directly after transfusion seems to be essential to induce the beneficial effect of blood transfusions . This has been demonstrated for a transfusion of whole blood 14 days before transplantation . A single transfusion of 100 ml of whole blood 14 days before transplantation could effectively prolong graft survival if immunosuppression with azathioprine and prednisone was started on the day of transfusion . No improvement in graft survival has been found with such a transfusion if preoperative immunosuppression has been omitted.

Scand J Haematol, 1982 Apr, 28(4), 293 - 305
Distribution of T-cell subsets identified by monoclonal antibodies in cell suspensions from lymph node biopsies of human B-cell lymphomas; Kvaloy S et al.; The distribution of T-cell subsets has been examined in cell suspensions from lymph node biopsies from 37 non-Hodgkin lymphomas (NHL) of B-cell origin and 8 cases of Hodgkin's disease (HD) . T-inducer/helper cells (T4+ cells) and T-suppressor/cytotoxic cells (T8+ cells) were identified by the monoclonal antibodies OKT4 and OKT8, respectively . Compared with reactive lymph nodes the T-cell subset distribution was aberrant in 18/37 (48.6%) of the B-cell lymphomas . The T-suppressor subset was dominating cells in 14/37 (37.8%) and the T-helper subset in 4/37 (10.8%) of the cases examined . In patients with leukaemic disease, a concordant T-cell subset distribution in lymph nodes and PBL was observed in 5 out of 6 cases . HLA-DR antigen was examined in a selected number of cases with a heteroantiserum . In general, the T-cells from neoplastic tissues expressed an increased percentage of HLA-DR antigen . This was most pronounced in cases with a dominance of T8+ cells, in which the majority of T8+ cells expressed HLA-DR antigen . These findings suggest that an increased proportion of activated T-suppressor cells are present in a proportion of B-cell lymphomas . Possible clinical and biological implications are discussed.

Proc Natl Acad Sci U S A, 1982 Apr, 79(8), 2722 - 5
Pluripotential hemopoietic stem cells in adult mouse brain; Bartlett PF; Single cell suspensions of adult mouse brain were shown to contain large numbers of pluripotential hemopoietic stem cells as detected by the ability to form hemopoietic colonies in the spleens of irradiated hosts . These colony forming unit, spleen (CFU-s) cells derived from brain gave rise to colonies identical in morphology and histology to those of bone marrow-derived CFU-s . The average number of CFU-s obtained per 10(5) dissociated adult brain cells was 14, whereas other adult tissues such as lung, kidney, heart, and thymus contained insignificant CFU-s levels when tested . As the level of CFU-s in adult blood is less than 1 per 10(6) nucleated cells, blood contamination does not contribute to the high levels found in adult brain . Individual spleen colonies isolated from irradiated CBA (H-2k) recipients injected with (BALB/c x CBA)F1 (H-2d x H-2k) brain cells were shown by immunofluorescence to contain cells bearing surface H-2d molecules, thus indicating that the colonies arose from the brain cell inoculum and were not endogenously derived . The surface phenotype of brain- and bone marrow-derived CFU-s was found to differ in that brain CFU-s could be inhibited by prior incubation with a monoclonal antibrain antibody B2A2, whereas bone marrow CFU-s were not . Further differences were found between brain and bone marrow CFU-s in the congenitally anemic Wf/Wf mice . These mice were shown to have a very few CFU-s in the adult bone marrow, whereas the brain contained normal adult levels . The large number of hemopoietic stem cells in the brain may indicate an essential requirement for the continual generation of cells such as microglia or phagocytic cells, without the disruption of the blood-brain barrier.

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