J Neurol Sci, 1981 Mar, 49(3), 335 - 40 Epileptic discharges produced in monkeys by injection of spleen cells from rabbits immunised with monkey brain; Baltz ML et al.; Cell suspension from the spleen of rabbits immunised with monkey brain gave rise to epileptic discharges in the monkey when injected intracortically, except in the case of one injection from a rabbit not recently given a booster treatment . In contrast, cell suspensions from the spleen of unimmunised rabbits, or of a rabbit immunised with a non-brain material, in all cases failed to be effective . The epileptic discharges began about 2 weeks after injection, were variable in frequency, and lasted throughout the period of recording. J Invest Dermatol, 1981 Mar, 76(3), 190 - 2 Some biochemical properties of pemphigoid antigen bound to the surface of dissociated epidermal basal cells; Grekin PM et al.; Bullous pemphigoid antigen (BP Ag) is a cell surface marker of epidermal basal cells . The functional role of this molecule is unknown . Epidermal cell suspensions obtained by trypsinization of skin show a population of epidermal basal cells with a polar rim of antigen as demonstrated by indirect immunofluorescence technique . This study shows that treatment of these cells suspensions with a variety of proteolytic and glycosidic enzymes failed to remove the antigen from these basal cells . BP Ag was also stable upon incubation with distilled water, Triton X-100, PBS, and 1 M NaCl . Treatment of epidermal basal cells with 2 N NaSCN, 1% periodic acid, and 4 M urea, as well as acidic pH or 56 degrees C temperature, abolished the reactivity of these cells with BP antibodies. Scand J Haematol, 1981 Mar, 26(3), 221 - 34 Spontaneous (H)-thymidine uptake in histological subgroups of human B-cell lymphomas; Kvaloy S et al.; Spontaneous (3H)-TdR incorporation was studied in cell suspensions from 64 patients with monoclonal B-cell neoplasias . Among various incubation periods a long-term (20 h) assay with (3H)-TdR had the greatest discriminatory power versus non-neoplastic lymph node cell suspensions . The (3H)-TdR uptake correlated positively with cell volume, nuclear volume, and cells in S + G + M phase of the cell cycle . A high degree of heterogeneity with regard to (3H)-TdR incorporation was found within several histological groups of the Kiel classification, especially in low-grade malignant lymphomas of centroblastic/centrocytic origin and in the lymphoplasmacytoid groups . In highly malignant lymphomas (3H)-TdR uptake was statistically, significantly higher than in lymphomas of low-grade malignancy . Patients with localized disease (stages I and II) and those with "B'-symptoms showed increased incorporation of radioactive thymidine as compared to patients with disseminated mined by histopathology and spontaneous uptake of (3H)-TdR. Undersea Biomed Res, 1981 Mar, 8(1), 59 - 62 Method for separation of cells and suspending medium at high pressure; Goldinger JM et al.; We describe a pressure chamber designed for measuring fluxes and equilibrium distributions of substances between cells in suspension and the extracellular medium at pressures up to 400 ATA . The pressure chamber contains a filtration apparatus capable of taking four separate samples of cell-free medium from a cell suspension at timed intervals, without necessity of decompression . In addition, a high pressure injection system makes possible the addition to a cell suspension of precise quantities of agents such as ionophores or metabolic inhibitors, for observations of their effects on cell functions at pressure . The chamber of filtration apparatus functioned successfully in preliminary experiments designed to measure the equilibrium distribution of 36Cl- in human erythrocytes at high pressure. J Microsc, 1981 Mar, 121(Pt 3), 273 - 82 The design and use of a simple device for rapid quench-freezing of biological samples; Handley DA et al.; The detailed design of a simple device for rapid quench-freezing of biological samples under reproducible conditions is presented . With spring-augmented descent, sample immersion velocity of 10 m s-1 into a cryogenic liquid is achieved . Biological samples, loaded in Balzers planchets, Denton holders, or a newly designed 'titanium envelope', are suitable for rapid-freezing with this device . Using 4 micrometers titanium foil, light weight (1 mg) streamlined holders can easily be made to enclose cell suspensions or tissue samples . The foil envelope is designed for efficient heat dissipation while protecting the sample from possible impact or flow distortions occurring from spring-augmented immersion . Human erythrocytes, quench-frozen in the titanium envelope, were prepared for electron microscopy by the freeze-substitution technique . Two opposing 25--30 micrometers surface zones were frozen in the apparent absence of ice . The extended depth of cryofixation is attributed to the advantages of thin foil in the titanium envelope design and the use of rapid-immersion technique. Br J Haematol, 1981 Mar, 47(3), 345 - 52 Inhibitor of in vitro neutrophil migration in sera of children with homozygous sickle cell gene during pain crisis; Akenzua GI et al.; There is conflicting evidence for a causal relationship between infection and haematological crisis of sickle cell disease . To find out whether changes in leucotaxis occur during pain crisis, in-vitro neutrophil migration was determined in 38 children with Hb SS during steady state and during pain crisis . Migrations of neutrophils of sickle cell patients was 29 +/ 12 microns in steady state and 27.5 +/- 10.5 microns during pain crisis . These rates were comparable to migration of neutrophils of control children with normal haemoglobin of 34 +/- 9.6 microns . However, with addition of autologous serum to the cell suspension, neutrophil migration of patients in pain crisis was significantly retarded (16 +/- 13 microns) as compared to those in steady state (26 +/- 10.2 microns) and control children (28.7 +/- 10 microns) . Sera of children in pain crisis also inhibited migration of neutrophils of healthy adults with normal Hb . Pooled normal plasma reversed inhibitory action of pain crisis serum on autologous and homologous neutrophil migration; but pain crisis plasma did not . Chemotactic effect of sera of Hb SS children in steady state or pain crisis and control children on neutrophils of eight adults with normal Hb were similar and comparable to that of pooled normal serum . Thus, children with sickle cell disease develop chemotactic inhibitor(s) in their circulation during pain crisis . They may lead to defective leucotaxis and enhanced susceptibility to infection. Eur J Immunol, 1981 Mar, 11(3), 221 - 8 Different macrophage populations distinguished by means of fluorescent polysaccharides . Recognition and properties of marginal-zone macrophages; Humphrey JH et al.; Fluorescent conjugates of hydroxyethyl (OEt) starch of Ficoll are selectively ingested and retained in vivo by spleen marginal-zone (MZ) and lymph node marginal-sinus macrophages of mice, whereas similar conjugates of type 3 pneumococcal capsular polysaccharide (SIII) are generally retained by macrophages (Kupffer cells, histiocytes, macrophages of spleen, lymph nodes, bone marrow and peritoneal cavity) . MZ and other macrophages are readily identifiable by fluorescence after injection in vivo of OEt starch and SIII labeled with tetraethylrhodamine isothiocyanate and fluorescein isothiocyanate . Collagenase digestion was required for recovery of intact MZ macrophages from spleen in single cell suspensions and for maximum yields of other macrophages . MZ macrophages are larger and morphologically distinct from other macrophages, but resemble them in respect of EA, EAC receptors and acid phosphates and nonspecific esterase content and are equally radio-resistant . The appear normal in CBA/N nude and in beige mice . Freshly isolated MZ macrophages in suspension have adherent lymphocytes, dispersible by EDTA treatment, with B but not T cell markers . It is suggested that selective adherence to MZ macrophages is a factor in determining B cell traffic . MZ macrophages did not have demonstrable surface I-A or I-EC antigens . Only 4--8% of other spleen macrophages freshly isolated by collagenase treatment expressed I-A in the same preparation, whereas 35% of other cells (lymphocytes and blasts) reacted with monoclonal anti-I-A and anti-I-EC . After adherence to glass or plastic, 40% or more red-pulp, but not MZ macrophages, became I-A-positive . When taken from mice recently restimulated with sheep erythrocytes, half the red-pulp macrophages expressed I-A even before adherence . The relation of MZ to other macrophages is not known . However, their properties are consistent with the demonstrated ability of thymus-independent antigens selectively taken up by these cells to elicit long-lasting IgM antibody responses by direct interaction with B cells . The unexpected observation that only a small proportion of spleen macrophages freshly isolated from unstimulated mice had detectable surface I-A, but that this proportion was much increased after attachment to plastic, is discussed in relation to the possibility that macrophages do not express surface Ia antigens unless they have been stimulated. Eur J Immunol, 1981 Mar, 11(3), 200 - 6 Immunofluorescence studies on the expression of VH a allotypes by pre-B and B cells of homozygous and heterozygous rabbits; Gathings WE et al.; Fluorochrome-conjugated antibodies specific for C mu determinants and VH a allotypes were used to examine pre-B cells and B lymphocytes for expression of these markers . The majority of mu+ bone marrow pre-B cells were shown to express a allotypic determinants in conjunction with C mu . Both pre-B and B cells from a2 a3 heterozygous rabbits showed allelic exclusion of these allotypes . Pre-B cells expressing the a2 or a3 specificities appeared to be generated in approximately equal numbers in heterozygotes, while B lymphocytes expressing a3 appeared to undergo preferential clonal expansion very early in development . It was also observed that rabbit bone marrow and blood contained a population of myeloid cells which, in a2 a3 heterozygotes, stained for both a2 and a3 determinants . The frequencies of this cell type, which exhibited bright immunofluorescence staining for a allotypes, could not be reduced by prolonged incubation at 37 degrees C but were readily reduced after cell suspensions were treated with low pH buffer . It is concluded that these myeloid cells bear high avidity Fc receptors for serum immunoglobulin and may be the culprits in studies which have found production of two a or b allotypes by individual B lymphocytes. Am J Vet Res, 1981 Mar, 42(3), 483 - 6 Fractionation of Marek's disease virus-induced lymphoma by velocity sedimentation and association of infectivity with cellular fractions with and without tumor antigen expression; Sharma JM; Cell suspensions of lymphomas induced by Marek's disease (MD) virus were fractionated by sedimentation at unit gravity on a continuous gradient of bovine fetal serum . Cells in various fractions were examined for MD tumor-associated surface antigen (MATSA) by indirect immunofluorescence, using specific antibody, and for viral infectivity by cocultivating fractionated cells with permissive monolayer cells of duck embryo fibroblasts . Most MATSA-bearing cells in the lymphomas sedimented at a sedimentation velocity of greater than 3.0 mm/hour, whereas smaller, slow-sedimentating cells were generally devoid of MATSA expression . Viral infectivity was associated with MATSA-bearing and MATSA-lacking fractions . Within the limits of the experimental procedures used, this observation provided evidence that presence of MD virus genome in lymphocytes doses not result in concurrent expression of detectable MATSA and that MATSA likely represents another stage of interaction between MD virus and certain lymphocytes. Hypertension, 1981 Mar-Apr, 3(2), 157 - 67 Responses of juxtaglomerular cell suspensions to various stimuli; Khayat A et al.; Cell suspensions were prepared from rat renal cortical tissue by dispersion with 0.1% collagenase . Unit gravity sedimentation in a 1%-4% Ficoll gradient resulted in a single-cell suspension enriched in juxtaglomerular (JG) cells . Both the cellular renin activity and the amount of renin released into the supernatant increased with time when the suspensions were incubated for 1 hour at 37 degrees C in tissue culture medium . These cells responded to epinephrine and norepinephrine by increasing both synthesis and release of renin . The response was blocked by timolol but not by phenoxybenzamine . Cell suspensions prepared in the same manner but using 0.25% trypsin as the dispersing enzyme neither synthesized nor released renin into the tissue culture medium when similarly incubated . Trypsin-dispersed cells did not respond to catecholamine stimulation . Renin synthesis and release in collagenase-dispersed JG cells were unaltered by changes in Na, K, or Ca ion concentrations . Angiotensin II inhibited release, while saline extracts of clipped kidney from renal hypertensive rats stimulated renin release by these cells. Int J Lepr Other Mycobact Dis, 1981 Mar, 49(1), 49 - 56 Occurrence of gamma-glutamyl transpeptidase activity in several mycobacteria including Mycobacterium leprae; Shetty KT et al.; gamma-Glutamyl transpeptidase (gamma-GT) activity, which catalyzes the transfer of the "gamma-glutamyl" group of gamma-glutamyl compounds to several dipeptide and amino acid acceptors, was found to be present in several mycobacteria, including M . leprae, both in cell suspensions and in cell-free sonicates . Glycyl D-amino acids were active as acceptors, particularly glycyl-D-alanine and alpha, epsilon-diaminopimelic acid, among the amino acids . Two mycobacterial isolates obtained from biopsy material of lepromatous patients also exhibited similar enzyme activity . The need for further work to delineate the possible role of gamma-GT in mycobacterial metabolism is strongly indicated. Tumori, 1981 Feb 28, 67(1), 1 - 7 Cell and cell-free transplantation experiments with a mouse reticulum cell sarcoma; Covelli V et al.; Quantitative cellular assays of the transplantability of 19 spontaneous reticulum cell sarcomas (RCS) of the BC3F1 mouse were carried out by injecting serial dilutions of monodispersed cells into syngeneic hosts through the intravenous, intracranial and subcutaneous routes . A clear relationship was found between the size of the inoculum and the incidence of tumour takes by all routes, but even for inocula of 5 million cells only 13 tumors grew in one or more of the transplanted recipients . The intravenous route of injection was found to be the most effective, both in terms of the absolute number of tumor takes and the number of cells necessary to produce a given level of takes . The survival of the animals injected intravenously was related to the number of cells received in that earlier deaths occurred with the most concentrated cell suspensions . Attempts to transmit the tumor by cell-free extracts injected into newborn or 1-month-old syngeneic hosts failed to substantiate the possible presence of a specific leukemogenic agent. Int J Cancer, 1981 Feb 15, 27(2), 243 - 8 Systemic and in situ natural killer and suppressor cell activities in mice bearing progressively growing murine sarcoma-virus-induced tumors; Gerson J et al.; The activity of systemic and in situ natural killer (NK) cells was measured in 3- to 5-week-old CBA mice with progressively growing murine sarcoma virus (MSV)-induced tumors . Splenic NK activity was depressed in tumor-bearing mice . NK activity was quite low or not detectable in unfractionated cell suspensions from the tumors 10-12 days after inoculation of virus, but activity was observed upon depletion of phagocytic and/or adherent cells . To determine whether this in situ inhibition of NK activity was correlated with the presence of previously described suppressor cells, subpopulations of cells from the tumors were tested for their ability to suppress various responses of normal spleen cells: NK activity, lymphoproliferative responses to concanavalin A, and proliferation-independent production of macrophage inhibition factor . A similar pattern of suppressor activity was seen in all of the assays, indicating the presence of pleiotropic suppressor cells in progressively growing MSV-induced tumors. J Lab Clin Med, 1981 Feb, 97(2), 179 - 86 The nature of the prostaglandin-producing mononuclear cell in human peripheral blood; Bankhurst AD et al.; The purpose of this investigation was to identify the human peripheral mononuclear cell responsible for PGE production . The prostaglandin-producing cell had the following characteristics: (1) was glass adherent; (2) was removed by techniques that effectively removed phagocytic cells (carbonyl iron); and (3) was not an E-rosette cell . Furthermore, the peripheral mononuclear cells that contain cytoplasmic PGE by indirect immunofluorescence also ingest latex particles . B and T lymphoid cell lines did not produce large quantities of PGE . Mononuclear cells separated on discontinuous BSA gradients varied greatly in their ability to produce prostaglandin (low density > high density) . These data constitute strong evidence that the majority of PGE produced in peripheral mononuclear cell suspensions emanates from a population of low-density monocytes. Cancer Res, 1981 Feb, 41(2), 433 - 7 Differences in cell density associated with differences in lung-colonizing ability of B16 melanoma cells; Baniyash M et al.; The B16 melanoma-derived low lung-colonizing variant B16-F1 and the high lung-colonizing variant B16-F10 retained their differential lung-colonizing abilities throughout at least 35 serial s.c . transplant generations . The majority of the cells originating from solid B16-F1 tumors had a higher density than did cells originating from solid B16-F10 tumors . Cell suspensions of unselected solid B16 melanomas contained two major subpopulations differing in their cell density . The subpopulation with the lower cell density was more efficient in lung tumor colony formation, following i.v . administration, than was the high-density subpopulation . Cloned tumors from low-density B16 cells were more efficient in lung colony formation than were cloned tumors from high-density cells. J Neurosci, 1981 Feb, 1(2), 204 - 17 Separation of cell types from embryonic chicken and rat spinal cord: characterization of motoneuron-enriched fractions; Schnaar RI et al.; Single cell suspensions prepared from embryonic chick or rat spinal cords were separated into morphologically and functionally distinct subpopulation based on their buoyant densities The lightest fraction (F-1) was highly enriched for cells containing the enzyme choline acetyltransferase (CAT), a marker for developing motoneurons . The morphology biochemistry, and in vitro development of this and other spinal cord cell fractions isolated by the outlined procedure were investigated . Spinal cords, dissected from 6-day chick or 12-day rat embryos, were dissociated with trypsin and applied to iso-osmotic metrizamide density gradients . After brief centrifugation, biochemical analysis revealed that cholinergic cells migrated to lower densities than other spinal cord cells . The use of discontinuous density gradients allowed rapid and simple isolation of three fractions of viable cells (designated F-1 to F-3, lowest to highest density) . Characterization of chicken and rat embryo cell fractions gave similar results . The cells in Fraction 1 were large with prominent nuclei and nucleoli, while those in F-2 and F-3 were distinctly smaller . Fraction 1 was highly enriched for cholinergic cells . The CAT specific activity (CAT/cell) was increased 400% in Fraction 1 compared to unfractionated cells, while CAT specific activity in F-2 and F-3 was reduced to 25% and less than 4% that of unfractionated cells, respectively . The recovery of cholinergic cells using this procedure was much better than with other published procedures; greater than half the spinal cord CAT activity was routinely recovered in the enriched fraction . The cholinergic-enriched cells (F-1) were unique in their in vitro growth characteristics . All fractions had neuronal cells, while non-neuronal cells were distributed primarily in F-3, fewer in F-2, and were essentially absent from F-1 . Neurons in F-2 and F-3 remained viable under a variety of conditions, most of which were not supportive of F-1 cell survival . The cholinergic-enriched F-1 cells survived and developed only in the presence of muscle cells or in muscle-conditioned medium on highly adhesive substrata . Large, multipolar neurons predominated under these conditions . The method described provides a means of characterizing the factors involved in the development of distinct populations of cells from the embryonic spinal cord. Endocrinol Jpn, 1981 Feb, 28(1), 95 - 100 A trial for the improvement of the in vitro bioassay for human luteinizing hormones; Suginami H et al.; In order to improve the sensitivity and the reliability of the in vitro bioassay for human luteinizing hormone, various strains of male mice at various chronological ages were sacrificed to obtained dispersed testes interstitial cell preparations . They consisted of DDN, DDY, DBA, C57, C3H, ICR, and BAL/C at 5, 7 and 9 weeks of age . The C57 Black strain, especially at 7 and 9 weeks of age, gave the most satisfactory standard dilution curve with a sensitivity of less than 12.5 microIU/tube, with a wide assayable range and with a parallelism to the sample dilution curve . Improvement of the assay is expected with the use of the C57 Black strain as the source of the cell suspension. Clin Sci (Lond), 1981 Feb, 60(2), 185 - 90 No functional difference of the two iron-binding sites of human transferrin in vitro; Van der Heul C et al.; 1 . According to the Fletcher-Huehns hypothesis there exists a functional difference between the two iron-binding sites of transferrin . 2 . The aim of the study presented was to evaluate this hypothesis in a homogeneous system, with human bone marrow cells and pure human monoferric transferrins A and B . 3 . For this reason normal human bone marrow cells were incubated with human monoferric transferrin . The monoferric transferrins A and B were obtained by selective labelling at different pH of apotransferrin followed by preparative isoelectric focusing in granulated gels . The uptake of iron by the cell suspensions from monoferric transferrins A and B was equal . 4 . In a heterogeneous but more active system for the removal of iron from human transferrin in vitro the two human monoferric transferrins did not show any significant functional differences . 5 . No support for the Fletcher-Huehns hypothesis could be obtained. Clin Exp Immunol, 1981 Feb, 43(2), 362 - 9 Prognostic significance of immunological tests in lung cancer; Check IJ et al.; We performed a battery of tests on peripheral blood samples from 94 patients with lung cancer to determine the extent to which immunological depression was due to abnormal lymphocyte function, as compared to changes in the number of lymphoid cells in the peripheral blood or in the efficiency of purification of cells in Ficoll-Hypaque gradients in preparation for testing . The percentage of lymphocytes in the gradient-derived cell suspension (%LG) was the most informative test . It decreased significantly with advancing stage of cancer and could predict survival of patients with uniform stage . The %LG correlated with survival better than any other test when multivariate analyses of all test combinations were performed . Low values of %LG reflected both the depressed lymphocyte counts and the altered buoyant density of the leucocytes of many patients with advanced cancer . A large proportion of the depression in other immune function tests was statistically attributed to changes in %LG . We concluded that this simple measurement provides valuable information about patients with lung cancer. J Natl Cancer Inst, 1981 Feb, 66(2), 355 - 62 Isolation of oval cells and transitional cells from the livers of rats fed the carcinogen DL-ethionine; Sells MA et al.; For the characterization of the metabolic and biologic properties of oval cells (i.e., cells emerging in the livers of rats treated with chemical carcinogens due to proliferation of bile ductular and/or duct cells) and transitional cells (i.e., cells having properties intermediate between those of oval cells and hepatocytes), these cells were isolated from the livers of Sprague-Dawley rats fed DL-ethionine for 4-5 weeks . The livers were dissociated into single cells by perfusion in situ with collagenase, and total cell suspensions were allowed to stand at unit gravity for 10 minutes to separate parenchymal (hepatocytes) from nonparenchymal cells . Nonparenchymal cells were centrifuged in linear gradients of Metrizamide (8-24% wt/vol), and 2-ml fractions were collected from the gradients . The cells in the fractions were defined by light microscopy, electron microscopy, and histochemical and immunofluorescence methods . A cell isolate was thus obtained consisting of Kupffer's cells (approximately 20%), bile ductular and/or duct cells and oval cells (approximately 30%), and transitional cells (approximately 50%) . A twofold enrichment of bile ductular and/or duct cells and their derivatives was achieved over that found in the nonparenchymal cell fraction before isopyknic gradient centrifugation. Thymus, 1981 Feb, 2(4-5), 225 - 33 Lymphocyte subsets in human thymus: expression of IgM-Fc receptor by peanut agglutinin positive and negative thymocytes; Musiani P et al.; The number of cells bearing receptors for the Fc portion of IgM (TM) and IgG (TG) has been evaluated in the human thymus . After overnight incubation the TM cells were 26.2 +/- 2.1% (mean +/- SE) in unfractionated thymus cell suspension, 23.4 +/- 3.8% in peanut agglutinin positive (PNA+) and 32.6 +/- 3.2% in PNA-negative (PNA-) fraction . PNA+ and PNA- fractions mainly consist of cortical and medullary thymocytes, respectively . The presence of TG was negligible . The fine structural features of TM cells from the thymus were very similar to those reported for TM lymphocytes from peripheral blood . In short-term culture a time-dependent increase of TM cells was observed . After 96 h of culture more than 60% of the thymocytes were shown to bear the Fc receptor of IgM . At the same time PNA+ and PNA- fractions expressed about 70% and 42% of TM cells respectively . The number of TG remained negligible . Our data indicate that in the human thymus there exists a distinct, quantitatively well-represented TM subset . Most probably, the peripheral TM cells derive from this subset which is present in both the cortex and the medulla of the thymus . Finally, the higher blastic responses to PHA exhibited by TM cells from PNA-subpopulation suggest that the most advanced intrathymic maturative step is attained by this distinct subset. Immunology, 1981 Feb, 42(2), 307 - 11 The guinea-pig complement-activating lymphocyte surface component (GPCA): a mouse T-cell marker distinct from Thy-1; Kierszenbaum F et al.; Mouse lymphocytes selectively expressing sensitivity to antibody-independent complement-mediated lysis by normal guinea-pig serum have been previously shown to be identical with T cells . This correlation has raised the question addressed in this work of whether or not the complement-activating surface component and Thy-1 were the same marker . S.49 mouse lymphoblastoid cells, sensitive to killing by anti-Thy-1.2 antibodies plus a source of complement activity devoid of non-specific cytotoxicity, were not sensitive to the non-specific, antibody-independent lytic effects of normal guinea-pig serum . Furthermore, rat thymus cell suspensions containing 93%--95% Thy-1-bearing cells were only partially susceptible (20%--35%) to guinea-pig serum cytotoxicity . Young rat thymus cells virtually devoid of guinea-pig serum-sensitive cells (less than 2%) were readily lysed by antirat Thy-1 and complement (greater than 98%) . While these results do not exclude the possibility that Thy-1 may constitute an indirect requirement for T cells to manifest sensitivity to GPS cytotoxicity, it is clear that this antigen does not confer such sensitivity by itself . Therefore, GPCA--the guinea-pig complement-activating marker of murine T cells--is not identical with Thy-1 and represents a distinct surface component of T lymphocytes which manifests itself in terms of a non-specific, but selective ability to activate complement. Cell Biol Int Rep, 1981 Feb, 5(2), 125 - 32 Growth and surface properties of dispase dissociated human fibroblasts; Cassiman JJ et al.; The dissociation of human fibroblast cultures with the bacterial neutral protease (Dispase II) was monitored by viability and growth measurement and was compared to the effect of trypsin and EDTA . Cell suspensions with high viability and excellent growth were obtained after 10 min incubation at 37 degrees C in 4 U/ml dispase in 0.02% EDTA . A two to three-fold increase in mitotic index occurred in the cultures within 48 h after dispase dissociation . The initial rate of aggregation was comparable to that of trypsin or EDTA dissociated cells, but attachment to a substratum and agglutination by Wheat Germ Agglutinin were markedly enhanced . The results indicate that dispase-EDTA provides a valuable alternative to the enzymatic dissociation with trypsin . Moreover, it is an additional tool for the dissociation of cultured cells and for the study of the surface properties of single cells. Boll Soc Ital Biol Sper, 1981 Jan 15, 57(1), 8 - 14 {Separation of osteoclasts from bird medullary bone by sedimentation at unit gravity}; Teti A et al.; A procedure for the isolation of osteoclasts from the heterogeneus population of medullary bone and marrow cells is presented . Cell suspensions were prepared from medullary bone of laying hens by mechanical dispersion . Following an osmotic shock and two 90 min unit gravity sedimentations, fractions highly enriched in osteoclasts were collected and subsequently cultivated in MEM with FCS . Ara C was added to the cultures for 48h to remove all the cells entering the mitotic cycle . The advantages and the possible applications of the method are discussed. Biochim Biophys Acta, 1981 Jan 8, 640(1), 223 - 30 Role of Ca2+ and Mg2+ in neutrophil hexose transport; O'Flaherty JT et al.; The influence of extracellular Ca2+ and Mg2+ on the transport of 2-deoxy-{3H}glucose into human polymorphonuclear neutrophils was studied . Omission of these cations from the cell suspensions had little effect on resting hexose uptake . Furthermore, the addition of the bivalent cation chelator, EDTA, depressed uptake only slightly . Similarly, neither cation was essential for the enhanced 2-deoxy-D-{3H}glucose uptake stimulated by two chemotactic factors (C5a and N-formylmethionylleucylphenylalanine) and arachidonic acid: enhanced uptake was only partially depressed by the omission of Ca2+ and Mg2+ from the suspensions and was still prominent in the presence of EDTA . Two other neutrophil stimulants, the ionophores, A23187 and ionomycin, also enhanced hexose uptake but their actions were heavily dependent upon extracellular bivalent cations and were totally abrogated by EDTA . In all instances, extracellular Ca2+, but not Mg2+, supported optimal enhanced hexose transport induced by stimuli . Activation of 2-deoxy-D-{3H}glucose uptake by each of the five stimuli was totally blocked by cytochalasin B (a blocker of carrier-mediated hexose transport) and D-glucose but not by L-glucose . The data indicate, therefore, that a variety of neutrophil stimulants activate carrier-mediated hexose transport . Although this transport can be triggered by the movement of extracellular Ca2+ into the cell (as exemplified by the action of the two ionophores), such Ca2+ movement is not required for the actions of chemotactic factors or arachidonic acid . Other mechanisms, such as a rearrangement of intracellular Ca2+, may be involved in mediating the activation of hexose transport induced by the latter stimuli. Cell Tissue Kinet . 1981 Jan;14(1):8590. An in vitro model of hematopoietic injury in chronic hypoplastic anemia; Fitchen JH et al.; Mice treated with high-dose busulfan develop a 'latent' form of bone marrow failure characterized by near-normal peripheral blood counts and marrow cellularity, but marked reductions in marrow pluripotent stem cells (CFUs) and myeloid progenitor cells (CFUc) . Spleen cell suspensions from control and 'latent' mice were placed in liquid culture in the presence of colony-stimulating activity . Cells were harvested at intervals up to 14 days and sub-cultured in agar to assay for CFUc . Baseline splenic CFUc did not differ significantly between control and 'latent' mice . Splenic CFUc from control mice increased 50-fold and reached a peak at day 10 in liquid culture . In contrast, splenic CFUc from 'latent' mice increased only 7-fold and reached a peak at day 3 . Our results indicate that although splenic CFUc are present in normal numbers in 'latent' mice, their proliferative capacity is markedly reduced, either as the result of defective CFUc self-renewal or defective feed-in from CFUs or both. Cell Tissue Kinet, 1981 Jan, 14(1), 39 - 52 A method for measuring the generation time and length of DNA synthesizing phase of clonogenic cells in a heterogenous population; Wu AM; Information on the cell cycle of progenitor cells in haemopoietic tissue is useful for understanding population control under physiological and abnormal conditions . Unfortunately, methods that have been developed for measuring cell cycle parameters are applicable only to cells of homogenous populations and not to morphologically non-recognizable progenitor cells such as colony forming units (CFU) that are present at low frequency in a heterogenous population . To circumvent this difficulty, a method was developed to measure CFU cell cycle parameters based on specific killing of cells in S phase by {3H}thymidine ({3H}TdR) . This was done by estimating the number of CFU killed following exposure of the cell suspension to {3H}TdR for various time periods . since cycling CFU are continuously entering S phase, a linear curve relating the percentage CFU-kill to the length of exposure of the cells to {3H}TdR in culture can be obtained . The slope of the curve (percentage kill/hr) indicates the rate that CFU enter the S phase and travel through the cell cycle . The inverse of this value will then represent a time period for CFU to move through a complete cell cycle (generation time) . The length of S phase can then be obtained by multiplying generation time by the fraction of cells in S phase at time zero . This method has been used to measure generation time and length of S phase of three kinds of haemopoietic progenitor cells: mouse granulocyte-macrophage CFU, human T lymphocyte CFU and CFU from regenerating mouse spleens . This method should be applicable to any normal or neoplastic clonogenic cell populations and the latter could be either of haematological or of solid tumour origin. Cancer Res, 1981 Jan, 41(1), 168 - 75 Isolation and characterization of epithelial cell types from the normal rat colon; Chopra DP et al.; Populations enriched in proliferative, mucous goblet, and absorptive cells were isolated by a method of repeated time dissociation of the colonic mucosa from the adult rat . Five sequential cell suspensions were obtained by rotating the everted colon on 0.2% trypsin solution in Eagle's minimum essential medium at 30 degrees for 20 min each time . Approximately 3.7 x 10(7) cells were obtained from each colon, and 87% of the cells were found viable . The results, based on the {3H}-thymidine-labeling index, {3H}thymidine incorporation, thymidine kinase activity, and histochemical and ultrastructural observations, indicate the Cell Suspension I, separated from the luminal surface, contains 82 +/- 9% (S.D.) absorptive cells; Suspension III, separated from the middle level of the crypts, contains 80 +/- 7% mucous cells . Suspension V, isolated from the crypt base, contains the majority of proliferative epithelial cells (85 +/- 10%) . This method provides a suitable tool for a variety of studies in colon carcinogenesis. Acta Physiol Acad Sci Hung, 1981, 58(4), 311 - 5 Effect of cyclophosphamide on the acute phase of experimental myocardial infarction in rats; Lepran I et al.; The effect of a single dose of the cytotoxic drug cyclophosphamide on the severity of acute myocardial infarction in conscious rats has been studied . One and 4 days after the i.p . injection of 100 mg kg-1 of the drug, the survival rate of rats subjected to coronary ligation was significantly increased . The occurrence of fatal arrhythmias was markedly reduced . The peripheral white blood cell count was profoundly lowered . On the first day after pretreatment, moderate granulocytosis with severe lymphopenia occurred . Four days following the administration of cyclophosphamide, marked granulocytopenia also ensued . In this stage, restoration of white blood cell count by cell suspension prepared from the spleen of normal rats did not significantly affect the cardioprotective effect of cyclophosphamide . The data provide additional evidence that protein synthesis is involved in the early phase of myocardial infarction . The significance of leukocytes in the phenomenon is briefly discussed. Scan Electron Microsc, 1981, 4, 55 - 60 Disappearance of microvilli accompanying radiation-induced interphase death of rat thymocytes; Yamada T et al.; Thymocytes are radiosensitive and show interphase death (immediate death) after irradiation . The present investigation was undertaken to characterize by scanning electron microscopy the shape of thymocytes irradiated in vitro with 1 kR X-rays and determine the cell surface changes that accompany the development of interphase death . Exposing the cells to X-rays caused disappearance of the microvilli, usually observed with viable cells . Association of dead thymocytes with porous surface without any digitations was confirmed by the examination of enriched dead cell populations prepared by centrifuging the cell suspension in Percoll density gradient. Curr Eye Res, 1981-82, 1(10), 579 - 89 Interaction of bovine pigment epithelium cells, photoreceptor outer segments, and interphotoreceptor matrix: a model for retinal adhesion; Adler AJ et al.; A model system for retinal adhesion, consisting of interacting bovine pigment, epithelium (PE) cells and photoreceptor outer segments, was utilized to examine any adhesive role of the interphotoreceptor matrix . PE cells were dissociated by trypsin treatment of the eyecup, and were allowed to replenish their surface proteins as single cells in spinner culture . They were then placed into short-term, slowly rotating suspension cultures, where their rapid aggregation as a function of time could be studied . Outer segments exhibited no tendency to interact with one another, or with PE cells, in suspension culture . Extracellular interphotoreceptor matrix from adult bovine eyes was isolated by rinsing the apposing surfaces of neural retina and PE . When this matrix material was added to the cell suspension cultures, adhesion between PE cells and outer segments in mixed cultures was not enhances . However, PE reaggregation itself appeared to be augmented . The principal matrix glycoprotein, obtained by concanavalin A affinity chromatography, displayed adhesive properties similar to those of the interphotoreceptor matrix . Thus, under the conditions of these in vitro experiments, no evidence could be obtained that either cell surface molecules or interphotoreceptor matrix plays a role in retinal adhesion. Microbiol Immunol, 1981, 25(12), 1327 - 34 A simplified micro-method for cytotoxicity testing using a flat-type titration plate for the detection of H-2 antigens; Shiroishi T et al.; A new micro-method for cytotoxicity testing using a flat-type titration plate has been developed . This micro-method is so simple that it involves no washing and staining procedures . In this method, cell viability is judged on the basis of the refractility change in the lymphocytes seen under the phase-contrast microscope . This micro-method has higher sensitivity and reproducibility than the dye exclusion method and 51Cr-release assay . Fixation of cells with glutaraldehyde and tight covering of the plate allow us to read the result even after 3 weeks . These advantage of the new method enable us to deal with a large number of samples at one time . We also introduce a rapid technique for the preparation of a cell suspension from the lymph node without killing the animal. Microbiol Immunol, 1981, 25(11), 1109 - 18 Selective and differential media for isolation and tentative identification of each species of Pityrosporum residing on normal or diseased human skin; Ushijima T et al.; Selective and differential media were designed for each species of Pityrosporum; P . pachydermatis, P . ovale, and P . orbiculare in order to make feasible a quantitative cultivation . Medium for P . pachydermatis (medium A) was composed of 1% trypticase peptone (BBL), 0.5% yeast extract (BBL), 0.3% glucose, 0.2% NaCl, 1.2% KH2 PO4 (anhydrous), 1.5% agar, 0.01% ampicillin, and 0.025% cycloheximide with a pH of 5.5 . Medium for P . ovale (medium B) was medium A supplemented with 0.05% sodium acetate (anhydrous), 0.2% Tween 80, and 0.025% (selective medium) or 0.075% (differential medium) sodium laurate . Medium for P . orbiculare was medium B (devoid of laurate) supplemented with 2% olive oil, 0.25% glycerol, 0.25% gall powder, 0.05% sodium palmitate, 0.05% sodium stearate, 0.05% sodium oleate and 8% (selective medium) or 10% (differential medium) sodium lactate and an increase in Tween to 1% . For isolation of Pityrosporum, specimens were suspended in 0.1% Tween 80 solution and inoculated onto agar plates of three selective media . The plates were incubated aerobically at 37 C for 8-10 days under conditions of prevention of water loss from the media . The plating efficiency of each selective medium, expressed as a ratio of cultural counts to microscopic counts was generally over 70% . Species of Pityrosporum could also be identified when we inoculated the cell suspension onto differential agar plates and incubated the preparations at 37 C for 7 days. Acta Biol Med Ger, 1981, 40(6), 801 - 9 {Production of primary cells cultures by a semi-industrial technic . II . Use of active trypsin solutions for effective cell yields}; Beyer R et al.; For effective cell yields in semi-industrial production of primary cell suspensions, the use of trypsin solutions with high enzyme activity has been proved to be an essential requirement . Enzyme solutions with a constant high activity may be obtained by using an optimal filtration method . Filtration results using the deep and membrane filtration method to sterilize trypsin solutions are demonstrated and discussed . A number of trypsin samples taken from the "Leidholdt-Firm" were tested concerning cell yields in the so-called 'Biomix-Procedure' . A correlation could be shown to exist between enzyme activity and cell yield . The so tested trypsin-L-samples out of the years 1976/78 were proved to be first-rate preparations for routine production of primary piglet and calf kidney cell cultures. Acta Biol Med Ger, 1981, 40(6), 739 - 42 Spreading of red blood cell suspensions on paper as a simple test of cell deformability; Tannert C et al.; Spreading of red blood cells on paper is a simple and rapid measure of erythrocyte deformability . It requires a small number of cells and is accurate enough to be applied to studies of deformability alterations caused by heat damage, metabolic depletion or storage of the cells . It is suggested as a test of red blood cell viability. Acta Biol Med Ger, 1981, 40(4-5), 717 - 20 Blood preservation considering the possibilities, the patient and the purse; Hogman CF; An efficient motivation for the clinician to use blood components instead of whole blood is that the components have improved quality . Systems of integrally connected plastic bags makes the preparation easy and safe . This means that the blood can be collected in a solution which is not at the same time the storage medium . The anticoagulant may--but must not necessarily--be one of the conventional solutions, ACD or CPD . Adenine and possibly guanosine should preferably be added to a separate red cell suspension medium which can simply consist of sodium chloride, adenine and glucose . In order to avoid elevated hemolysis, sucrose or mannitol can be enclosed in the medium, if the protein concentration is low . Control of pH by a buffering system or an ion-exchange device is a necessity if the liquid storage shall be possible for more than 5--6 weeks . An interesting new possibility to improve the storage conditions is to introduce a chemical coupling of heparin onto the inner surface of the plastic bag. J Immunol Methods, 1981, 47(1), 31 - 8 Monocyte purification with counterflow centrifugation monitored by continuous flow cytometry; De Mulder PH et al.; Continuous monitoring of cell light scatter during counterflow centrifugation of a mononuclear cell suspension allows counting and size recognition of the cell types elutriated . With this method an optimal separation point between monocytes and lymphocytes, determined for each individual donor, may be established . With a constant flow of 15 ml/min this separation point is found at centrifugal velocities ranging from 2348 to 2444 rpm (n = 10) . From 50 ml venous blood, 84.1% +/- 4.1% (15.7 +/- 8.6 x 10(6)) of all elutriated monocytes, with a purity of 92.4% +/- 1.4%, is collected in a volume of 50 +/- 1 ml . In the same run, 92% +/- 4.3% of the lymphocytes is gathered in one fraction with a purity of 98.9% +/- 0.7% . After counterflow centrifugation, 91.6 +/- 10.5% of the cells loaded is recovered; viability exceeds 98%. Bioelectromagnetics, 1981, 2(3), 203 - 15 In vitro microwave effects on human neutrophil precursor cells (CFU-C); Ottenbreit MJ et al.; Human marrow cells were irradiated with 2450-MHz CW microwaves in a fluid-filled waveguide irradiation system . Cell exposure was conducted by placing a marrow cell suspension in 20-microliter glass microcapillary tubes were positioned in the exposure chamber, and irradiated at power densities from 31 to 1,000 mW/cm2 (with corresponding specific absorption rates of 62 to 2,000 mW/g) for 15 minutes . The temperature of the sample was maintained at a fixed point . Sham-irradiated (SI) and microwave-irradiated (MWI) cells were cultured in a methylcellulose culture system for neutrophil colony proliferation . There was no reduction in neutrophil colony number on days 6-7 or 12-14 in cells exposed at 31 or 62 mW/cm2, but as the power density was increased to 1,000 mW/cm2, there was a reduction in colony number of MWI cells compared with SI cells . The microwave interaction with the human neutrophil colony-forming cells was apparently not related to temperature rise, or to the state of cells cycle, and was irreversible. Biosci Rep, 1981 Jan, 1(1), 79 - 87 Continuous replication of potato spindle tuber viroid (PSTV) in permanent cell cultures of potato and tomato; Muhlbach HP et al.; The continuous replication of potato spindle tuber viroid (PSTV) in callus cultures from PSTV-infected wild-type potato (Solanum demissum L.) and tomato (Lycopersicon peruvianum L . Mill) plants and in cell suspensions derived from potato protoplasts (Solanum tuberosum L.) inoculated in vitro is described . The persistence of PSTV replication in these cell lines through at least 14 subculture passages, which corresponds to a continuous replication over a period of more than one year, was demonstrated by infectivity assay and by polyacrylamide-gel electrophoresis of isolated nucleic acids . This continuous synthesis de novo of PSTV was substantiated by the incorporation of {3H}uridine and of {32P}orthophosphate into viroid RNA. Z Parasitenkd, 1981, 65(2), 131 - 6 Fine structure of the oocyst wall and excystation of Eimeria procyonis from the American raccoon (Procyon lotor); Duszynski DW et al.; Oocysts of Eimeria procyonis, from the American raccoon (procyon lotor), were broken, added to a cell suspension, fixed in Karnovsky's fluid, and studied with the electron microscope . The oocyst wall has three layers: a thin electron-dense inner layer (8-15 nm), an electron-lucent middle layer (25-35 nm), and a thick outer layer (120-140 nm) . The outer layer has an electron-dense inner portion and an electron-lucent outer portion that contains membrane-bound vesicles . When exposed to a trypsin-sodium taurocholate fluid, sporozoites excysted from most sporocysts which were 35-43 months old, but not from sporocysts that looked normal and were 106 months old . Excysted sporozoites measured 13-16 x 3-4 (mean 14.3 x 3.2) micrometer, usually had two refractile bodies, and had a nucleus with a prominent nucleolus. J Membr Biol, 1981, 62(1-2), 149 - 60 Amino Acid Transport and stimulation by substrates in the absence of a Na2+ electrochemical potential gradient; Heinz A et al.; Uptake of alpha-aminoisobutyric acid (AIB) was examined in Ehrlich ascites tumor cells treated with the cation-exchange ionophore nigericin (20 microgram/ml) . Membrane voltages were measured using the voltage-sensitive dye diethyloxadicarbocyanine (DOCC) . In normal phosphate-buffered media, nigericin changed the distribution ratios of Na+ and K+ (the ratio of intra- to extracellular concentrations) nearly to unity, but AIB was still accumulated to a distribution ratio of approximately 9.0 . When all but 40 mM Na+ in the medium was replaced by choline, nigericin resulted in K+ loss and Na+ gain and both cation distribution ratios approached 2.8-3.4, as would be expected if both ions were distributing near electrochemical equilibrium with a membrane voltage in the range of -28 to -33 mV . This conclusion was supported by the observation that the addition of 5 X 10(-7) M valinomycin to the nigericin-treated cell suspension produced no change in DOCC absorbance . In spite of the apparent zero electrochemical potential gradients for Na+ and K+, AIB was accumulated to a distribution ratio of 5.4 in the low-Na+ medium . Addition of 0.1 mM oubain or 50 microM vanadate did not alter the extent of AIB accumulation as would have been expected if a large component of the membrane voltage were due to electrogenic operation of the (Na+ + K+)-ATPase . Addition of lactate, pyruvate or glucose increased the AIB distribution ratios to 11.9, 9.4 and 15.3, respectively . The effect of glucose could be explained, at least in part, by an enhanced Na+ electrochemical potential gradient . However, neither lactate nor pyruvate produced any change either in membrane voltage or the intracellular Na+ concentration . Therefore, these results confirm the existence of a metabolic energy source which is coupled to AIB accumulation and operates in addition to the Na+ co-transport mechanism, and which is augmented by metabolic substrates such as lactate and pyruvate. Folia Biol (Praha), 1981, 27(4), 284 - 8 On the mechanism of intercellular adhesion; Peknicova J et al.; Using a 2 X 2 design, the collection rates of cells from radioactively labelled single-cell suspension on cell-coated collecting surfaces were tested for a possible H-2 effect on intercellular adhesiveness . In five donor-host combinations differing in the H-2 complex, no straight advantage of syngeneic over allogeneic relationship could unequivocally be proved . Trypsin was deliberately omitted in preparing the single-cell suspension; it was shown that the cell suspension processed in this way affected the mechanism of intercellular adhesiveness to a degree at least comparable to the role played by the properties of the collecting layer cells. Folia Biol (Praha), 1981, 27(4), 255 - 64 Growth of normal and tumour haemopoietic cells on glass coverslips in peritoneal cavity of mice . II . Immunological identification of bone marrow colony-forming cells; Rozinova EN et al.; The effect of rabbit anti-mouse brain serum and monospecific antibodies to an antigen of erythroblasts, Ag-Eb, on cells forming haemopoietic colonies on glass coverslips was studied . These colonies developed on the fibroblast-macrophage layer formed on glass coverslips which had been inserted into the peritoneal cavity of mice . when bone marrow cell suspensions were treated with RAMBS and the complement, or with anti-Ag-Eb antibodies plus complement, the CFU-GC colony-forming ability remained unaffected . In the examined cell suspensions, RAMBS inactivated 44% of CFU-S . When bone marrow cell suspensions were treated with anti-Ag-Eb antibodies, the number of immature erythroid cells was reduced to 75% . The CFU-GC are presumed to resemble the committed (unipotent) haemopoietic progenitor cells. Carcinogenesis, 1981, 2(3), 195 - 8 The cellular DNA content and profile of dimethyl nitrosamine induced marine lung tumors in relation to their histopathology; Digernes V; A total of 61 lung tumors induced in NMRI mice by the carcinogen dimethyl nitrosamine were classified as solid adenomas or papillary tumors . Single cell suspensions from each of these tumors were analyzed by flow cytometry according to DNA content . The adenomas contained stemlines having a diploid DNA content, a finding which is in accordance with the benign nature of these tumors . In the papillary tumors heteroploid stemlines were often found in addition to diploid cells, this seems to be a sign of their malignant properties . The significance of the heteroploidy and the clustering of stemlines at the near diploid and the tetraploid levels in these tumors is discussed. J Immunol Methods, 1981, 43(1), 87 - 93 Criteria of cell killing in vitro; Pullen GR et al.; The attachment of cells to plastic tissue culture plates is a widely used criterion of cell viability in microcytotoxicity assays . When cell suspensions of primary human colon carcinoma and melanoma cells which were of low initial viability (assessed by trypan blue exclusion) were allowed to adhere to tissue culture plates, many of the adhering cells did not satisfy other criteria of cell viability . They did not stain with fluorescein diacetate but did stain with propidium iodide and fluorescein-labelled antinuclear antibodies . Using complement-mediated cytotoxicity, relatively weak activity was inconsistently demonstrated against these cells . On the other hand, strong activity was always demonstrated against highly viable cells from one colon carcinoma and two melanoma cell lines . Immunologically mediated damage to these cells was demonstrated readily by loss of the ability to stain with fluorescein diacetate, but not by cell detachment. Cytometry, 1981 Jan, 1(4), 272 - 8 Intracellular binding of fluorescein in lymphocytes; Meisingset KK et al.; The fluorescence characteristics of intracellular fluorescein, formed by the hydrolysis of fluorescein diacetate in peripheral human lymphocytes, were studied by fluorometry on cell suspensions and compared to those of albumin bound and free fluorescein in solution . The absorption and fluorescence spectra of both intracellular fluorescein and fluorescein in aqueous solutions of albumin and glycerol were red shifted by 2-10 nm as compared to the spectra of fluorescein in phosphate buffered saline . The fluorescence polarization (P) of both intracellular fluorescein and a mixture of albumin-bound and free fluorescein showed a decrease towards low emission wavelengths and an increase toward high excitation wavelengths . The results were found to be consistent with a simple model assuming that part of the intracellular fluorescein is dissolved in the aqueous phase of the cytoplasm, giving P less than 0.1, while the rest is bound to macromolecules, giving P = 0.33 . The fraction of bound intracellular fluorescein was estimated to be about 70% . Fluorescein was found to bind with high affinity and more rigidly (P = 0.43) to albumin than to intracellular macromolecules in general. J Hyg Epidemiol Microbiol Immunol, 1981, 25(2), 121 - 33 Use of physiological and genetic algal tests for the hygienic evaluation of pollutants in waters . Part I . 226Ra studies; Havlik B et al.; After exposure for several days of intensively growing algal cultures to a radium concentration of 10(-6) g.1(-1) the physiological and genetic responses of Chlorella kessleri and Scenedesmus obliquus cells could be clearly demonstrated . Of physiological characteristics, we recorded the frequency of cells with a reduced number of autospores, the length of the lag phase and cell survival . Of genetic responses, the frequencies of delayed lethal and sublethal effects, the frequencies of minute colonies, colonies with big cells and scabby colonies, and the frequencies of pigmentation changes were followed . The above concentration produced no toxic effect . No simple and clear-cut correlation was found between the recorded responses and the amount of accumulated radium in algal cells . For an interpretation of the mutagenic effect we had to assume a homogeneous radioactive irradiation of the mixed cell suspension . We had also to take into account the participation of repair systems of algae with regard to their survival or manifestation of mutagenic changes in individuals in which mutagenesis had been induced. Curr Med Res Opin, 1981, 7(6), 352 - 8 A new method of measuring red cell deformability and the effects of pentoxifylline; Isogai Y et al.; The effect of pentoxifylline on red cell deformability was studied using a new device of a polycarbonate membrane filter ("Nuclepore') and a differential pressure type transducer . Differential pressure fluctuations due to the passage of red cells through the "Nuclepore' membrane were recorded as a differential pressure curve of red cell filtration . Red cell deformability was measured with a diluted red cell suspension (10 x 10(4)/mm3) in this buffered NaCl solution of normal osmotic pressure, 300m0sm/l as a suspending medium . Blood samples were taken from 9 healthy subjects and 26 patients with diabetes mellitus, liver disease and miscellaneous diseases . The ability of pentoxifylline to modify red cell deformability was investigated using concentrations of 20 micrograms/ml and 40 micrograms/ml in diluted red cell suspension which was incubated for 2 hours of 37 degrees C . Changes in red cell deformability were estimated from the gap between two differential pressure curves recorded in blood suspension with or without pentoxifylline . Addition of pentoxifylline increases red cell deformability in both groups of blood suspension in healthy subjects and patients . A significant increase in red cell deformability was observed in 40% of samples with the addition of 29 micrograms/ml pentoxifylline and in 77% of those with 40 micrograms/ml, respectively. J Physiol, 1981 Jan, 310, 321 - 36 Effects of external cations on calcium efflux from single cells of the guinea-pig taenia coli and porcine coronary artery; Hirata M et al.; 1 . Single smooth muscle cells were prepared from the guinea-pig taenia coli and the porcine coronary artery by treatment with collagenase, in order to measure the 45Ca flux with special reference to the effects of external cations . 2 . In excess {K}o solution, cell suspensions prepared from both tissues showed an increased 45Ca uptake within 3-5 min . In Na-free solution, the cells prepared from taenia coli, but not from coronary artery showed an increased 45Ca uptake . The Ca uptake of the cells paralleled with the tension increase recorded from tissues of both species . 3 . The efflux of 45Ca into Ca-free EGTA containing solution was markedly increased by {Na}o in cells from the taenia coli, but not in cells from the coronary artery . 4 . The {Na}o-activated 45Ca efflux from cells of the taenia coli was slightly larger in Ca-free solution than in the Ca-containing (10)-4) M) solution . Depolarization of membranes produced by excess {K}o did not effect the {Na}o-activated 45Ca efflux . 5 . Increase in {Na}i by treatment with K-free solution suppressed the {Na}o-activated 45Ca efflux in the taenia coli . Re-addition of {K}o reactivated the {Na}o-activated 45Ca efflux . This re-activation was blocked by ouabain . 6 . The efflux of 45Ca was slightly activated by {Ca}o in cells from the taenia coli . This {ca}o-activated 45Ca efflux was larger in Na-free solution than in Na-containing solution, thus suggesting interactions between {Na}o and {Ca}o on the Ca efflux . 7 . In cells from the taenia coli, 45Ca efflux could still be observed in nominally Na-and Ca-free solution . This residual 45Ca efflux made a large contribution to the total 45Ca efflux . 8 . When 45Ca uptake was measured in Na-free (Tris) solution, the {Na}o-activated, {Ca}o-activated and residual 45Ca effluxes of cells from the taenia coli were accelerated, non-selectively . 9 . These results obtained with cells prepared from the guinea-pig taenia coli are comparable to the Ca2+ efflux mechanism seen in the squid axon . However, maintenance of low concentrations of {Ca}i seems to require not only the above three 45Ca efflux mechanisms, but also Ca sequestering mechanisms in the cell. J Immunol Methods, 1981, 41(3), 289 - 301 Separation of AKR mouse thymus lymphoma from normal thymic cells by centrifugal elutriation; Meistrich ML et al.; The rapid separation of large numbers of viable thymus cells from AKR mice bearing transplanted or spontaneous thymic lymphomas was achieved by centrifugal elutriation . Separation of more than 3 x 10(8) cells from either a transplanted lymphoma, designated 720, or from a spontaneous thymic lymphoma, required only 15 min . Unfractionated thymus cells obtained from mice bearing the transplanted lymphoma consisted of 80% lymphoma cells (by immunofluorescence for the viral protein, gp70) and 20% normal cells . Fractions of slowly sedimenting cells consisted almost exclusively of normal cells (95%) with modal volumes of 95 micrometers cubed . Fractions of rapidly sedimenting cells consisted of 95% tumor cells with volumes of 150-400 micrometers cubed . The slowly sedimenting cells were almost exclusively (98%) in the GO- or G1-phase . Fractions of rapidly sedimenting cells contained up to 55% S-phase and up to 30% G2-phase cells . Intermediate fractions contained mixtures of normal cells and small GO- or G1-phase tumor cells . Thymidine uptake by the separated cells was determined . The fractions containing normal cells showed little thymidine uptake after 4 and 48 h in culture, while the fractions of tumor cells showed high levels of incorporation . In contrast to the high levels of thymidine uptake by the tumor cell fractions after 48 h in culture, there was little uptake by the unseparated cell suspension, suggesting a possible interaction between normal and tumor cells during the culture period. Somatic Cell Genet, 1981 Jan, 7(1), 43 - 57 Quantitative cell fusion: derivation and application of theoretical models; Rohme D et al.; Cell fusion experiments were made on ten cell lines representing seven mammalian species, using inactivated Sendai virus . The extent of fusion was determined microscopically and tabulated as frequencies of cell with different numbers of nuclei . Expected distributions were derived theoretically under certain assumptions concerning the fusion process . A random model was assumed according to which the tendency to fuse depends only on the cell size, expressed as the number of nuclei present in the cell . Three distributions were derived, which were referred to as the simple, additive, and multiplicative models . The additive model pertained to fusions made in cell suspensions and the multiplicative one mainly to fusions in fibroblast monolayers. Scand J Rheumatol, 1981, 10(1), 55 - 61 Characterization of antibody-dependent cell-mediated cytotoxicity in rheumatoid synovial fluid; Abrahamsen TG; Synovial fluids of 15 rheumatoid arthritis patients, 11 juvenile rheumatoid arthritis patients, and 6 patients with other chronic inflammatory joint diseases were studied . The isolated mononuclear cells, mainly monocyte/macrophages and lymphocytes, were preincubated overnight and then cultured with antibody-sensitized 51Cr-labelled chicken erythrocytes . There was no significant difference in the cytotoxicity between mononuclear cells from synovial fluid and peripheral blood of the patients . Depletion of adherent cells of the mononuclear cell suspensions by nylon wool column fractionation or by incubation on plastic dishes and depletion of cells which had phagocytosed colloidal iron had no substantial effect on the cytotoxicity whereas depletion of cells forming rosettes with human erythrocytes sensitized with human IgG (EA-RFC) practically abolished the activity . This shows that in rheumatoid synovial fluids the net effect of monocyte/macrophages in this in vitro antibody-dependent cytotoxic assay is small . Thus, the main effector cells in these mononuclear cell suspensions are lymphocytes which probably exhibit surface-bound Fc-receptors. J Cell Biol, 1981 Jan, 88(1), 138 - 48 Studies on cell adhesion and recognition . II . The kinetics of cell adhesion and cell spreading on surfaces coated with carbohydrate-reactive proteins (glycosidases and lectins) and fibronectin; Carter WG et al.; The kinetics of cell attachment and cell spreading on the coated surfaces of two classes of carbohydrate-reactive proteins, enzymes and lectins, have been compared with those on fibronectin-coated surfaces with the following results: (a) A remarkable similarity between the kinetics of cell attachment to fibronectin-coated and glycosidase-coated surfaces was found . In contrast, cell attachment kinetics induced by lectin- and galactose oxidase-coated surfaces, in general, were strikingly different from those on fibronectin and glycosidase surfaces . The distinction between fibronectin- or glycosidase- and lectin- or galactose oxidase (an enzyme with lectin-type characteristics)-coated surfaces was further supported by the finding that cytochalasin B and EDTA inhibited cell attachment to fibronectin- and glycosidase-coated surfaces but not lectin-coated surfaces . (b) Fibronectin, if labeled and added to a cell suspension, showed only low or negligible interaction with the cell surface . However, fibronectin absorbed on plastic surfaces showed a high cell-attaching activity . It is assumed that fibronectin coated on plastic surfaces may form polyvalent attachment sites in contrast to its lower valency in aqueous solution . (c) Various inhibitors of cell attachment to both fibronectin-, galactose oxidase-, and lectin-coated surfaces were effective only during the first few minutes of the adhesion assay, after which time the attached cells became insensitive to the inhibitors . It is suggested that the initial specific recognition on either lectin-type or fibronectin-type surfaces is followed by an active cell-dependent attachment process . The primary role of the adhesion surface is to stimulate the cell-dependent attachment response . (d) Cells attached on tetravalent concanavalin A (Con A) spread very rapidly and quantitatively, whereas divalent succinyl Con A and monovalent Con A were effective stimulators of cell attachment but not cell spreading . Cross-linking of succinyl Con A restored the cell spreading activity . Tetravalent Con A surfaces specifically bind soluble glycoproteins, whereas succinyl Con A has a greatly reduced ability to bind the same glycoproteins . These results suggest that cross-linking of cell surface glycoproteins by the multivalent adhesive surface may trigger the cellular reaction leading to cell spreading. Pathol Annu, 1981, 16 Pt 1, 101 - 37 Diagnostic electron microscopy . I . Hematology: differential diagnosis of acute lymphoblastic and acute myeloblastic leukemia . Use of ultrastructural peroxidase cytochemistry and routine electron microscopic technology; Dvorak AM et al.; We have described in detail a number of ultrastructural methods which we have found to be useful for the evaluation of hematologic cases submitted to our diagnostic electron microscopic unit . The techniques include the preparation of peripheral blood for study as both buffy coat and cell suspension specimens and the preparation of bone marrow spicules . Ultrastructural methods for the demonstration of glycogen and peroxidase are detailed . The study of such material includes light microscopic study of plastic-embedded, alkaline-Giemsa-stained one micron sections as well as ultrastructural studies . All hematological cases submitted for ultrastructural analysis in a two-year period were reviewed and are presented here . The identification of individual mature cells was relatively simple using light microscopy . Populations of blasts could also easily be recognized . Further differentiation of blasts, primarily lymphoblasts or myeloblasts, was done using ultrastructural cytochemistry where needed . These techniques can easily be done in electron microscopy units concerned with diagnostic work . We submit that pathologists and hematologists should have access to the diagnostic tools described here in order to manage patients with acute leukemia. J Immunol Methods, 1981, 47(1), 39 - 48 Simultaneous analysis of cell surface antigens and cell morphology using monoclonal antibodies conjugated to fluorescent microspheres; Mirro J Jr et al.; A new method for simultaneous analysis of cell surface antigens and cell morphology using monoclonal antibodies conjugated to fluorescent microspheres ('immunospheres') is described . Wright's staining was performed on cells after reaction with immunospheres, and a direct correlation of cell surface antigen expression and cell morphology was made . Mild formalin fixation of cells inhibited phagocytosis of microspheres, which is a potential source of confusion in the analysis of cell surface binding . Rapid, accurate analysis of cell surface antigen expression in single cell suspensions of heterogeneous human hematopoietic and lymphoid cell populations was facilitated by this method. Ciba Found Symp, 1981, 84, 246 - 64 Histophysiology of follicular structures and germinal centres in relation to B cell differentiation; Nieuwenhuis P et al.; A historical review of germinal centres since their original description in 1885 by Flemming is given, followed by a description of the germinal centre reaction . Microenvironmental aspects are discussed, special attention being paid to the role of antigen in the induction and further regulation of germinal centre activity . Germinal centres give rise to a new population of lymphocytes (germinal centre-derived cells) . Their immune capacities were studied using the rabbit appendix as an experimental model . The possible properties of the cell from which a germinal centre develops (germinal centre precursor cell) were investigated using germinal centre cell suspensions from the rabbit appendix and bone marrow and thoracic duct lymphocytes in the rat in adoptive transfer systems . From these experiments a working hypothesis was formulated for the possible role of germinal centres in B cell differentiation . Essentially, germinal centres are thought not only to be sites of selective amplification of antigen-reactive but still immature B cells, resulting in the generation of memory cells, but also to contribute to the overall pool of B cells by polyclonal amplification of B cells of specificities unrelated to the antigen inducing the germinal centre. Cancer Biochem Biophys, 1981, 5(3), 153 - 61 Three dimensional reconstruction of disseminated cancer modules; Chawla SD et al.; A computer system which can be used to perform three dimensional reconstruction of dispersed cancer growth from serial sections is described . The system was used to reconstruct and analyze melanoma nodules growing in the left upper lobes of the lungs of seven mice following intravenous inoculation of a single cell suspension of melanoma cells . The analysis shows a great variability in the tumor size within a single mouse, and also in different mice . The preponderance of tumors (40 out of 50) were subpleural and extended to the surface of the lung at some point . An analysis of the spatial distribution of the tumors was performed which indicates that the tumors may not be randomly dispersed in the lungs . The mechanisms which could lead to the observed results are discussed. J Immunol Methods, 1981, 43(3), 269 - 75 Propidium iodide as a nuclear marker in immunofluorescence . II . Use with cellular identification and viability studies; Yeh CJ et al.; The use of a fluorescence method employing propidium iodide to determine nuclear morphology of different cell types by epi-illumination in membrane immunofluorescence is described . Its usefulness is illustrated by differentiating nucleated and anuclear transferrin receptor-bearing cells as well as by simultaneously determining the viability of cell suspension without requiring a change from fluorescence microscopy, this causing no loss of visual accommodation. Natl Inst Anim Health Q (Tokyo), 1981 Spring, 21(1), 21 - 5 Production of heat-stable enterotoxic component by Escherichia coli strains enteropathogenic for swine; Kashiwazaki M et al.; Forty-nine strains of Escherichia coli isolated from piglets with or without neonatal diarrhea were all negative for both classical heat-labile and heat-stable enterotoxins when examined by the 18-hour rabbit ileal loop test with culture supernatants, the mouse Y1 adrenal cell test, and the infant mouse test . Of them, seven strains from diarrhea were positive only when tested in porcine ileal loops challenged with living bacterial cell suspensions or culture supernatants . The enterotoxic component produced by them was resistant to heating at 80 degrees C for 20 minutes . It presented a rapid onset of action with a long duration, although the amount of accumulated fluid in the porcine ileal loops was small . These porcine strains of E . coli which has been positive only for the porcine ileal loop test were designated "ST pig loop" or "STpl" strains. Acta Neurochir (Wien), 1981, 55(3-4), 181 - 200 The utilization of native glioma antigens in the assessment of cellular and humoral immune responses in malignant glioma patients; Apuzzo ML et al.; Cellular and humoral components of the immune response have been studied preoperatively, concurrently, and serially in patients with malignant glial neoplasms . In order to assess titres of circulating antibodies to tumour cell constituents an indirect immunofluorescent technique was applied to single cell suspensions and snap frozen cell smears . In an allogeneic system, 49% of 47 test and 7% of 124 control sera gave a positive response to cytoplasmic components . The leucocyte adherence inhibition assay was applied to study 39 test and 64 control patients . Significant non-adherence of leukocytes was observed in 77% of test cases . Control parameters indicated specificity of the response . Simultaneous assessment in 28 test patients yielded a positive response for one or both assays in 89% of cases.. Endocrinology, 1981 Jan, 108(1), 18 - 26 Preparation of smooth muscle cell suspensions from the rabbit oviduct and prostaglandin binding analysis; Riehl RM et al.; The preparation, viability, and prostaglandin (PG) binding characteristics of dispersed smooth muscle cells from the rabbit oviduct have been determined . Cell suspensions were prepared by enzymatic degradation of the intracellular matrix and subsequent mechanical dispersion with a wide bore pipette . The digestion media consisted of a modified Hanks' Balanced Salt Solution, pH 7.1, containing 1.6 U/mg wet wt elastase and 8 U/mg wet wt collagenase . This method provided a yield of single smooth muscle cells of approximately 3 x 10(6) cells/100 mg wet wt within 3-4 h of organ removal . Cell viability was determined by trypan blue dye exclusion, retention of lactate dehydrogenase, absence of 57Co-EDTA uptake, and ouabain sensitivity of cationic transport . Roughly 80% of the isolated cells remained viable after the digestion procedure . The dispersed cells specifically bound {3H}PGE2 and {3H}PGF2 alpha . Scatchard analysis of the binding data revealed separate homogenous populations of high affinity sites for both PGE2 and PGF2 alpha . The equilibrium dissociation constants and total sites per cell were 0.55 nM and 11,332 for PGE2 and 0.19 nM and 5,154 for PGF2 alpha, respectively . Specific labeled PG binding was inhibited in a concentration-dependent manner by increasing amounts of unlabeled PG . Inhibition of labeled PGE2 binding by unlabeled PGF2 alpha, and vice versa were negligible, except at high concentrations . The results indicate that smooth muscle cells can be enzymatically dispersed from the rabbit oviduct with minimal damage . Also, these cells possess distinct specific binding sites for PGE2 and PGF2 alpha that differ in regard to affinity and total number of sites per cell. Can J Surg, 1981 Jan, 24(1), 39 - 44 Studies into the mechanism of reversal of experimental acute hepatic failure by hepatocyte transplantation . 1; Makowka L et al.; The authors reported previously that single cell suspensions of syngeneic, allogeneic and xenogeneic hepatocytes can significantly improve survival in a rat model of acute hepatic failure induced by D-galactosamine . This report explores the mechanism by which hepatocyte transplantation reverses the toxin-induced hepatic necrosis . Radioautographic studies indicated that intraperitoneally administered hepatocytes labelled with tritiated thymidine did not repopulate the injured recipient liver . Hepatocytes irradiated with 10 000 rad (i.e., the cells were nonreplicating) also resulted in a significant (P less than 0.001) increase in animal survival when given to rats treated with D-galactosamine . Experiments with subcellular fractions of hepatocytes demonstrated that an intact cell was not required and that a heat stable "factor" (or factors) present in the cytosol fraction, which is not insulin or glucagon, is responsible for the increase in survival observed . This factor appears to increase the rate of endogenous regeneration of the injured recipient liver. Int Arch Allergy Appl Immunol, 1981, 64(1), 60 - 66 Identification and functional characterization of monocytes in rheumatoid synovial fluid; Thorsteinsson L et al.; Mononuclear cell suspension from synovial fluids of 13 patients with rheumatoid arthritis and 5 patients with juvenile rheumatoid arthritis contained on average of 1.5 and 1.3% cytotoxic plaque-forming cells, respectively, when tested against a monolayer of antibody-sensitized sheep erythrocytes . The plaque-forming ability was completely abolished after removal of cells which had phagocytosed carbonyl iron . The highest proportion of plaque-forming cells was found in the cell suspensions that contained the highest percentage of peroxidase-positive mononuclear cells . The plaque-forming activity was almost completely inhibited by human IgG . Cell suspensions from patients treated with corticosteroids contained the highest proportions of plaque-forming cells and peroxidase-positive mononuclear cells compared with patients not receiving such treatment . Our findings indicate that the plaque-forming leukocytes in rheumatoid synovial fluid are monocytes. Int Arch Allergy Appl Immunol, 1981, 66(4), 372 - 81 T lymphocyte subpopulations and pokeweed mitogen-induced immunoglobulin synthesis in vitro by mononuclear cells from psoriasis patients; Gu SQ et al.; T-lymphocyte subpopulations and pokeweed mitogen(PWM)-induced immunoglobulin (Ig) synthesis of mononuclear cells in vitro were studied in patients with psoriasis as well as in age- and sex-matched normal blood donors . The frequences of T cells with receptors for IgG (Tg) and T cells with receptors for IgM (Tm) were examined . The IgG and IgM synthesis in mononuclear cell suspensions in the presence of different amounts of PWM was determined . The percentage of total T cells (rosetting with neuroaminidase-treated sheep red blood cells), including high and low affinity T lymphocytes showed no difference between the patient group and controls . The patients with psoriasis had a significantly higher mean proportion of Tg cells than the normal donors whereas there was no significant difference in the proportions of Tm cells between these two groups . The PWM-induced Ig synthesis in the mononuclear cell suspension seemed lower in patients with psoriasis than in controls, the difference being statistically significant when the results were expressed as ratios of Ig amounts present in the supernatants of PWM-stimulated and non-stimulated cultures (index of stimulation). Cell Tissue Res, 1981, 218(1), 59 - 73 Germinal centers and the B-cell system . VI . Migration pattern of germinal-center cells of the rabbit appendix; Opstelten D et al.; The migration pattern of germinal center cells of the rabbit appendix was studied and compared with that of appendix dome cells, spleen cells, thymus cells and thoracic duct lymphocytes . To discriminate T- and B-cell migration pathways, normal or T-cell-depleted rabbits were used as donors . Cell suspensions were labeled in vitro with 3H-leucine followed by intravenous transfer . The migration of labeled cells in lymphoid organs was studied using autoradiography, particular attention being paid to the spleen of the recipient . B-cells from the appendix dome, spleen and thoracic-duct lymph migrate to primary follicles or the corona of secondary follicles via thymus-dependent areas of peripheral lymphoid organs . In contrast, a B-cell subpopulation from the germinal centers of the appendix migrates to the center of splenic primary follicles and into germinal centers . The migration of germinal center cells to splenic follicle centers is not enhanced by specific antigens . The migration properties of B-cells, possibly changing during differentiation, may be instrumental in the two types of immune reactions, i.e., plasma-cell reaction and germinal-center reaction. Scand J Clin Lab Invest Suppl, 1981, 156, 171 - 3 Red cell filterability in diabetes; Isogai Y et al.; A negative pressure red cell filteration technique using polycarbonate sieve (Nuclepore) was devised for the observation of red cell filterability (RCF) in diabetes . A significant increase of HbA1 was suggested to be closely correlated to the decrease of RCF . A greater decrease of RCF was observed in diabetics with advanced retinopathy . The increased 2,3-DPG of red cells has been observed in diabetes, but P50 changed much less than might be expected . Improved RCF was observed when Pentoxifylline was added into red cell suspensions . This suggests that Pentoxifylline will have a good effect on microcirculation where impared circulation is observed. Nippon Seikeigeka Gakkai Zasshi, 1981 Jan, 55(1), 45 - 58 {Experimental studies on ascites tumor of human osteosarcoma in nude mice (author's transl)}; Yamada Y; Six human osteosarcomas were transplanted subcutaneously in the back of nude mice and three of the six were found to be serially transplantable . Two of three transplantable solid tumors were called DOS tumor and KOS tumor . The 4th generation of DOS tumor and the 1st generation of KOS tumor grew to invade the peritoneum and tumor cells were spilt spontaneously into the abdominal cavity . These two cases were transformed to ascites tumor, called DOS-NM ascites tumor and KOS-NM ascites tumor . They were found to be serially transplantable by an intraperitoneal injection of free tumor cells (10-20 x 10(5) cells) . DOS-NM and KOS-NM ascites tumor cells were analysed to have human karyotype and alkaline phosphatase activity . Ascites tumor cells were studied under light and electron microscopes . A large number of elongated and ballooned mitochondria were characteristically seen in the cytoplasm of two ascites tumor cells by electron microscope . Development of rough surfaced endoplasmic reticulum and Golgi's apparatus was poor, and no production of collagen fiber was seen in these materials . There is no report referring to ascites tumor of human osteosarcoma, therefore ascites tumor of osteosarcoma was made deliberately and the following results were obtained . 1) Several blocks of DOS tumor were implanted intraperitoneally into 5 nude mice, and only solid tumors grew in the abdominal cavity without ascites . 2) Minced DOS tumor was implanted subcutaneously close to the peritoneum and 2 out of 6 nude mice developed ascites tumor . 3) Cell suspension of DOS tumor was injected intraperitoneally and 3 out of 4 nude mice developed ascites tumors . As mentioned above, human osteosarcoma was successfully induced to ascites tumor (DOS-NM), which was established earlier compared with solid tumor (DOS) . Characteristics of human osteosarcoma are easily examined with ascites tumor and in vivo screening test of anticancer drugs can be quantitatively done . Therefore ascites tumor is considered to be a valuable experimental model. Ric Clin Lab, 1981, 11 Suppl 1, 109 - 16 Filtrability of whole blood and erythrocyte suspensions under the influence of several anticoagulants; Schroer R et al.; Red cell deformability was frequently estimated by filtration tests . Filtrability tests with whole blood samples reveal striking differences in flow rate depending on the anticoagulant used . To clarify the reasons for this filtrability of whole blood samples and of red cell suspensions with similar packed cell volume were compared under the influence of EDTA-Na2 1.5 mg/ml, heparin 75 U/ml, and citrate Na 0.38% (w/v) . Furthermore, after use in whole blood filtration test filters were examined by scanning electron microscopy (SEM) . Filtration tests with red cell suspensions in presence of the different anticoagulants revealed no significant differences . This suggests that the differences in filtrability of differently anticoagulated whole blood samples are not reflecting differences in red cell deformability but different reactivity of the coagulation or the platelet system . SEM examination of filters after filtration tests with heparinised whole blood samples showed large platelet adhesions covering the filters and blocking the filter pores to a great extent . Since this is not found in case of EDTA-anticoagulated samples this anticoagulation is recommended for filtration tests of whole blood samples, if red cell deformability cannot be examined with platelet-free red cell suspensions. Acta Haematol, 1981, 66(4), 238 - 43 Metabolism of ethanol by human bone marrow cells; Wickramasinghe SN et al.; The ability of human bone marrow cells to metabolise has been studied . After incubation of suspensions of bone marrow cells with {1(-14)C} ethanol (0.1 mg/ml) for 1.5 h, radioactivity was present in the evolved CO(2) and in the cellular lipids . Only a part of the 14CO2 generated was dependent on the presence of contaminating blood cells in the marrow cell suspensions . These findings indicate that one or more cell types in the marrow are capable of oxidising ethanol to acetaldehyde and acetate . Our date raise the interesting possibility that the effects of chronic alcoholism on the bone marrow may not be caused by a direct action of ethanol, but may be related to some intracellular or extracellular changes resulting from the metabolism of ethanol within the marrow cells themselves. Acta Biol Med Ger, 1981, 40(7-8), 973 - 7 Evaluation of a rapid fractionation procedure of rabbit reticulocytes after digitonin or hypotonic hemolysis for the study of enzyme and metabolite distributions; Spengler V et al.; A rapid centrifugation procedure following cell lysis with either digitonin or short hypotonic shock in EDTA containing solutions was evaluated, applicable for investigations of enzyme and metabolite compartmentations in intact rabbit reticulocytes . The application of the digitonin disruption seems to be restricted to cell suspensions containing up to 40% reticulocytes only, whereas the hypotonic lysis can be used with practically pure reticulocytes, too . The distribution of markers revealed that an almost complete cell disruption, sufficient separation into pellet and supernatant fraction and satisfactory preservation of mitochondrial intactness could be achieved under appropriate conditions . The suitability of the proposed method in studies on reticulocyte energy metabolism is further supported by the almost insignificant ATP splitting during the entire procedure. Acta Biomed Ateneo Parmense, 1981, 52(5), 179 - 86 {The use of autologous bone marrow transplants in malignant hemopathies and solid tumors}; Manna A et al.; Bone marrow collected from patients with hematologic malignancies were cryopreserved using DMSO as a cryoprotective agent . The growth kinetics of hemopoietic stem cells frozen to -196 degrees C were monitored immediately after thawing by the semisolid agar CFU-C assay and 2 different methods of cell reconstitution were compared . In the first way, thawed cells were plated after the removal of DMSO by washing the cell suspension, in the second, cell suspensions were cultured after a simple 1:1 dilution of DMSO with medium . The number of CFU-C per 2 X 10(5) cells plated was higher washing out the DMSO in all the groups studied . However, the absolute numbers of CFU-C contained in the whole ampoules after the freezing procedures was approximately the same with both methods . It is concluded that washing the cells only apparently yielded a better cloning efficiency, suggesting that such a procedure led to a higher mature nucleated cell loss with the consequence of a CFU-C concentration . This trend seems particularly evident in the AML and CML patients. Microbios, 1981, 31(125-126), 171 - 81 Phase-shift markers in the genus Bordetella: loss of cytochrome d-629 in phase IV variants; Ezzell JW et al.; The respiratory electron transport chain of Bordetella pertussis was examined in whole cell suspensions using difference spectra obtained at room temperature . Phase I (virulent) strains were found to possess cytochromes a-603, b-560, c-550, d-629 and cytochrome o . Cytochrome c-553, previously reported (Sutherland, 1963) was not detected and was assumed to be masked by the alpha-peaks for c-550 and b-560 . Phase IV (avirulent) strains and C-mode cells (phase I strains grown in the presence of 20 mM MgSO4) were deficient in cytochrome d-629 and appeared to have higher levels of cytochromes a-603, b-560, c-550 and cytochrome o . Preliminary data indicate that B . parapertussis and B . bronchiseptica have electron transport chains similar to that of B . pertussis. J Immunol Methods, 1981, 47(3), 295 - 302 Detection of H-2 and Sendai virus antigens by chemiluminescence; Peterhans E et al.; Two monoclonal antibodies reacting with Ia9 and H2.2, respectively, have been added to spleen cell suspensions prepared from mice of different H-2 haplotypes . Cellular light emission was monitored in a liquid scintillation spectrometer operated in the out-of-coincidence mode . The antibodies stimulated chemiluminescence (CL) in cells possessing the target antigenic determinant, but were inactive in cells lacking the determinant . In addition, CL could be stimulated in Sendai virus-infected spleen cells with anti-Sendai antibodies . The results demonstrate that CL measurement is a sensitive and simple method for the detection of cell surface antigens and for the screening of antibodies reacting with these antigens. Histochemistry, 1981, 73(1), 131 - 6 A quantitative approach to trace the corticotrophs in culture after adrenalectomy; Rappay G et al.; Anterior pituitaries of adrenalectomized and sham operated adult rats were dispersed by trypsin and cultured for 4 and 8 days . Adrenalectomy caused a moderate increase in number of corticotrophs in both zero-time cell suspensions and cultures . There was a pronounced elevation of immunoreactive ACTH content in both cells and media and an enhanced secretory response to stimulation of cultures with stalk-median eminence extract containing cortiocotropin releasing (CRF) activity . Some cells identified as corticotrophs by a specific immunostaining incorporated tritiated thymidine into their nuclei suggesting their ability to enter the cell cycle . The relatively smaller increase in number of ACTH cells and the considerably higher ACTH producing capacity of the corticotrophs after adrenalectomy seem to be inconsistent with the quantal response model of hormone secretion recently introduced by Rodbard. Dev Pharmacol Ther, 1981, 2(1), 55 - 66 Direct phosphorylation effects of epinephrine on the plasma membranes of intact rat fat cells; Kang ES et al.; Collagenase-treated isolated cells, prepared from epididymal fat pads of Sprague-Dawley rats exposed to 32Pi or (gamma-32P)-ATP, result in differences in label incorporation into peptides as determined by autoradiography of dried SDS polyacrylamide gels . 32Pi-labelled cells respond to epinephrine by increase in labeling of a 67,000-dalton band, presumably the activated lipase . (gamma-32P)-ATP exposed cells gave a dose-dependent increase in a 53,000-dalton band, a finding shared by cells exposed to cAMP in the absence of epinephrine . However, whereas, cAMP also significantly increased the labeling of an 18,000-dalton band, epinephrine had only a minor effect in the labeling of the 18,000-dalton component . Also, the degree of labeling of a 42,000-dalton band is diminished after epinephrine compared to unstimulated cells . By contrast, cAMP does not affect the labeling of the 42,000-dalton component . The localization of the 53,000- and the 18,000-dalton peptides as well as the enzyme(s) that catalyze their phosphorylation on the external surface of the fat cell is supported by studies using a number of macromolecular probes as well as by subcellular fractionation studies . The absence of such phosphopeptides or kinase activity in the infranatants of such cell suspensions eliminates the possibility that these phenomena are the result of leakage of cytoplasmic components . Thus, epinephrine appears to have effect and do not appear to be explicable simply by the release of cAMP to the extracellular compartment. J Invest Dermatol, 1981 Jan, 76(1), 53 - 5 Normal and psoriatic keratinocytes and fibroblasts compared in culture; Baden HP et al.; The ability of psoriatic fibroblasts to stimulate growth of epidermal cells was studied by comparing the capacity of murine 3T3 cells, normal fibroblasts and psoriatic fibroblasts to act as feeder layers for cultured normal human keratinocytes . 3T3 cells consistently gave shorter times to confluency than normal or psoriatic cells which were about the same . The SDS polyacrylamide gel electrophoretic pattern of the fibrous protein was identical irrespective of the feeder layer use . Psoriatic epidermal cells could be grown from single cell suspensions by using 10(-9) M cholera toxin in the medium . The cultured psoriatic keratinocytes grew identically to normal cells and made the same fibrous protein . The psoriatic keratinocytes also grew better on 3T3 cells than normal or psoriatic fibroblasts. J Natl Cancer Inst, 1981 Jan, 66(1), 85 - 7 A modified enzymatic technique for production of cell suspensions of a murine fibrosarcoma; Penning JJ et al.; Collagenase, elastase, and mild mechanical procedures were used to obtain numerous viable single cells in suspension from a murine fibrosarcoma of C57BL/6 origin . This technique produced suspensions with minimal debris . The suspensions were more than 90% viable and yielded over 1.25 x 10(8) cells/g of solid tumor. Scan Electron Microsc, 1981, (Pt 2), 83 - 6, 82 Mounting of biological microsamples on protein coats for TaOTO non-coated scanning electron microscopy; Ohtsuka A et al.; Gelatin-coated glass cover slips were immersed in a mixture of glutaraldehyde and tannic acid, and rinsed in bovine or human serum . The gelatin-coated cover slips were first placed in a human blood cell suspension, and subsequently placed in a solution of glutaraldehyde or osmium tetroxide . This preliminary mounting of the blood cells eliminated troublesome centrifugations in the tannin-osmium-thiocarbohydrazide-osmium (TaOTO) impregnation . The blood cells seldom detached from the cover slips even when the specimens were dehydrated with ethanol and critical point dried with liquid carbon dioxide . Both the mounting media as well as the mounted cells were intensely stained by the TaOTO method, and exhibited no charging in the scanning electron microscope even at acceleration voltages of 25 kV . The scanning images of the white blood cells with their microvilli were clearly visible with sharp contrast. Cancer Chemother Pharmacol, 1981, 6(3), 273 - 7 Review of experience with interferon and drug sensitivity testing of ovarian carcinoma in semisolid agar culture; Epstein LB et al.; Studies were performed with a semisolid agar culture system to determine the in vitro sensitivity of human ovarian carcinoma to human leukocyte interferon (IFN-alpha) and standard chemotherapeutic agents (adriamycin, cis-platinum, hexamethylmelamine, melphalan, and velban) . Growth in culture occurred in 67% of samples derived from ascitic fluid and 71% of these exhibited reduction in tumor colony number by greater than or equal to 25% in response to 300 units interferon/ml . The responsiveness of the ascitic fluid samples to interferon ran parallel to the overall responsiveness to the other agents tested . Sensitivity to interferon was not related to the histology or grade of the tumor or to the stage of the disease . As compared with cell suspensions prepared from ascitic fluid samples, solid tumor samples had markedly lower viability, 39% vs 89%, and had more tumor cells, 81% vs 28% . Also, colonies derived from solid tumor samples were less sensitive to interferon and more sensitive to cis-platinum and adriamycin than were ascitic fluid-derived colonies . Three of four ascitic fluid samples showed a reduction in tumor colony number of greater than or equal to 25%, whereas none of the solid tumor samples obtained from the same donors were affected by interferon to that degree . Retrospective analysis of drug testing (exclusive of interferon) for in vitro: in vivo correlations revealed that in 58% of 12 evaluable situations, when a single drug (or at least one of a group of drugs) gave a positive response in vitro, stabilization or regression of the tumor occurred in vivo after treatment with the drug . These studies prove the utility of the semisolid agar culture system for assessing the antiproliferative effects of interferon against ovarian carcinoma, and will be of utility in the future for assessing whether various types of interferon have the same degree, range, and mechanism of action of antitumor effect. Scand J Immunol, 1981, 13(6), 605 - 8 Production of immune interferon (type II) in cocultures of mouse peritoneal macrophages and syngeneic tumour cells; Olstad R et al.; Mouse peritoneal macrophages were activated by coculture with syngeneic methylcholanthrene-induced sarcoma cells {7}, which grow as an ascites tumour in C3D2 mice {3} . During 4-5 days of cocultivation tumour cells progressively died, leaving highly activated macrophages . Cell-free supernatant harvested from the cultures contained larger amounts of interferon than either macrophages or tumour cells cultivated alone . Peak activity of interferon occurred on day 2 . Non-adherent cells cultivated together with tumour cells did not produce interferon . Removal of all non-adherent cells from macrophage cultures and host cells from tumour cell suspension did not abolish interferon production . The macrophages thus seem to be the interferon-producing cells, but the possibility that the very few remaining lymphocytes may cooperate with the macrophages in interferon production cannot be totally ruled out . The interferon produced could not be inactivated by antibodies against virus-induced interferon and was destroyed by treatment at pH 2, indicating that the interferon was not of the alpha or beta type. Arch Dermatol Res, 1981, 271(3), 245 - 57 Changes in suspensions of human keratinocytes due to trypsin; Barton SP et al.; Epidermal cell suspensions derived from keratotomed samples of normal human skin have been studied by light microscopy and by scanning and transmission electron microscope techniques . The method allows detailed inspection of the keratinocyte membranes . However, the trypsin used to obtain the suspensions limits the usefulness of the technique as some of the features observed - including invagination of desmosomes, vacuolation, and redistribution of tonofibrils - may be attributable to this proteolytic enzyme. Arch Dermatol Res, 1981, 271(1), 73 - 82 Pemphigus, pemphigoid, and epidermal upper-cytoplasmic antigens: changes in expression in cultured human keratinocytes; Faure M et al.; In an approach of epidermal differentiation, the expression of pemphigus, bullous pemphigoid, and upper-cytoplasmic epidermal antigens was studied in human keratinocytes in culture . The cells were cultured without feeder cells, dermal tissue, or collagen at an acid pH (5.6--5.8) similar to that of the surface of the skin in vivo . Cell suspensions from fresh trypsinized skin and primary, secondary, and tertiary cultures were tested by indirect immunofluorescence for the presence of each antigen using human sera from patients with pemphigus, bullous pemphigoid, and human sera with antibodies against upper-cytoplasmic antigens . Normal sera and cultured human normal fibroblasts and melanoma cells were used as controls . Pemphigus and pemphigoid antigens were found to be expressed, and synthesized by keratinocytes in vitro . The expression to upper-cytoplasmic antigens decreased with time in culture, and they were absent in secondary or tertiary cultures, while expressed by 45--65% of cells prepared from fresh skin . Both upper-cytoplasmic and pemphigoid antigens can be used to type subpopulations of human epidermal cells; however, these findings suggest that epidermal differentiation in vitro differs from that which occurs in vivo. Carcinogenesis, 1981, 2(7), 581 - 7 Metabolism of benzo{a}pyrene and DNA adduct formation in cultured human epidermal keratinocytes; Theall G et al.; Cultured human epidermal cells which require no feeder layer were used to study metabolism of benzo{a}pyrene (BP) and DNA adduct formation . The cultures were prepared from a single cell suspension and maintained at a pH of 5.9--6.2 . At 2 microM BP some cell toxicity was observed, and substantial cell death occurred at 4 microM BP . The metabolism and DNA binding of BP were followed from 6 to 48 h of incubation . High pressure liquid chromatography (h.p.l.c.) revealed that BP was metabolized into 9,10-diol, 7,8-diol, quinones, phenols and tetraols of BP . The DNA binding levels increased linearly up to 28 h of incubation . At 18 h, the level of DNA binding at 0.4 microM BP was 1.5 x 10(-6) mol BP/mol DNA and increased to 6.0 x 10(-6) mol BP/mol DNA at a dose of 4 microM BP . Analysis of the DNA adducts by h.p.l.c . indicates that the 2'-deoxy-N2-(7,8,9,10-tetrahydro-7 beta-,8 alpha, 9 alpha-trihydroxybenzo{a}pyrene-10-yl) guanosine was the predominant adduct formed in cells exposed to BP . The prevalence of the minor DNA adducts varies as a function of the source of the primary skin cells . These results confirm that human cells with no feeder layer metabolize BP and the resultant DNA damage is similar to that found in other mammalian systems. Acta Virol, 1981 Jan, 25(1), 10 - 8 Cell-mediated immunity in flavivirus infections . I . Induction of cytotoxic T lymphocytes in mice by an attenuated virus from the tick-borne encephalitis complex and its group-reactive character; Gajdosova E et al.; The lytic activity of splenocytes from C3H mice immunized with a highly attenuated line of Langat virus {tick-borne encephalitis (TBE) complex} was determined by the in vitro 51Cr release assay on TBE virus (western subtype)-infected target L929 cells . After the spleen cell suspensions were depleted of T cells with anti-mouse theta serum, or of B cells with anti-mouse immunoglobulin, the cytotoxic effect was found dependent on the presence of T lymphocytes . During immunization with Langat virus, cytotoxic T lymphocytes were generated, specific for both the foreign virus-specified antigen and the self component of the host system studied, coded by the K allele of the H-2 complex . The peak of T lymphocyte cytotoxic response was attained on the 6th day after administration of the live virus . T lymphocytes from mice, single-shot immunized with a flavivirus, displayed distinct cross-reactive lysis when studied on target cells infected with other flaviviruses but not on cells infected with an alphavirus. Acta Derm Venereol, 1981, 61(1), 58 - 61 Dissociation of suction blister roof epidermis with trypsin and desoxyribonuclease into viable single cells; Kariniemi AL et al.; Human epidermis, obtained in vivo by the suction blister method, was dissociated with trypsin and desoxyribonuclease into a single-cell suspension . Autoradiographic analysis of the blister roof epidermis and of the epidermal cell suspension was performed to show that neither the suction procedure nor the enzymatic dissociation affected DNA synthesis of the epidermal cells. J Supramol Struct Cell Biochem, 1981, 15(2), 169 - 76 Release of erythropoietin from macrophages by treatment with silica; Rich IN et al.; An erythropoietic stimulating factor (ESF) can be shown to be released from preincubated macrophage-containing cell suspensions from mice by the macrophage-specific, cytotoxic agent, silica . A concentrated silica-treated spleen cell supernatant containing ESF is shown to cause a dose-dependent increase in 59Fe incorporation into red blood cells using the in vivo polycythemic mouse bioassay . The ESF from the same supernatant can also be neutralized by anti-erythropoietin . A second concentrated supernatant fractionated using wheat germ lectin-Sepharose 6MB and compared to either unfractionated or fractionated step III erythropoietin (Ep), tested in vitro using the erythroid colony-forming technique and 12-day fetal liver as target cells, indicates parallelism of all linear dose-response lines . This, together with the in vivo data, strongly suggests that the ESF released from macrophages treated with silica is, in fact, Ep . Substituting Ca2+ ions for fetal calf serum in the preincubation procedure results in the same activity being released compared to the presence of 1% or 20% fetal calf serum. Biomedicine, 1980 Dec, 32(4), 185 - 8 Detection of lymphocyte stimulation by flow cytometry: differences in cells from patients with and without neoplasia; Hartmann W et al.; Fluorescence polarization measurements were performed to detect PHA-stimulation of peripheral lymphocytes in an early state . Single cell fluorescence was measured by flow cytometry using an epi-illumination design to avoid the influence of background fluorescence and multiscattering . By these arrangements the accuracy in the determination of P, the degree of polarization, could be improved over the usual cell suspension measurements . Fluorescence polarization measurements were performed on density specified human lymphocytes after incubation with fluorescein diacetate with and without preincubation with phytohemagglutinin (PHA) . A group of healthy donors had an average polarization of P = 0.17 before and P = 0.12 after PHA-stimulation, whereas a group of patients with malignant disease showed an average polarization of P = 0.14 before and after PHA-stimulation. Biochim Biophys Acta, 1980 Dec 1, 633(2), 162 - 75 Elicitor modulation of the turnover of L-phenylalanine ammonia-lyase in French bean cell suspension cultures; Lawton MA et al.; (1) The mechanisms underlying the transient increase in phenylalanine ammonia-lyase activity during phaseollin accumulation in cell suspension cultures of Dwarf French bean (Phaseolus volgaris) have been investigated using density labelling with 3H from 2H2O coupled with residual analysis of the equilibrium distribution of enzyme activity in high-resolution KBr density gradients . (2) The resolution achieved in this system is sufficient to allow quantitative analysis of the relative proportions of light, unlabelled, pre-existing enzyme and heavy, labelled, newly synthesised enzyme . (3) Elicitor released by heat treatment of cell walls of Colletotrichum lindemuthianum, the causal agent of anthracnose disease of French bean, caused a marked but transient increase in phenylalanine ammonia-lyase activity concomitant with the onset of phaseollin accumulation in the bean cultures . The induction of enzyme activity was highly dependent on elicitor concentration, with maximum induction occurring in two discrete concentration ranges; at an intermediate elicitor concentration, or at supra-optimal elicitor concentrations, no enzyme induction was observed . (4) At low concentrations of elicitor the induction of enzyme was entirely a result of elicitor stimulation of the rate of de novo enzyme production . In contrast, at higher elicitor concentrations the increase in enzyme activity was accompanied by a marked apparent stabilization of the enzyme in vivo, and the rapid but transient increase in enzyme activity was achieved by a programme of reciprocal changes in the rate constant for de novo enzyme production and the rate constant for removal of enzyme activity . Such reciprocal control of the rates of enzyme production and removal may be crucial in determining the magnitude and duration of the phytoalexin defense response . (5) Information on the specific activity of 2H label in the amino acid pools was obtained from analysis of the equilibrium distribution of residual, labelled activity . This showed directly that the turnover of the amino acid pools was fast relative to the turnover of phenylalanine ammonia-lyase and, hence, the rate of enzyme labelling was not limited by the rate of labelling of the amino acid pools . Elicitor treatment had no effect on the specific activity of label in the amino acid pools from which phenylalanine ammonia-lyase is synthesised. Blood, 1980 Dec, 56(6), 1111 - 9 Cytochemically demonstrable B-glucuronidase activity in normal and neoplastic human lymphoid cells; Machin GA et al.; Mononuclear cell suspensions were prepared from 40 normal peripheral blood and lymphoid tissue specimens and 42 neoplastic specimens obtained from patients with malignant lymphoma and lymphocytic leukemia . These suspensions were analyzed for la antigens, surface immunoglobulin (Slg), sheep erythrocyte (E) rosette formation and, in some instances, acid alpha-naphthyl acetate esterase (ANAE) activity . The results of these studies were correlated with the expression of cytochemically demonstrable BG activity . The percentage of BG+ lymphocytes was found to be comparable, within 10%, to the percentage of E+ (T) cells in the majority of normal, non-neoplastic peripheral blood, tonsil, spleen, and lymph node specimens examined . Occasionally, the percentage of E+ cells exceeded the percentage of BG+ cells by 20% or more, suggesting the presence of an E+BG- T cell subpopulation . BG+ B lymphocytes were only demonstrated in 1 of 40 non-neoplastic lymphoid specimens . The neoplastic B cells in each of 14 B cell (la+Slg+E-) lymphomas were BG- . However, a variable proportion of the neoplastic cells isolated from 6 cases of B cell chronic lymphocytic leukemia and neoplastic plasma cells isolated from 7 cases of multiple myeloma expressed BG activity . Thus, it appears that both normal and