Cell suspensions

Eur J Immunol, 1975 Apr, 5(4), 274 - 81
Antigens on mouse and rat lymphocytes recognized by rabbit antiserum against rat brain: the quantitative analysis of a xenogeneic antiserum; Morris RJ et al.; Quantitative assays for measuring the binding of xenogeneic antiserum to dispersed cell suspensions are described . Cells were incubated with unlabeled xenogeneic antiserum and the antibody bound measured indirectly by a second binding step with 125I-labeled anti-immunoglobulin antibody . This indirect radioactive binding assay was calibrated by measuring, with a radioimmunoassay, the true amount of antibody bound in the first step . With these methods one can measure the strength of antisera, quantitate the number of antigenic sites, and partially differentiate determinants being recognized on cell surfaces . The binding of rabbit anti-rat brain antiserum to rat and mouse lymphocytes was analyzed in detail . After absorption of the antiserum with rat liver, the antibody remaining recognized lymphocyte antigens that were distributed among various rat and mouse tissues in quantities identical to Thy-1.1 antigen . Thus, at saturation, 670 000 Ig molecules from liver-absorbed rabbit antiserum were bound per rat thymocyte, and the antiserum bound to 90%, 21%, 4% and 2% of rat thymocytes, spleen, lymph node and thoracic duct lymphocytes, respectively . With mouse tissues, 90% 24% and 50% of thymocytes, spleen and lymph node cells, respectively, were labeled . In rat brain the concentration of xenoantigen increased with age, while in thymocytes the full adult amount was present at birth . Three antigenic determinants could be defined with the liver absorbed rabbit antiserum: the Thy-1.1 antigen, a rat specific antigen and an antigen cross-reacting between rat and mouse tissues . All 3 may be on the Thy-1 molecule . The anti-brain antiserum contained about 0.05 mg/ml of antibody specific to these xenoantigens.

Naunyn Schmiedebergs Arch Pharmacol, 1975 Mar 25, 287(2), 141 - 56
Histamine release, formation of prostaglandin-like activity (SRS-C) and mast cell degranulation by the direct lytic factor (DLF) and phospholipase A of cobra venom; Damerau B et al.; Cobra venom, alone and in combination, on mast cell degranulation, histamine release and formation of prostaglandin-like activity (SRS-C) was studied in perfused guinea-pig lungs and in mast cell-containing rat peritoneal cell suspensions . For comparison, the effect of equivalent doses of whole cobra venom was investigated . 1 . Cobra venom caused mast cell degranulation, histamine release and SRS-C formation in both systems . For comparable effects much higher doses had to be used in guine-pig lungs than in rat peritoneal cell suspensions . 2 . Phase A showed little degranulation of mast cells in both systems, a limited histamine release in rat peritoneal cell suspensions and none in perfused guinea-pig lungs . It caused a considerable SRS-C formation in both, lung tissue and peritoneal cell suspensions . 3 . DLF caused histamine release, SRS-C formation and mast cell degranulation in both systems; in rat peritoneal cell suspensions it acted almost as strong as equivalent doses of cobra venom, in guinea pig lungs it was much less active . 4 . In rat peritoneal cell suspensions the effects of DLF and phase A in combination did not exceed the sum of their single effects . In guinea-pig lungs these two substances interacted in a potentiating synergism . It is concluded that DLF is the main cytotoxic principle of cobra venom, whereas ph-ase A alone is not cytotoxic . The difference in the synergism of DLF and ph-ase A between rat peritoneal cells and guinea-pig lungs may be due to two different actions of DLF and species differences as regards sensitivity against these actions.

Eur J Biochem, 1975 Mar 17, 52(2), 311 - 20
Isoenzymes of p-coumarate: CoA ligase from cell suspension cultures of Glycine max; Knobloch KH et al.; Two isoenzymes of p-coumarate: CoA ligase were isolated from cell suspension cultures of soybean (Glycine max L., var . Mandarin) . Separation and partial purification of the enzymes were achieved by precipitation with MnCl2 and (NH4)2SO4, and column chromatography on DEAE-cellulose, Sephadex G-100 and hydroxyapatite . The isoenzymes had approximately the same molecular weight, but differed significantly with respect to their substrate specificity, their inhibition constants for AMP, their dependence on pH and ionic strength for optimum activity, and their fractionation pattern during the purification procedure or upon analytical disc-gel electrophoresis . Both coumarate: CoA ligases were specific for the activation of various substituted cinnamic acids . Of the cinnamic acids tested, ferulic, sinapic, 5-hydroxyferulic, p-coumaric, and caffeic acids were the substrates with the lowest apparent Km values (on all the order of 1 to 4 x 10(-5) M) for isoenzyme 1 . The lowest apparent Km values (from about 1 to 9 x 10(-5) M) for isoenzyme 2 were obtained for caffeic, p-coumaric, m-coumaric, and o-coumaric acids . Sinapic acid and several methoxycinnamic acids were efficient substrates of isoenzyme 1 but were not activated at all by isoenzyme 2 . The possible roles of the two p-coumarate: CoA ligase isoenzymes in the phenylpropanoid metabolism of the cell cultures are discussed.

Eur J Biochem, 1975 Mar 17, 52(2), 265 - 71
Bacterial respiration-linked proton translocation and its relationship to respiratory-chain composition; Jones CW et al.; 1 . The relationship between chain composition and the efficiency of respiration-linked proton translocation was studied in nine bacterial species of widely differing taxonomic and ecological status . 2 . All the bacteria investigated contained respiratory chain dehydrogenases, ubiquinone and/or menaquinone, cytochrome b and cytochrome oxidase aa3 and/or o . In addition, some of these organisms also contained pyridine nucleotide transhydrogenase and/or cytochrome c . 3 . leads to H+/O ratios of whole cell suspensions oxidising endogenous substrates were in the approximate range 4-8 mol H+ translocated per g-atom oxygen consumed . It was concluded from the observed leads to H+/O ratios of cells loaded with specific substrates that proton-translocating loops 1 and 2 were present in all of the organisms investigated, but that loops 0 and 3 were dependent upon the presence of pyridine nucleotide transhydrogenase and cytochrome c respectively . 4 . The wide range in energy conservation efficiency which was observed in these organisms is discussed in relation to their respiratory chain composition and natural habitat.

Biochim Biophys Acta, 1975 Mar 14, 385(1), 81 - 7
NMR study of -17-O from H2-17-O in human erythrocytes; Shporer M et al.; Human erythrocytes were incubated in a Ringer's solution enriched with 10--18 per cent H2-17-O . The longitudinal relaxation time (T1) of the -17-O was determined separately in samples of red cell suspensions, packed cells, and supernatant . The longitudinal relaxation of -17-O in erythrocyte suspensions was non-exponential, reflecting water exchange across the cell membranes as well as relaxation processes inside and outside the cell . The T1 of intracellular -17-O is 4--5 times shorter than in the supernatant, similar to the enhancement of proton relaxation by hemoglobin in erythrocytes and free solution at the frequency applied (8.13 MHz) . This datum is consistent with tht thesis that hemoglovin modifies the NMR relaxation behavior of water inside cells and in free solution in the same way . The rate constant (kx) for water exchange was calculated to be 60 and 107 s- minus 1 at 25 and at 37 degrees C, respectively . The apparent activation energy for kx over the temperature range 23--37 degrees C was 8.7 plus or minus 1.0 kcal/mole.

Biochim Biophys Acta, 1975 Mar 14, 385(1), 88 - 100
Stimulatory effect of gonadotropins on the synthesis of adenosine 3': 5'-cyclic monophosphate and progesterone by suspensions of rat ovarian interstitial cells; Kawano A et al.; A cell suspension was prepared from immature rat ovaries by treatment with trypsin and collagenase . The isolated cells were capable of converting {8-14-C}adenine to cyclic {-14-C}AMP and the rate of this conversion was stimulated in vitro by luteinizing hormone and human chorionic gonadotropine, but not by prolactin, norepinephrine, dopamine or albumin . The accumulation of progesterone was also measured in these cells by radioimmunoassay . In vitro addition of luteinizing hormone and human chorionic gonadotropine, but not by prolactin, norepinephrine, dopamine or albumin . The accumulation of progesterone was also measured in these cells by radioimmunoassay . In vitro addition of luteinizing hormone stimulated the accumulation of radioimmuno-assayable progesterone . The conversion of {8-14-C}adenine to cyclic {-14-C}AMP showed a rapid increase during the first 30 min of the incubation period when luteinizing hormone was added to the incubation medium . Progesterone accumulation in response to the same dose of luteinizing hormone showed a lag period for the first 30 min of incubation after which there was an increase up to 2 h . The luteinizing hormone-induced progesterone accumulation was sensitive to puromycin, but there was no effect on the luteinizing hormone-induced increase in cyclic {-14-C}AMP formation from {8-14-C}-adenine . Actinomycin D also inhibited the luteinizing hormone-induced progesterone accumulation in rat ovarian interstitial cell suspension is preceded by an increased accumulation of cyclic AMP and that the accumulation of steroid under the influence of luteinizing hormone involve processes sensitive to puromycin and antinomycin D.

Cancer, 1975 Mar, 35(3), 580 - 7
Further observations of Fc receptors in human malignant tissue and normal lymphoid tissue; Tonder O et al.; Twenty human malignant solid tumors of various histologic types were tested for the presence of Fc receptor using cryostat sections or single cell suspensions of fresh tissue . Sheep erythrocytes sensitized by various amounts of rabbit IgG antibodies served as indicator cells (EA) . All tumors possessed Fc receptor, but to varying degrees; eight reacted more strongly than normal spleen without any relation to histologic type . The tumors which gave the strongest reactions in sections also formed the highest percent of EA rosettes in suspensions, thus indicating surface localization of receptors . The reactions with spleen sections localized to the B cell and monocytic areas; the latter also showed high avidity in reactions with uncomplexed IgG . Rabbit antisera to tumors, spleen, and peripheral lymphocytes (polyvalent ALS) in inhibited the reactions, while a T-cell-specific ALS did not . Absorptions of the antisera with lymphocytes or tissue sediments of spleen and tumors removed the inhibiting activity, tissue sediments of muscle and kidney only reduced the titers . Again, results with spleen sections paralleled those obtained with tumor sections . Apparently, the tumor Fc receptor is very similar to the Fc receptors present in normal lymphoreticular tissues.

Clin Exp Immunol, 1975 Mar, 19(3), 459 - 74
The different migratory characteristics of lymphocyte populations from a whole spleen transplant; Parrott DM et al.; Spleens from AS x BN donor rats labelled in vivo by multiple doses of {3H}thymidine were transplanted into syngeneic recipients by anastomosis to the abdominal great vessels . The recipients were killed 1-5 days after receiving the whole spleen transplants and the numbers and location of the {3H}thymidine-labelled cells which had migrated from the labelled donor spleen traced by means of autoradiographs of sections, imprints and smears of various recipient lymphoid tissues . These results were compared with the migration pattern of labelled dissociated spleen cell suspensions injected intravenously . The latter consists almost entirely of small lymphocytes which migrate to T or B areas of recipient spleen, lymph nodes and Peyer's patches . The labelled whole spleens also contained cells which migrated to the T and B areas of recipient lymphoid tissues, but in addition contained many lymphoid cells which migrated to the red pulp of the recipient spleen and to the lamina propria of the gut . These experiments showed, therefore, that the spleen contains mobile elements which have not been detected by transfer of spleen cell suspensions.

J Cell Sci, 1975 Mar, 17(3), 371 - 9
Effects of cytochalasin B on the aggregation, electrophoretic mobility and surface morphology of chick neural retina cells; Jones GE; Over a concentration range of o-5-10 mug/cm-3, cytochalasin B caused a biphasic change in the electrophoretic mobility of disaggregated neural retina cells . An initial rise in anodal mobility at low concentrations of the drug was transformed into a reduction in the mobility below that of the control at a concentration of 10 mug/cm-3 . The effect of cytochalasin B was found to be reversible by washing treated cells in cytochalasin B-free media . This was investigated at a concentration of cytochalasin at which the greatest difference existed between the mobilities of the control and experimental cell suspensions . Reaggregation of cell dispersions failed to show any significant difference in the rate of aggregation between cytochalasin B-treated cells and the control . Scanning electron microscopy of cells fixed while in suspension also showed little significant change in the surface morphology upon application of cytochalasin B . In high concentrations of the drug cells appeared somewhat smoother in outline, but no correlation was found between changes in surface morphology and the variations in cell electrophoretic mobility . It is concluded that the observed changes in electrophoretic mobility may be attributed to a binding of cytochalasin B to the cell membrane . This lends support to the hypothesis that the primary site of action of cytochalasin B may be the plasma membrane.

J Cell Sci, 1975 Mar, 17(3), 337 - 48
Kinetic aspects of synchrony in suspension cultures of Acer pseudoplatanus L; Gould AR et al.; Synchronous divisions have been induced in cell suspension cultures of sycamore (Acer pseudoplatanus L.) initiated at low density from stationary phase, nitrate-starved cells . The pattern of division and DNA replication synchrony is unusual in that it does not show progressive decay but rather a sharp transition to asynchrony . Feulgen densitometry data indicate that in terms of an S-phase synchrony index, DNA replication becomes more highly synchronized as cell density rises and interphase duration is reduced . In comparison with asynchronous cultures the duration of mitosis appears to be reduced during synchronous growth . The persistence of division and S-phase synchrony and the apparent shortening of interphase and mitosis in these sycamore cultures suggests the operation of intercellular entrainment . The departure from synchrony during the interphase which precedes initiation of asynchronous division remains unexplained.

Blood, 1975 Mar, 45(3), 427 - 33
Erythropoiesis in steel mutant mice: effects of erythropoietin in vitro; Chui DH et al.; Adult SI/SI-d mutant mice have severe macrocytic, normochromic anemia . Moreover these animals are unresponsive to the stimulation of erythropoietin in vivo . By means of a bone marrow cell suspension culture system, the present investigation shows that in adult SI/SI-d marrow, there are cells capable of responding in vitro to erythropoietin in a normal fashion . Moreover, the erythropoietin present in SI/SI-d serum is biologically active in vitro without any prior biochemical modification . These observations support the suggestion that there is a defect in differentiation in the erythroid cell lines of SI/SI-d mice in vivo due to an abnormal hemopoietic microenvironment.

Proc Soc Exp Biol Med, 1975 Mar, 148(3), 808 - 12
Failure of peritoneal exudate macrophages to reverse immunologic impairment by Friend leukemia virus; Ceglowski WS et al.; Transfer experiments with peritoneal exudate macrophages from normal donor mice were performed to determine if a defect of normal macrophage function or activity was a major or contributing factor to the immunosuppression characterizing leukemia virus infection of mice . Challenge immunization of Friend leukemia virus-infected mice with sheep erythrocytes resulted in markedly depressed hemolytic antibody responses, as compared to responses of normal noninfected mice . When PE cell suspensions rich in macrophages were transferred from normal donor mice to leukemia virus infected recipients there was no affect on the FLV-induced impairment of the immune response . Similar transfer of PE cells to normal uninfected mice generally resulted in a moderate depression of the expected immune response . In no case did the PE cells enhance the immune responses in normal or virus-infected mice.

Arzneimittelforschung, 1975 Mar, 25(3), 378 - 82
Effect of cytostatic drugs on nucleic acid synthesis in mouse thymocytes and chick embryo cells; Hakansson L; Cell suspensions of either mouse thymocytes or chick embryo cells were incubated with labelled precursors after preincubation with different cytostatic drugs . The total time of in vitro incubation was four hours . The ratio between the amount of incorporated labelled precursors and the total amount of DNA was determined and used to measure the cytostatic effect . All drugs used caused a marked depression of this ratio . The cytostatic drugs had no effect on the phosphorylation of thymidine.

Am J Med, 1975 Mar, 58(3), 307 - 13
Immunologic and morphologic studies of T cell lymphoma; Mann RB et al.; Thymic-independent (B) lymphocytes, thymic-independent (T) lymphocytes and histiocytes may be distinguished by the presence of certain surface markers . In addition B and T lymphocytes have been reported to show distinctive surface architecture by scanning electron microscopy . Neoplastic cells from a lymph node and cerebrospinal fluid of a patient with a diffuse malignant lymphoma of the poorly differentiated lymphocytic type were examined in frozen sections and cell suspensions for the presence of surface immunoglobulin and the antigen-antibody-complement (IgMEAC) receptor of B lymphocytes, the presence of the cytophilic antibody (IgGEA) receptor of histiocytes and the ability to form nonimmune rosettes with sheep red blood cells (E) characteristic of T lymphocytes . Cells from the lymph node were also studied by scanning electron microscopy . The majority of neoplastic cells from the lymph node and cerebrospinal fluid formed rosettes with E, but lacked surface immunoglobulin and failed to bind IgMEAC or IgGEA . By scanning electron microscopy the neoplastic cells, although larger in diameter, showed surface architecture similar to normal lymphocytes with a varying number of surface microvilli . These studies suggest that the malignant cells of this lymphoma are of thymic type.

J Exp Med, 1975 Mar 1, 141(3), 664 - 80
Early cellular events in a systemic graft-vs.-host reaction . I . The migration of responding and nonresponding donor lymphocytes; Atkins RC et al.; A systemic graft-vs.-host (GVH) reaction was initiated by the intravenous injection of parental strain thoracic duct lymphocytes (TDL) into irradiated F1 hybrid recipients with in-dwelling thoracic duct cannulae . The migration of the donor lymphocytes was followed by labeling them in vitro with either {3H} or {14C}uridine and measuring radioactivity by scintillation counting of the spleen and lymph nodes of the recipients removed 24 h after injection and in TDL collected throughout this period . The localization of labeled cells was always compared to that of a reference population of nonreactive lymphocytes, e.g . F1 hybrid, labeled with the alternative isotope (Fig . 1) . A consistent surplus of the reactive label was found in the spleen which was balanced by a deficit of the reactive label in TDL; lymph nodes gave intermediate values . The same distribution pattern was noted when the reference population was a specifically unresponsive population of the parental strain . This differential distribution depends on recognition of the recipient's Ag-B antigens because when normal lymphocytes were injected together with specifically unresponsive lymphocytes into a "third party" F1 hybrid (against which both populations were reactive) there was no surplus of the normal cells in the spleen and no deficit in the lymph . Moreover in an Ag-B identical strain combination there was no detectable difference in the distribution of reactive and nonreactive populations . The distribution of a labeled reaction population can be accounted for if a substantial minority of cells are immobilized in the spleen and lymph nodes as a consequence of antigen recognition (Fig . 3) . When the donor cells in the spleen were assayed 24 h after injection there was paradoxically a slight reduction in their specific GVH activity, which is at least partly because they are under-represented in a single cell suspension . The size of the splenic surplus (23%) and the thoracic duct deficit (12%) suggested that the minority of nonimmune lymphocytes which recognize each Ag-B complex carry 12% of the radioactive label in the original population . It is argued that this provides a near estimate of the frequency of T lymphocytes which can recognize each Ag-B antigenic complex.

Pediatr Res, 1975 Mar, 9(3), 143 - 6
Thymidine kinase activity in cerebrospinal fluid of rabbits with Herpesvirus hominis encephalitis; Hilty MD et al.; The rabbit model of Herpesvirus hominis (HVH) encephalitis was utilized to determine whether thymidine (TdR) kinase (EC . 2.7.1.2.1), AN EARLY ENZYME IN THe HVH replicative cycle, is present in the cerebrospinal fluid (CSF) of rabbits with HVH encephalitis . The mean TdR kinase activity in CSF from infected rabbits was 2.30 plus or minus 1.69 pmol product/120 min/100 mul CSF, while the mean TdR kinase activity of uninfected CSF was 0.34 plus or minus 0.25 . The enzyme appeared to be cell associated since the TdR kinase activity in the sedimented cell suspension was 5.08 plus or minus 2.93 (seven values) while the supernatant activity was 0.14 plus or minus 0.31 (seven values) . To allow a comparison of TdR kinase from infected rabbit CSF with HVH and rabbit mononuclear cell TdR kinase, the mean ratio of TdR kinase activity at pH 6.0/pH 8.0, for infected rabbit CSF is intermediate between the ratio for HVH TdR kinase (P less than 0.001) and rabbit mononuclear cell TdR kinase (P less than 0.05).

Endocrinology, 1975 Mar, 96(3), 792 - 6
A short method for the isolation of somatotrophs from the rat pituitary gland; Snyder G et al.; Application of a density gradient centrifugation procedure for the isolation of somatotrophs from dispersed rat anterior pituitary cells is described . Two sequential discontinuous gradients of bovine serum albumin are used to enrich the somatotroph population from a frequency of 38% in the initial cell suspension to a final purity of 85% . The isolated somatotrophs retain both histochemical and ultrastructural integrity throughout the isolation procedure . The isolated somatotrophs also release immunoassayable growth hormone when incubated with 6 mM dibutyryl 3',5'-cyclic AMP . The method offers the advantages of being both simple and short.

Cancer Res, 1975 Mar, 35(3), 825 - 31
Heritability of the phytohemagglutinin responsiveness of lymphocytes and its relationship to leukemogenesis; Heiniger HJ et al.; Upon analyses of 59 inbred strains, F-1 hybrids, and congenic-resistant mouse strains, the strain distribution pattern of the stimulation of perippheral mouse lymphocytes by phytohemagglutinin was established using a micromethod . The family of DBA mice were the lowest responders of phytohemagglutinin, whereas the C57 family responded best . Strain PL/J exhibited the best response . The response of lymphocytes to the lectin is governed by more than two but less than five major genes of unknown linkage . No direct association to the H-2 histocompatibility complex was found, although an indirect influence of this locus could not be excluded . All high-leukemia strains are good responders to phytohemagglutinin . None of the low-responder-group strains exhibit spontaneous leukemia . No correlation of the response of lymphocytes to the expression of the type C RNA genome could be established . Cell suspensions from animals exhibiting clinical signs of leukemia responded only weakly or not all to the lectin.

Science . 1975 Feb 21;187(4177):656.
Transmissible mink encephalopathy: infectivity of corneal epithelium; Marsh RF et al.; Corneal epithelium from hamsters dying of transmissible mink encephalopathy contained a virus titer of 10-4.8 times the 50 percent lethal dose (10-4.8 LD50) per 0.05 milliliter when assayed as a cell suspension derived directly from the infected animal . After one passage in tissue culture, an equivalent concentration of cells contained only 10-0.8 LD50 per 0.05 milliliter. . It is concluded that corneal tissues are infectious; the infectivity may be mainly associated with free nerve endings . However, the most important immediate inference is that corneas from human beings affected with Creuzfeldt-Jakob disease are likely to be lethal if transplanted to healthy recipients.

Science, 1975 Feb 14, 187(4176), 542 - 4
Toxicology and pharmacological action of anabaena flos-aquae toxin; Carmichael WW et al.; Calves, rats, ducks, and goldfish given lethal oral doses of bacteria-free lyophilized cell suspensions of toxic Anabaena flos-aquae died as a result of respiratory arrest . Experiments with selected animals and pharmacological preparations showed that the main effect of the toxin was production of a sustained postsynaptic depolarizing neuromuscular blockade.

Ann Immunol (Paris), 1975 Feb-Mar, 126(2), 121 - 35
Regulation of lymphocyte responses in vitro . VII . - "Cell contact inhibition", a possible regulatory mechanism for lymphocyte activation; Bernard DP et al.; Potentiation by cytochalasin B (CB) of (3H)thymidine incorporation in lymphoid cell suspensions stimulated by phytohemagglutinin (PHA) is counteracted by adherent cells . The drug has no effect on the production of the macrophage product lymphocyte activating factor (LAF) and does not interfere with its potentiating activity . CB appears to inhibit agglutination of lymphocytes . Other factors (absence of serum, macrophages, mechanical disturbance of the cultures) diminishing the size of aggregates increase the response of lymphocytes . Responses to less agglutinating mitogens are less or not enhanced by CB . Factors increasing agglutination or/and cell contacts (high cell density, wheat germ agglutinin) diminish lymphocyte activation and make it susceptible to CB potentiating effect . The results suggest that topoinhibition (cell contact inhibition) is an important mechanism regulating the lymphocyte response to mitogens in culture . Its effect could be modulated by macrophages and certain serum factors, and CB would potentiate these responses by reducing contact inhibition.

J Appl Physiol, 1975 Feb, 38(2), 315 - 20
A new method for the measurement of percent oxyhemoglobin; Pittman RN et al.; A new method is presented for the spectrophotometric determination of percent oxyhemoglobin (percent saturation) in whole blood . The method is based on a theory of light absorption and scattering by particulate suspensions and requires the measurement of optical densities (D) of blood at three closely spaced wavelengths . The contribution of scattering to the optical density at each wavelength is determined from optical density values at two isosbestic wavelengths (546 and 520 nm) and the optical density at the third wavelength (555 nm) is related to the extent of oxygenation of the hemoglobin . The wavelength independence of the scattering contribution (B) induced by red cells was established from 510 and 575 nm . The optical density of red blood cell suspensions was measured with a spectrophotometer and a linear relationship was found between percent saturation and the corrected optical density ratio, (D555 B)/(D546 B) . For a given saturation, this ratio was independent of optical path length (0.1-2 mm) and hematocrit (3-50%).

Am J Pathol, 1975 Feb, 78(2), 277 - 84
Acceleration of amyloidosis by syngeneic spleen cells from normal donors; Axelrad M et al.; Intravenous administration of syngeneic spleen cells from normal donors was found to markedly shorten the induction time for casein-associated splenic amyloid diposition in the mouse . The effect of intravenous donor cells seemed purely one of acceleration; it did not provoke amyloid deposition either by itself or in combination with independently ineffective heterologous proteins . The accelerator effect did not depend on the viability of the cell suspension, and after physical disruption of the cells all the accelerator activity seemed localized to the sedimentable fraction of damaged cells, nuclei and coarse debris . It is suggested that the accelerator acts through affecting the function of perifollicular splenic macrophages.

J Gen Microbiol, 1975 Feb, 86(2), 319 - 26
Studies on the physiological significance of the lack of a pyruvate dehydrogenase complex in Hyphomicrobium sp; Harder W et al.; Hyphomicrobium X was grown in media containing either methanol or ethanol as a carbon and energy source, with or without additional organic carbon sources . The organism transported pyruvate, malate and succinate into the cells, and incorporated their carbon skeletons into cellular material, but when each of these compounds was added as sole carbon and energy source none supported growth of the organism . Enzymic analysis of crude cell-free extracts failed to detect either a complete pyruvate dehydrogenase complex or an active E1 component . Furthermore, oxygen uptake experiments with whole cell suspensions did not show any oxidation of pyruvate, succinate or malate . The distribution of radioactivity amongst the amino acids in hydrolysates of cell protein obtained from organisms grown in the presence of {14C}pyruvate, {14C}acetate or {14C}succinate indicated that the organism is limited in its ability to metabolize pyruvate . Growth in the presence of {14C}pyruvate resulted in 93% of the total radioactivity recovered being associated with amino acids derived directly from pyruvate . In contrast, growth in the presence of {14C}acetate or {14C}succinate resulted in more-or-less uniform labelling of all biogenic classes of amino acids . These results are consistent with the lack of an active pyruvate dehydrogenase complex which would make it impossible for Hyphomicrobium X to convert pyruvate into acetyl-CoA and to generate energy from carbon compounds for which the energy metabolism relies on oxidation through tricarboxylic acid (TCA) cycle intermediates.

Br J Haematol, 1975 Feb, 29(2), 293 - 300
Blood leucocyte enzymes . II . Activities at 8-9 a.m . in cells of normal subjects, chronic lymphatic leukaemia and chronic myeloid leukaemia patients; Brok-Simoni F et al.; The activities of glucose-6-phosphate dehydrogenase (D-glucose-6-phosphate: NADP oxidoreductase, G6PD), 6-phosphogluconate dehydrogenase (6-phospho-D-gluconate: NADP oxidoreductase, 6PGD), hexokinase (ATP: D-hexose 6-phosphotransferase, Hx), lactate dehydrogenase (D-lactate: NAD oxidoreductase, LDH) . glutamate oxaloacetate transaminase (L-aspartate: 2 oxoglutarate aminotransferase, GOT) and dihydrofolate reductase (DHFR) were measured at 8 a.m . in leucocytes of healthy individuals and patients with chronic myeloid leukaemia (CML), chronic lymphatic leukaemia (CLL), myelofibrosis with myeloid metaplasia and polycythaemia vera . In view of the heterogeneity of the leucocyte populations in these conditions, the enzyme activities were correlated to the number of immature cells in CML and to the percentage of lymphocytes in CLL . No differences in the enzyme activities were found between the white cells of healthy individuals, myelofibrosis with myeloid metaplasia and polycythaemia vera . In CML the activities of all enzymes except GOT correlated directly with the number of immature cells; an inverse correlation with the number of lymphocytes was observed in CLL . GOT was the only enzyme whose activity correlated with the number of lymphocytes in the cell suspension . Furthermore, a significantly higher activity of this enzyme was found in Ficoll-isolated CLL lymphocytes as compared to normal lymphocytes.

J Lab Clin Med, 1975 Feb, 85(2), 329 - 36
Determination of hemoglobin-oxygen affinity on micro samples; Lyonnais J et al.; The spectrophotometric determination of hemoglobin-oxygen dissociation on dilute red cell suspensions has been modified by the use of a bicarbonate buffer containing bovine serum albumin . The red cell suspension is equillibrated with known ratios of air and nitrogen and 5 per cent carbon dioxide at atmospheric pressure using two gas pumps . The method enables the hemoglobin-oxygen dissociation curve to be measured on 10 to 20 mul of whole blood, and has given values for the P50 (the oxygen tension at 50 per cent hemoglobin-oxygen saturation) of 25.9 plus or minus 1.5 S . D . which are comparable to the value of 26.0 plus or minus 1.0 S . D . obtained by the mixing technique and by other methods . The addition of bovine albumin (35 mg . per cent or greater) increased the P50 by 3 mm . Hg above that obtained with buffer alone . The P50 value has been compared in bis-Tris and phosphate buffers with and without albumin, and in bicarbonate buffers with the addition of approximately equimolar concentrations of either human or bovine serum albumin or polyvinyl-pyrrolidone (PVP), and buffer alone . The effect of albumin on the P50 value was most marked in bicarbonate buffer, and was statistically significantly greater in the presence of human or bovine albumin than in buffer alone or buffer plus PVP . The addition of albumin could not be explained through an alteration of cell shape or volume, but may be an influence on intracellular pH.

Endocrinol Jpn, 1975 Feb, 22(1), 17 - 23
Effect of temperature during preparation of rat adrenal quarters and isolated cell suspension of their ACTH responsiveness; Morita Y et al.; The effect of temperature during preparation of rat adrenal quarters or isolated adrenal cell suspension on their response to ACTH was examined through a comparison of amounts of corticosterone produced after their incubation . The response to ACTH added in vitro was considerably higher when adrenal quarters and isolated adrenal cell suspension were prepared at room temperature (25 degree C) than when prepared at ice-cold . Endogenous steroidogenesis was not affected by the temperature . It seemed unlikely that this higher response to ACTH of adrenal quarters or isolated adrenal cell suspension prepared at room temperature was due to an activation of the cells . A possibility was discussed that cooling adrenal quarters or isolated adrenal cell suspension during the preparation may create an unphysiological state in some place to related the cell membrane.

J Immunol Methods, 1975 Feb, 6(4), 331 - 46
A hemolytic plaque assay for the detection of direct and indirect antibody-forming cells to keyhole limpet hemocyanin; Herscowitz HB et al.; A procedure is described for the in vitro enumeration of individual lymphoid cells producing antibody against Keyhole Limpet Hemocyanin (KLH) by a modification of the hemolytic plaque technique . Optimal conditions for the coupling of KLH to sheep erythrocytes by chromic chloride are described . Plaque-forming cells are detected in a liquid monolayer slide chamber assay system . The kinetics of the in vivo primary and secondary PFC responses of both rabbit popliteal lymph node cells and mouse spleen cells are described . Both direct (IgM) and indirect (IgG) plaque-forming cells can be enumerated . The method can also be used to detect an in vitro anamnestic response in dispersed rabbit lymph node cell suspensions . The method is simple, extremely sensitive and the results correlate with those previously obtained in which newly synthesized antibody was detected in similar systems using the coprecipitation assay.

J Biol Chem, 1975 Jan 10, 250(1), 171 - 4
Initiation by methionine of mouse immunoglobulin light chain containing NH-2terminal pyroglutamic acid; Prasad C et al.; The mechanism of biosynthesis of NH2-terminal pyroglutamic acid has been studied in a mouse plasmacytoma (RPC-20) which produces an immunoglobulin light (lambda) chain containing NH2-terminal pyroglutamic acid . To this end, initation of lambda chain synthesis in plasmacytoma cell suspensions has been investigated . The analysis of radioactive lambda chain synthesis by these cells was accomplished with an antibody preparation specific for the precipitation of lambda chain protein from total plasmacytoma protein . NH2-terminal analysis of plasmacytoma cells labeled with {35S}methionine showed that the ratio of radioactivity in NH2-terminal methionine to total incorporation in lambda chain was greater at 2 min of labeling than at 60 min . However, such a pattern of transient labeling of the NH2 terminus of the lambda chain was not obtained when cells were incubated with tritiated leucine, arginine, or tryptophan . The data indicate that methionine is the initiator amino acid for the synthesis of lambda chain containing NH2-terminal pyroglutamic acid.

Arch Geschwulstforsch, 1975, 45(8), 810 - 3
{The influence of the cellular composition of primary tumour cell suspensions on the in-vitro cultural growth rate with monolayer cultivation--a contribution to tumor-immunological interrelationships (author's transl)}; Schonfelder M; Quantitative differential analyses were performed on smears of primary suspensions of human malignant tumours prepared by tryptic digestion . We correlated the findings thus obtained with the in-vitro rate of positive cultivation in monolayer cultivation and clinical courses of tumour carriers . This way we found, that the ratio of lymphocytes to tumour cells in the cell suspension has significant influence on the in-vitro growth rate . We have explained the mutual reaction between lymphocytes and tumour cells as expression of a specific immunological reaction . The correlation with the clinical courses of tumour carriers underlines the in-vitro findings.

Basic Life Sci, 1975, 5B, 719 - 24
Direct evidence that pyrimidine dimers in DNA result in neoplastic transformation; Hart RW et al.; A specific test for the biological role of UV-induced pyrimidine dimers in DNA is photoreactivation (PR) . Fish contain large amounts of the PR enzyme . Portions of cell suspensions of tissue from various organs of the fish Poecilia formosa were exposed to UV radiation (254 nm), then injected into isogenic recipients . An incident fluence of 20 J/m2 resulted in 10% of the fish with large granulomas and 100% with thyroid carcinomas . If the irradiated cell suspension was illuminated with PR light before injection, the yields of both types of lesion were reduced approximately 10-Fold . If the PR light was given before the UV exposure, there was no reduction in the numbers of growths . These experiments show that pyrimidine dimers in DNA can lead to neoplastic transformation.

Eur Surg Res, 1975, 7(6), 326 - 40
Dextrans and the formation of pulmonary metastases after intravenous tumour cell injection in rats non-sensitive to dextran; Ivarsson L et al.; This study showed, that in a syngeneic tumour-host system in rats non-sensitive to dextran, dextran 40 and dextran 100 did not stimulate metastasis formation after intravenous tumour cell injection, neither when given as intravenous pretreatment nor when given in the tumour cell suspension . Dextran 1000 stimulated the formation of metastases in both these situations . Disturbed microcirculation, intravascular coagulation and effects upon the tumour cell membrane appear to be the main resons, why dextran 1000 stimulated metastasis formation under the experimental conditions used.

J Clin Invest, 1975 Jan, 55(1), 198 - 200
The effect of Tuftsin on the nitrous blue tetrazolium reduction of normal human polymorphonuclear leukocytes; Spirer Z et al.; The influence of Tuftsin, the synthetic phagocytosis-stimulating tetrapeptide (L-threonyl-L-lysyl-L-prolyl-L-arginine), on the nitrous blue tetrazolium (NBT) reduction by human polymorphonuclear leukocytes was investigated . It was found that this substance increases the NBT reduction by approximately as much as endotoxin . Other tetrapeptides do not share this property . When Tuftsin analogs are added to the cell suspension and incubated, they prevent the action of both Tuftsin and endotoxin but not of methylene blue . When washed of the analogs, the cells regain the property to be activated by both Tuftsin and endotoxin . It appears that methylene blue on one hand and Tuftsin and endotoxin on the other hand have different sites for their actions . We suggest that whereas methylene blue diffuses into the cell and acts directly upon the hexosemonophosphate shunt activation, Tuftsin and endotoxin appear to act on the cell membrane binding to specific receptors . By treating the cells with Tuftsin analogs, we probably block these receptors.

Clin Genet, 1975 Jan, 7(1), 29 - 36
Cultivated cells from diagnostic amniocentesis in second trimester pregnancies . II . Cytogenetic parameters as functions of clonal type and preparative technique; Hoehn H et al.; From a total of 418 primary amniotic fluid colonies, 5.5% were fibroblast-like (F), 33.7% epithelioid (E) and 60.8% had characteristics of what was previously shown to be the principal class of clonable amniotic fluid cells (AF) . Polyploidy occurred in all three categories, although both pure tetraploidy and mixoploidy were more frequent in E colonies . The incidence of non-constitutional chromosomal changes was identical in AF and E type colonies if primary spreads were analyzed in situ without prior trypsinization . Spreads from pooled cell suspensions showed higher base-line levels of both aneuploidy and structural changes . Analysis of individual colonies employing an in situ preparative technique is clearly the method of choice for a reliable cytogenetic prenatal diagnosis within the shortest possible period of time.

Pflugers Arch, 1975, 354(4), 319 - 37
Microrheology and light transmission of blood . IV . The kinetics of artificial red cell aggregation induced by Dextran; Volger E et al.; Employing both microscopic and photometric methods the rheology of pathological red cell aggregation was studied in model experiments . Suspensions of washed human red blood cells in dextran solutions containing rising concentrations of dextrans (M.W . 40000, 70000, 110000, 250000, 500000) were used . At low concentrations (less than 500 mg-%) of high molecular weight dextrans (greater than 70000) red cell suspensions formed aggregates similar to the ones found in normal human blood . At higher concentrations, the aggregates were similar to those observed in pathological human blood . The aggregates were studied under the condition of stasis, slow flow and at shear rate of their hydrodynamic dispersion . Besides, the flow behavior of the dispersed cells at high shear rates was studied . We found: 1 . In all samples the rate of spontaneous aggregate re-formation in stasis (following hydrodynamic desaggregation) rose with rising dextran concentration up to 5.0 g-% . 2 . The shear resistance of the aggregates, as measured by the shear stress necessary to keep them dispersed, rose up to concentrations of 2.5g-%, but fell at higher concentrations . 3 . Only with dextran of a molecular weight above 110000 coarse agglomerates could be produced at high concentrations . Loose elastic meshes were rapidly produced at high concentrations of Dx 70 . 4 . When subjected to steady state low shear (m sec-1) only the agglomerates, but not the meshes rapidly grew in size . Most of the aggregation kinetics recorded by photometry and microscopy evaded detection by viscometry.

Oncology, 1975, 31(1), 1 - 16
Brain changes and survival of animals with tumors implanted in the brain; Gershbein LL et al.; The effects of tumor cells implanted into the brain of animals on survival rates and gross and microscopic brain changes have been ascertained . Walker carcinosarcoma 256 cell suspensions were injected at several brain sites in rats and leukemia L1210 and P388 and Ehrlich ascites tumor cells, intracerebrally into BDF1 mice . Such neoplasms provided for rapid and rather predictable growth increments . The survival rates were dependent on the number of cells introduced, those receiving the higher counts succumbing first . The brains of the animals revealed fairly discrete space-occupying lesions and neurological symptoms became apparent only hours before death due to intracranial pressure.

J Immunol Methods, 1975, 8(1-2), 151 - 7
A comparative study on the uptake in vitro of {3H}thymidine, {131I}5-iodo-2'-deoxyuridine and {3H}deoxycytidine in mouse spleen cells using double isotope autoradiography; Rooijen NV; The uptake of 3H-labeled thymidine (3H-TdR), 131I-labelled 5-iodo-2'-deoxyuridine (131IUdR) and 3H-labelled deoxycytidine (3H-CdR) by mouse spleen cells in vitro was studied using an autoradiographic method for the separate detection of 3H and 131I in cell smears . The experiment was performed in two steps . In one half of a pooled spleen cell suspension uptake of 3H-TdR and 131IUdR was compared; in the other half of the suspension uptake of 3H-CdR and 131IUdR was compared . Only two nucleated spleen cells were found which had taken up only 3H-CdR . All other scanned cells were double labelled . Generally the nucleotides were taken up according to a rather fixed relation . It was further found that 3H-CdR and 3H-TdR were incorporated to about the same extent . However, cells which were incubated with 131IUdR and 3H-TdR took up the 131IUdR to a lesser extent than cells which were incubated with 131IUdR and 3H-CdR.

Int Arch Allergy Appl Immunol, 1975, 49(3), 358 - 70
Anaphylactoid-inflammation-promoting factor . An insulin-induced factor derived from non-sensitized lymphocytes increases anaphylactoid inflammation in rats; Koltai M et al.; The experiments reported here indicate that, when exposed to insulin, viable lymphocytes rapidly released into the incubation medium a factor capable of increasing the dextran-induced anaphylactoid reaction, but having no effect on the inflammatory response evoked by 5-HT . This pro-inflammatory factor was shown to be elaborated by cell suspensions derived from lymph nodes of rats, rabbits, pigs or calves as well as from human tonsils . Thymus cells showed no such activity . The pro-inflammatory factor was termed as anaphylactoid-inflammation-promoting factor (AIPF) . Its production depended upon the dose of insulin, and the time of exposure . AIPF was found to have an elution pattern in Sephadex G-100 gels similar to that of BSA (67,000 daltons) . The activity was abolished by heat or incubation with DNase or a-chymotrypsin, but was not influenced by RNase . AIPF by itself did not induce increased vascular permeability, and proved to be distinct from the permeability factors present in the lysate of lymph node cells.

Acta Pathol Microbiol Scand {A}, 1975 Jan, 83(1), 25 - 36
7,12-DMBA-induced rat mammary tumour studied for hormonal responsiveness in vitro . 1 . Short-term incubations of cell suspensions; Aspergren K; The effect of testosterone, progesterone, and 17-beta-oestradiol on the incorporation of tritiated thymidine into DNA was studied in vitro using cell suspensions from DMBA-induced rat mammary carcinomas and subcutaneous sarcomas . Both stimulatory and inhibitory effects of the hormones were seen . The two types of tumour reacted similarly . The validity of hormonal responsiveness in vitro compared with in vivo is discussed.

Vox Sang, 1975, 28(2), 81 - 9
Removal of leucocytes from whole blood and erythrocyte suspensions by filtration through cotton wool . IV . Immunization studies in rabbits; Engelfriet CP et al.; Hetero-immunization experiments in rabbits were performed to evaluate the quantity of leucocyte antigens present in red cell suspensions prepared by the filtration method of DIEPENHORST et al., in which no intact leucocytes could be detected . It was found that the quantity of leucocyte antigens is smallest in blood filtered immediately after taking . More leucocyte-antigenic material seems to be present in blood that is filtered after storage and in frozen-thawed red cell suspensions . Evidence was obtained that granulocyte-specific antigens are more strongly immunogenic in the rabbit than lymphocyte-specific antigens or antigens common to both cells.

J Natl Cancer Inst, 1975 Jan, 54(1), 233 - 4
A simple cell-suspension method for transplantation of V2 carcinoma; Berkowitz DM et al.; A simple and rapid method for the preparation of an essentially monodispersed cell suspension from the rabbit V2 carcinoma, suitable for either large or small amounts of tumor tissue, did not require specialized equipment and yielded relatively large numbers of viable cells . Cell suspensions prepared by this method could be used to approximate a dose-response curve relating the percentage of tumor takes to the number of cells inoculated and could allow quantitative correlations not possible when macroscopic tumor fragments are used for implantation . The method, or slight modifications of it, should prove suitable for most tissues with a significant connective tissue component.

Bibl Haematol, 1975, (40), 229 - 33
Regulation of normal hemopoiesis in the acute leukemia L5222; Hoelzer D et al.; Normal hemopoiesis becomes markedly depressed in rats during the development of the rat leukemia L5222 . Whether this could be due to the influence of a circulating humoral factor was investigated by comparing the growth of normal bone marrow cells in diffusion chambers implanted into the peritoneal cavity of normal or leukemic hosts . Similar growth in total cell numbers was observed in both groups of hosts, and no significant difference could be detected between them . In an attempt to exclude the possibility that an inhibitor either did not penetrate to the peritoneum or was inactivated by the chamber membrane, normal serum, or serum from a highly leukemic rat was mixed with the bone marrow cell suspension in the chambers . Again, no difference in growth between the 2 groups could be detected . Therefore, the influence of a circulating humoral factor in the depression of normal bone marrow hemopoiesis in this experimental leukemia seems to be ruled out, and the decrease may be attributable to local events in the bone marrow such as short-range factors or cellular interaction between the leukemic and normal hemopoietic cell populations.

Ser Haematol, 1975, 8(1), 4 - 21
Megakaryocytes in agar cultures of mouse bone marrow; Nakeff A et al.; Cytologic and immunofluorescent data demonstrate that cells morphologically resembling megakaryocytes appear in thin layer agar cultures of cellular fractions of mouse bone marrow and increase in number from zero to a maximum of approxiamately 60 per culture on day 6 . It would appear that these "candidate" megakaryocytes are derived from a morphologically unidentified precursor(s) since they are not present in the original cell suspension cultured but begin to appear 24 hours later . Similarities between the ultrastructure of "candidate" megakaryocytes and that of femoral marrow megakaryocytes support morphological similarities at the light microscopic level . Pretreatment of mice with either vinblastine and nitrogen mustard or antiplatelet serum has no effect on the number of "candidate" megakaryocytes appearing in cultures of their marrow . Preliminary data suggest that the rate of appearance of these cells in culture is responsive to serum from thrombocytopenic donors.

J Exp Med, 1975 Jan 1, 141(1), 1 - 10
Mass isolation and culture of rat kupffer cells; Munthe-Kaas AC et al.; Collagenase perfusion of the liver followed by pronase treatment of the cell suspension thus obtained gave a quantitative recovery of viable nonparenchymal liver cells (NPC) . From these NPC, Kupffer (K) cells can be purified by attachment to tissue culture dishes . Tail vein injection of carbon 1-2 h before liver perfusion permitted stepwise calculation as well as visualization of carbon-containing K cells . When these K cells have been put into tissue culture medium with serum and incubated overnight, they exhibit typical macrophage characteristics . Phase-contrast and transmission electron microscopy showed typical macrophage morphology and scanning electron microscopy revealed well-spread cells with cytoplasmic projections and ruffled membranes . Endocytosis studies using radioactive colloidal gold and inert latex particles also indicated that these cells are highly active in pinocytosis and phagocytosis . Further characterization of K cells is the identification of Fc receptor on their membranes . Studies on lysosomal enzymes showed that purified K cells possess higher specific activities in beta-glucuronidase, acid DNase, and cathepsin D than in purified parenchymal cells.

Immunology, 1975 Jan, 28(1), 161 - 70
A method for the separation of lymphocytes and plasma cells from the human palatine tonsil using sedimentation in an isokinetic gradient of Ficoll in tissue culture medium; Willson JK et al.; Several methods for the dissociation of human tonsils into cell suspensions were compared . Dissociation of tonsils using 0-25 per cent trypsin gave both the largest number of total cells and the largest number of plasma cells per gram of tonsil . Lymphocytes and plasma cells were separated in a previously described isokinetic gradient of Ficoll in tissue culture medium . In the purest gradient fractions, lymphocytes were 97-2 plus or minus 1-9 per cent of nucleated cells . The purest gradient fractions contained 43-1 plus or minus 5-9 per cent plasma cells . More than 95 per cent of purified lymphocytes and plasma cells excluded Trypan Blue.

J Natl Cancer Inst, 1975 Jan, 54(1), 11 - 21
Malignant lymphomas of follicular center cell origin in man . I . Immunologic studies; Leech JH et al.; Lymphomas with histologic features indicating a follicular center cell (FCC) origin were analyzed from 26 patients of a group of 45 consecutive non-Hodgkin's lymphoma patinets whose tumors were studied for B- and T-cell characteristics . They were compared with benign, reactive lymphoid tissue from 14 patients . Cell suspensions from biopsy material, blood, or bone marrow were examined for surface Ig and for rosette formation with sheep erythrocytes (E rosettes) . Of the 26 patients with FCC lymphomas, 22 had 40% or more Ig-bearing cells; all patients with FCC lymphoma tissues had 25% or less E rosette-forming cells . Cells from most FCC lymphomas of the cleaved type had surfac IgM; those from several FCC lymphomas had both IgM and IgD . Cells from lymphomas of noncleaved cell type had surface IgG or IgA . Light-chain analysis showed that cells from FCC lymphomas bore a predominant light-chain type, which indicated their monoclonal nature . Neoplastic cells from several FCC lymphomas synthesized the surface Ig which they bore . Reactive tissues usually contained fewer Ig-bearing and more E rosette-forming cells than FCC lymphomas; the Ig-bearing cells, with one exception, had a polyclonal distribution . Correlation of histologic and immunologic observations indicates that most lymphomas identified as FCC in origin by light micorscopic criteria mark as B cells with the use of immunologic techniques and that FCC lymphomas are the most common type of non-Hodgkin's lymphoma.

J Immunol, 1975 Jan, 114(1 Pt 1), 40 - 4
Differential IgA repopulation after transfer of autologous and allogeneic rabbit Peyer's patch cells; Rudzik O et al.; The distribution and differentiation of rabbit Peyer's patch cells were studied in lethally x-irradiated autologou, and allogeneic recipients, 6 days after i.v . transfer . Immunoglobulin Ig-containing cells were detected in both frozen tissue sections and fixed smears of single cell suspensions of spleens using monospecific anti-alpha, -mu, and -iota antisera in direct immunofluorescence . The tissues of lethally x-irradiated rabbits which had not received any cells were almost devoid of Ig-containing cells whereas the intestinal lamina propria of both autologous and allogeneic Peyer's patch cell recipients had approximately equal numbers of predominantly IgA-containing cells . In the spleen of allogeneic recipients, many large colonies of IgA-containing cells were seen and these cells made up 4.6 to 16.9% (mean 8%) of all spleen cell suspension . The spleens of autologous recipients contained fewer and smaller colonies of IgA cells which made up only 0.5% of all cells in spleen cell suspensions . The numbers of IgM- and IgG-containing spleen cells were small in all animals, however, recipients of allogeneic Peyer's patch cells had four to five times as many as either non-reconstituted lethally irradiated rabbits or autologous recipients . These data confirm that rabbit Peyer's patches are relatively rich in precursors of IgA-producing cells and suggest that histocompatibility differences may either potentiate differentiation of IgA production or lead to trapping of allogeneic Peyer's patch cells in the spleen . Although these experiments did not elucidate the mechanisms of differentiation of IgA-producing cells, the fact that no differences were seen between numbers of IgA cells in gut lamina propria after either autologous or allogeneic cell transfer suggest that different mechanisms may be responsible for appearance of these cells in gut and spleen.

Eur J Immunol, 1975 Jan, 5(1), 70 - 2
Receptors for IgG molecules on human lymphocytes forming spontaneous rosettes with sheep red cells; Ferrarini M et al.; A mixed rosette technique with sheep red blood cells (SRBC) and ox erythrocytes heavily coated with rabbit antibody (EA) was employed to simultaneously identify human peripheral blood T lymphocytes and IgG receptor-bearing cells . The findings of a noticeable proportion of "mixed rosettes" in peripheral blood lymphocytes freshly drawn from normal individuals and of an even higher number of these mixed rosettes in cell suspension kept in culture media supplemented with fetal calf serum provide evidence for the capacity of human T cells to express membrane receptors for antigen-antibody complexes.

Eur J Immunol, 1975 Jan, 5(1), 32 - 9
Surface morphology of murine B and T lymphocytes: A comparative study by scanning electron microscopy; Polliack A et al.; A variety of murine lymphocytes of known B or T derivation obtained from different lymphoid organs were prepared for scanning electron microscopy (SEM) by the critical point drying method after collecting the cells by aspiration onto silver membranes . Comparison of SEM appearances of cells prepared by this technique and serological classification according to surface antigens showed that most T cells had smooth surfaces with few microvilli, while many B lymphocytes were moderately to markedly villous . Further evidence for the above correlation was obtained by examining thymic cells and enriched B or T cell populations . Thymic cell suspensions containing less than 5% B cells showed over 80% generally smooth cells by SEM . Enriched T cell populations, obtained by mass cytolysis of lymph node preparations with anti-Ia or anti-Ig sera or by purification through nylon fiber columns, contained over 85% T cells, and more than 75% of them were of the smooth cell type . A similar correlation was noted for enriched B cell populations obtained by cytolysis of lymph node cells with anti-Thy-1 serum, and by lysis of EAC-rosettes . Over 90% of these cells were identified as B cells by immunologic methods and approximately 75% had moderate to markedly villous surfaces . The 15% difference can be accounted for by the existence of a subpopulation of smooth B cells . Direct observation of EAC-rosettes confirmed that most B cells had moderate to large numbers of surface microvilli and that less than 10% were smooth . It is possible that some of the smooth cells seen in enriched B cell populations may represent precursors or B lymphocytes at different stages of differentiation . These results indicate that murine T and B lymphocytes, like their human counterparts, can be recognized in many cases under the SEM on the basis of their surface morphology . Smoother B and more villous T cells are difficult to classify by SEM without parallel immunologic identification.

Eur J Immunol, 1975 Jan, 5(1), 23 - 6
Spontaneous release of T cell receptors for alloantigens . II . Induction of antibodies to T cell receptors; Ramseier H; Spleen cells from normal mice, when cultivated in vitro, released receptors or recognition structures (RS) for alloantigens into the surrounding medium . The spontaneous shedding of receptors was revealed by their ability to induce the formation of anti-recognition structure(anti-RS) antibodies upon injection into appropriate F1 hybrid recipients . Cell suspensions containing T and B lymphocytes and those containing T lymphocytes were capable of inducing anti-RS antibody formation, whereas suspensions devoid of T cells were incapable of doing so . Receptors shed from such cell suspensions during a 24-hour cultivation period gave exactly the same results . Cell-free culture supernatants, however, incited higher anti-RS antibody titers, presumably because of an accumulation of RS . The capacity of released T cell receptors to recognize alloantigens as determined in the PAR assay and their ability to induce anti-RS antibodies went roughly parallel.

Antonie Van Leeuwenhoek, 1975, 41(4), 421 - 9
Oxidation of organic C1 compounds by Hyphomicrobium spp; Harder W et al.; Washed cell suspensions of Hyphomicrobium spp . were able to oxidize methanol, formaldehyde and formate . This suggested that enzymes for the oxidation of these compounds were present . The pathway of the oxidation of methanol to carbon dioxide and water has been investigated using cell-free extracts . An ammonium-ion-activated, phenazine methosulphate-linked methanol dehydrogenase was detected . This enzyme has a dual substrate specificity for normal primary alcohols and formaldehyde . It has a high pH optimum for activity of 9.5 . The pathway is completed by an NAD-linked formate dehydrogenase . This enzyme is inhibited by low concentrations of potassium cyanide, copper sulphate and hypophosphite.

Scand J Immunol, 1975, 4(8), 823 - 30
Elution and characterization of lymphocytes from rheumatoid inflammatory tissue; Abrahamsen TG et al.; Lymphocytes were eluted from the synovial tissue of 19 patients with classical rheumatoid arthritis . The tissue was minced and dissociated by treatment with crude collagenase and DNase . The cell suspension obtained was filtered and incubated in plastic culture flasks overnight at 37 degrees C . The cells that did not adhere to the plastic surface were harvested and the lymphocytes further purified by the Ficoll-Isopaque gradient centrifugation technique . The lymphocyte yield varied from 0.64 to 32 times 10(6) cells . Differential counts showed on the average 85% lymphocytes, 12% mocrophage-like cells, and variable proportions of polymorphonuclear granulocytes, unclassified cells, and dead cells . An average of 77% of the cells were viable as assessed by the trypan blue exclusion test . This cell suspension was investigated for lymphocyte populations . T lymphocytes were predominant in all experiments (mean, 73.6%) . The mean percentage of B lymphocytes was 9.7%, whereas the proportion of Fc-receptor-bearing lymphocytes was on the average 6.0%.

Clin Exp Immunol, 1975 Jan, 19(1), 75 - 82
In vitro activation of human T and B lymphocytes by pokeweed mitogen; Mellstedt H; In previous in vitro studies the DNA synthesis in human blood lymphocytes induced by low concentration of PWM correlated with the percentage of bone marrow-derived (B) lymphocytes in cell suspensions . No correlation with lymphocyte subpopulations was noted when lymphocytes were activated by high concentrations of PWM . In this paper the hypothesis that low concentration of PWM mainly activates B lymphocytes was tested by measuring the {14C}thymidine incorporation into purified T or B lymphocytes from healthy donors . B lymphocytes were purified to 90--95% by buoyant density centrifugation of T lymphocytes rosetted with sheep red blood cells . T lymphocytes were enriched by passage of lymphocytes through an IgG-anti-IgG-coated column . Low concentration of PWM-stimulated B lymphocytes but not T lymphocytes, while high concentrations of the stimulant activated both cell types . It was also noted that the B- but not the T-lymphocyte fraction contained cells which synthesized DNA in the absence of PWM.

Int Arch Allergy Appl Immunol, 1975, 49(5), 607 - 14
Presence and kinetics of sensitized cells in different tissue compartments; Brocker EB et al.; The question was studied as to whether lymph nodes (LN), spleen (SP) and bone marrow (BM) contain different amounts of sensitized cells at various times after one or repeated immunizations . The following model was used: C57 mice were injected i.p . once, twice or three times with C3H cells . Between 3 and 20 days after the last immunization, LN, SP and BM cell suspensions were tested in vivo for their local cutaneous lymphocyte transfer reactivity in C3H mice and in vitro for their lymphocytotoxicity (partially after preincubation with anti-theta serum) . In both test systems similar observations were made: after one sensitization, reactive cells were demonstrated first in the LN and then in the SP, but only during the next few days; BM cell reactivity appeared later . In the first few days after two sensitizations SP cells reacted strongly whereas LN and BM cells reacted weakly; later BM cell reactivity increased while the others decreased . After three sensitizations reactivity of the SP cells predominated during the entire period of observations; the BM cells showed less, the LN cells no reactivity . These findings suggest dependance of T cell-mediated reactivity on (1) the origin of the cells from LN, SP or BM, (2) the time of the harvesting from these tissues, and (3) the degree of sensitization.

Ciba Found Symp, 1975, 0(29), 315 - 41
A molecular approach to retinotectal specificity; Marchase RB et al.; An assay is described to examine the hypothesis that retinal neurons adhere preferentially to that part of the optic tectum near theri normal synaptic termini . The method measures the adherence of isotopically labelled cell bodies from either the dorsal or ventral half of the neural retina of chick embryos to dorsal and ventral tectum halves . When a labelled cell suspension is prepared from a dorsal half-retina, more cells adhere to the ventral half of the tectum . When the cells are from the ventral part of the retina, more bind to the dorsal half of the tectum . This preferential adhesion mimics the retinotectal projection found in vivo and supports an interpretation of neuronal specificity dependent on cell surface adhesive properties . Molecular mokels are presented that utilize glycosyltransferases and their substrates as the basis for adhesive recognition . Two of these models suggest that quantitative changes in the distribution of transferases and their substrates determine retinotectal specificity . The third proposes qualitative variations in these molecules across the retina and tectum.

Eur J Immunol, 1975 Jan, 5(1), 47 - 53
Immunoglobulin heavy chain mRNA in mitogen-stimulated B cells; Stevens RH et al.; This paper relates the synthesis of DNA, immunoglobulin and heavy chain (H) mRNA in murine spleen cells following activation of B cells with lipopolysaccharide from E . coli (LPS) . Spleen cells (CBA/H mice) were cultivated with 10% FCS and 10 mug LPS/ml . 4 h pulses with {3H}thymidine showed that DNA synthesis was stimulated within the first day following LPS activation and exhibited a sharp peak at 24 h . The shape of the DNA synthesis curve suggests that the cells susceptible to LPS stimulation are activated in a synchronous manner . Stimulation of H-chain mRNA (H-mRNA) synthesis proceeded rapidly (within 6 h of LPS addition) and peaked around 24 h, in parallel to DNA synthesis . The H-mRNA was isolated and quantitated by making use of its interaction with IgG{1, 2} . The actual level of H-mRNA in the culture increased threefold during the first 24 h and then doubled within the next 48 h . Estimates of the actual number of H-mRNA were approximately 200 molecules H-m-RNA/cell on day 0 rising to 1800/cell on day 3 . In such a mixed cell population these figures will be accurate only within a factor of 2-3 (at least 35% B cells in spleen cell suspensions at the commencement of the culture, with up to 35-60% of plasma blasts by day 3 and 4 of LPS treatment) . Translation of the lymphoid cell mRNA in oocytes from Xenopus laevis demonstrated that stimulation of H-mRNA synthesis was restricted to mu-mRNA, although some gamma-mRNA was present in the original spleen cells . High levels of synthesis of immunoglobulin followed after a lag period of about 24 h following LPS addition peaking after 48 and 72 h; the proportional Ig production relative to total protein synthesis reached 26% on days 3 and 4 . Stimulation of Ig production was limited to IgM . Rapid stimulation of mitosis and H-mRNA synthesis thus precedes the maximum synthesis of Ig molecules, suggesting a translational block on H-mRNA during cell maturation . There was no apparent block on the transport of H-mRNA from the nucleus during early stages of activation.

Acta Histochem, 1975, 52(1), 79 - 87
{FDA-hydrolysis for the fluorometrical evidence of a cell membrane alteration in peritoneal macrophages (author's transl)}; Augsten K et al.; The enzymatic hydrolysis of fluoresceindiacetate (FDA) and the intracellular accumulation of fluorescein (fluorochromasia) by murine peritoneal macrophages were photometrically measured in cell suspensions . The intensity of fluorescence (excitation at lambda = 485 nm) was recorded by a right angel detector as a function of time, number of cells per ml, and FDA concentration, respectively . The meter response was found to be directly proportional up to the concentration of the fluorophor of 1.125 x 10(-6) M . In suspension the intensity caused by cells other than macrophages could be neglected . The rate of the fluorescein formation corresponds to the Michaelis-Menten constants (km = 8,34 x 10(-6) M.) Repeated in vivo administrations of some N-mustard benzimidazole derivatives were followed by competitive and non-competitive inhibition of the FDA-hydrolysis, respectively . Consequently, the fluorescence intensity varied, but without an concomitant adequate alteration of the non-specific (alpha-naphthyl acetate) esterase as was being measured by means of photometry of single macrophages in cell smears . Thus, this method seems to be suitable for quantitation of membrane (permeability) alteration induced experimentelly in macrophages.

Transplantation, 1975 Jan, 19(1), 42 - 7
Altered allotransplantability of BALB/c Leydig cell tumor after organ culture or cell suspension; Ninnemann JL et al.; In confirmation of prior studies, fragments of testicular interstitial (Leydig) cell tumors from BALB/c mice were found to be transplantable to H-2d (BALB/c and DBA/2) or allogeneic H-2q (DBA/1 and BUB/BnJ) recipients after organ culture, whereas noncultured fragments were accepted only in H-2d recipients . Lymphocyte cytotoxicity assays showed that while transplantability was altered by the organ culture process, tissue immunogenicity in vitro was unchanged . Cell suspensions from the same tumor were also found to be transplantable to both H-2d and H-2q recipients and occasionally even to H-2a and H-2k recipients, irrespective of whether the cells were obtained from fragments previously organ-cultured or from dispersed fresh tissue . We suggest, therefore, that rather than modifying cell surface antigens, it is possible that organ culture explanation serves to (1) allow free dispersal of the tumor cells which leads to progressive tumor growth, and/or (2) raise the ratio of dead to living cells in the transplanted fragments, which has previously been shown to direct host immune responses toward enhancement.

Tex Rep Biol Med, 1975, 33(3), 415 - 22
Morphological changes induced during fixation of Mycoplasma mycoides for electron microscopy; Furness G et al.; Only spherical mycoplasmas 0.6-0.9 mum diam were observed by darkfield microscopy in single cell suspensions prepared from exponential broth cultures of Mycoplasma mycoides var mycoides and Mycoplasma mycoides var capri . Similar cells were seen in pseudoreplicas by electron microscopy and they are considered characteristic of the morphology of M . mycoides . When the mycoplasmas were fixed by the addition of 10% formalin to the suspensions or washed, centrifuged cells were fixed by glutaraldehyde, filamentous and ring forms were observed in electron micrographs . These are not considered typical of the morphology of M . mycoides but are artifacts produced during the preparation of the mycoplasmas for electron microscopy.

Eur Neurol, 1975, 13(1), 19 - 30
Drainage of thoracic duct lymph in twelve patients with myasthenia gravis; Bergstrom K et al.; The effect of thoracic duct lymph drainage (5-34 days) in 12 patients with myasthenia gravis on muscular function has been followed for 5-43 months . Among the results obtained were: (1) During the drainage the myasthenic symptoms decreased markedly after 1-4 days and remained so during the drainage . (2) The doses of cholinesterase inhibitors had to be markedly reduced during the lymph drainage in eight patients . (3) Discontinuation of the lymph drainage increased the myasthenic symptoms within a few days . However, after a median observation time of 14 months with conventional treatment all but one of the patients had improved . (4) Retransfusion of the patients own cell-free lymph caused a worsening of the myasthenic symptoms . This effect could also be obtained following infusion of IgG preparations from the patients lymph . Three retransfusions of cell suspensions obtained from the thoracic duct lymph from two patients had no effects on their myasthenic symptoms . (5) It is suggested that thoracic duct lymph drainage can be combined with other forms of treatment in severe cases of myasthenia gravis.

Z Krebsforsch Klin Onkol Cancer Res Clin Oncol, 1975, 83(2), 97 - 104
Sensitivity tests of tumors to cytostatic agents . II . Investigation on human tumors; Mattern J et al.; Certain substances which influenced nucleic acid metabolism were found to have about the same cytostatic activity on human cells when measured in tissue culture experiments (cell numbers) or in short-term cultures (3-H-uridine incorporation in cell suspensions) . By treatment with a dose of cytostatics corresponding to 10 times therapeutic dose, chemosensitive tumors can be distinguished from non-responsive tumors . By using this in vitro test system to investigate the sensitivities of 100 human tumors, it is shown that 28 of these tumors were responsive to adriamycin, daunomycin and Actinomycin D . Good agreement was observed between these in vitro results and the literature data on clinical therapy using these particular substances.

Z Krebsforsch Klin Onkol Cancer Res Clin Oncol, 1975, 83(2), 85 - 96
Sensitivity tests of tumors to cytostatic agents . I . Comparative investigations on transplanted tumors in vivo and in vitro; Volm M et al.; With series of transplanted tumors, the activities of different cytostatic agents which directly influence the metabolism of nucleic acids (Actinomycin D, adriamycin, daunomycin, 5-fluorouracil, procarbazine, trenimon) was measured by determining 3-H-uridine incorporation in short-term (3hrs) incubations of tumor cell suspensions . Data obtained could be used to predict the response of each tumor to particular cytostatic agents in vivo . The activities of the cytostatic agents as determined using long-term tissue cultures (time of exposure of tumor cells to cytostatic agent 48 hrs were comparable to those obtained with the short-term test . In long-term cultures, determination of cell numbers gave results similar to those obtained by morphological evaluation . In SHORt-term test, differing sensitivities of tumors to cytostatics could be detected.

Folia Haematol Int Mag Klin Morphol Blutforsch, 1975, 102(4), 462 - 9
{Potency determination of horse anti-human-lymphocyte serum (ALS) by means of the lymphocyte resistance test, a modification of Schroder's leukocyte resistance test}; Mundt B et al.; The leukocyte resistance test of Schroder was used for the purpose of modifying the in vitro determination of cytotoxic potency of ALS during the course of immunization . In contrast to the original method, where the test was carried out with whole blood, the incubation was made with pure lymphocyte cell suspensions . The results obtained show that the extensive procedure of examination enables the cytotoxic effectiveness of antilymphocytic antibodies to be evaluated.

Int Arch Allergy Appl Immunol, 1975, 49(3), 285 - 92
Inhibition by nicotinamide of an homologous PCA reaction and antigen-induced histamine release from rat peritoneal mast cells; Wyczolkowska J et al.; The antigen-induced release of histamine both in vitro from rat peritoneal cell suspensions containing mast cells actively sensitized with IgE antibody and in vivo from passively sensitized skin can be inhibited by relatively high doses of nicotinamide . The action of nicotinamide on the cellular mechanism involved in the regulation of antigen-induced histamine release is discussed.

Acta Endocrinol Suppl (Copenh), 1975, 194, 37 - 54
Immunocytological methods for the identification and localization of antigens; Avrameas S; Immunocytologic techniques allow the study of antigenic cellular constituents by optical or electron microscopy, Antibodies labelled with tracers such as fluorochromes, enzymes, radioactive isotopes, or large electron-dense molecules form the basis of these techniques . The immunochemical procedures involved in the preparation of such conjugates are well defined . Satisfactory results, however, also depend upon the proper preparation and processing of the tissue or cell suspension being studied.

Int Arch Allergy Appl Immunol, 1975, 48(4), 513 - 29
Ontogeny of lymphoid cell surface determinants in the chicken; Albini B et al.; Five antigenic lymphoid cell surface determinants (LCSD) were detected in hatched chickens using specific antisera . These LCSD were: thymus-specific surface determinants, bursa-specific non-immunoglobulin determinants, IgM-specific determants, IgG-specific determinants, and IgA-specific determinants (ASD) . Viable cell suspension of embryonic yolk sac, bursa, thymus and spleen were tested by means of indirect or direct immunofluorescent staining procedures for the presence and frequency of LCSD during maturation . Experiments performed with liver cells . brain cells and red blood cells of embryos confirmed the specificities of the antisera used for determinants present on cells of lymphoid tissues . The results showed LCSD to occur on yolk sac cells on the 5th to 7th embryonic day (ED) . This suggests the presence of a stem cell pre-committed for the lymphoid cell line already in the yolk sac . Furthermore, findings are reported indicating the presence of distinct lympoid stem cell populations or maturation stages in the yolk sac, which may be responsible for either populating the thymus or the bursa . The finding of ASD-bearing cells early in ontogenesis of the lymphoid system suggests the presence of two specificities in anti-chicken IgA sera, one of which may be directed against an antigenic site on a rudimentary immunoglobulin molecule, which becomes lost or hidden in later maturation . Studies on the bursa and the thymus show that covering, hiding, or loss of antigenic determinants plays an important role in lymphoid cell differentiation . Furthermore, the spleen is reached by B-determined stem cells as early as the bursa, but these stem cells seem not to proliferate in the former to any considerable extent until hatching . Finally, the sequence of the appearance of immunoglobulin classes as proposed by other authors is confirmed with reservaitons concerning IgA, and it is suggested that immunoglobulins are detectable earlier on cell surfaces than intracytoplasmatically.

Int Arch Allergy Appl Immunol, 1975, 48(3), 372 - 82
Observations on rabbit thymocytes and peripheral T cells . I . Anomalous results in the mixed antiglobulin reaction caused by non-specific adsorption of sheep immunoglobulin; Wilson AB et al.; The "single-stage" mixed antiglobulin reaction (MAR) was carried out with rabbit thymocytes . This test involved treating the cells with either sheep or goat anti-rabbit globulin sera, and subsequently reacting them with indicator erythrocytes coated with rabbit immunoglobulin (Ig) so as to form rosettes . An unexpectedly high number (up to 38%) of thymocytes reacted, although the rosettes were weaker than those given by peripheral B lymphocytes . When blood and lymph node lymphocytes or thymus cells which had already been treated with sheep anti-rabbit globulin serum were subsequently exposed to rabbit anti-sheep Ig serum and then rosetted with indicator cells coated with ox Ig (cross-reacts with sheep Ig) almost 100% reaction was obtained in each of the cell suspensions . This was designated the "two-stage" MAR . The anomalous results, both in the one-stage and two-stage MAR, were abolished by pepsin-treating the sheep anti-rabbit globulin serum; thus indicating that sheep Ig is adsorbed non-specifically via the Fc part of the molecules to the surface of rabbit thymocytes and peripheral T lymphocytes.

Ann Immunol Hung, 1975, 18, 53 - 6
Study on the interaction of lymphotropic cytostatic agent treatment and graft-versus-host (GVH) reaction in mice; Anderlik P et al.; F1 hybrid mice (C57BL X C3H) suffering from graft-versus-host disease, were treated with dianhydrodulcit (DAD) a lymphotrop cytostatic agent . 15 mg/kg of the compound was injected intraperitoneally, 8 days following the administration of the parental spleen cell suspension . The interaction between the two interventions is discussed on the basis of the rate and time course of deaths observed is the individual groups, as well as on the state of the lymphoid organs of the succumbed and sacrificed animals.

J Lipid Res, 1972 Mar, 13(2), 234 - 43
Fat mobilization in vitro and in vivo in the genetically obese Zucker rat "fatty"; Zucker LM; Fat mobilization was studied in vitro with epididymal fat pad tissue and also with cell suspensions from epididymal, retroperitoneal, and subcutaneous fat from the obese mutant "fatty" (fafa) and control rats of four different ages . Fat mobilization per cell in response to epinephrine was well above normal in young "fatties"; in older "fatties" the output per cell fell to near normal, but the much greater number of fat cells per rat indicated that the fat mobilizing capacity of the older "fatty" is well above normal . The "fatty" showed normal reactions to epinephrine in vivo: hyperglycemia, glycogenolysis, lipolysis with elevated free fatty acids and glycerol, and increased entry of free fatty acids into muscle and liver . Response was at least as great in "fatty" as in control animals . Metabolic indices-levels of circulating free fatty acids, glycerol, and in some cases glucose and lipid-determined at various ages in fed "fatties" and controls, and at intervals during prolonged fasting (70 days), were consistent with a picture of excessive adipose tissue lipolysis, excessive reesterification in the adipose tissue, fat mobilization in excess of need, and return of the excess to the adipose tissue via lipoproteins.

J Lipid Res, 1971 Sep, 12(5), 545 - 52
Utilization of free fatty acids complexed to human plasma lipoproteins by mammalian cell suspensions; Spector AA et al.; The purpose of this study was to determine whether lipoprotein-bound free fatty acid could be utilized by isolated mammalian cells . Ehrlich ascites tumor cells were incubated in vitro with radioactive free fatty acids that were bound to human plasma lipoproteins . Under these conditions, lipoprotein-bound free fatty acids were readily taken up by the cells . After 2 min of incubation with free fatty acids bound to low density lipoproteins, most of the radioactivity that was associated with the cells was in the form of free fatty acids . As the incubation continued, increasing amounts of radioactivity were incorporated into CO(2) and cell lipids, particularly phospholipids . Most of the free fatty acid uptake was the result of fatty acid transfer from low density lipoproteins to the cell, not from irreversible incorporation of the intact free fatty acid-low density lipoprotein complex . Fatty acid uptake increased as the ratio of free fatty acid to low density lipoprotein was raised . When albumin was added to the medium, free fatty acid uptake decreased . A large percentage of the newly incorporated cellular radioactivity was released into the medium if the cells were exposed subsequently to a solution containing albumin . Most of the released radioactivity was in the form of free fatty acid . The results with this experimental model suggest that lipoprotein-bound free fatty acid, like albumin-bound free fatty acid, is readily available for uptake by isolated cells . The mechanism of free fatty acid utilization by the Ehrlich cell is similar when either low density lipoprotein or serum albumin serves as the fatty acid carrier.

J Lipid Res, 1969 May, 10(3), 253 - 9
Effects of prolonged incubation and cell concentration on lipogenesis from glucose in isolated human omental fat cells; Goldrick RB et al.; The behavior of human omental fat cells in vitro has been examined in order to define conditions under which glucose is converted to glyceride-glycerol and glyceride fatty acids . Synthesis of glyceride fatty acids from glucose reached maximal rates only after several hours of incubation in Krebs-Henseleit bicarbonate buffer, with or without added bovine albumin . Conversion of glucose to glyceride fatty acids was readily demonstrable with concentrated cell suspensions and was stimulated 3- to 8-fold by insulin . With dilute cell suspensions, little fatty acid was synthesized even after prolonged incubation in the presence of insulin . Conversion of glucose to glyceride-glycerol was linear during 6-hr incubations in buffer and unaffected by the concentration of the cell suspension . In the presence of bovine albumin, glyceride-glycerol synthesis was readily demonstrable at all cell concentrations used, although synthesis was faster in dilute suspensions . Thus, different incubation conditions produce widely divergent patterns of glucose metabolism in human omental fat cells.

J Lipid Res, 1968 Jan, 9(1), 110 - 9
Methods for the determination of adipose cell size in man and animals; Hirsch J et al.; Four methods for the sizing of adipose cells in small samples of human or animal adipose tissue are compared . These methods depend on the preparation of cell suspensions by incubation of the tissue with collagenase or by prolonged fixation with osmium tetroxide and separation of the fixed cells . A Coulter electronic counter was used to count and size the suspended cells and a Zeiss particle size analyzer for the sizing of cells in photomicrographs . The use of the Coulter counter to count cells in a suspension derived from a known amount of tissue and subjected to osmium tetroxide fixation is recommended for accuracy and general applicability to adipose cells of all sizes in man and animals.

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