Cell suspensions

J Immunol, 1975 Sep, 115(3), 827 - 33
Subpopulations of human thymus cells differing in their capacity to form stable E-rosettes and in their immunologic reactivity; Galili U et al.; The majority of human thymus cells from young donors form stable E-rosettes with sheep red blood cells (SRBC) that do not distintegrate after prolonged incubation at 37 degrees C . With advancing age the proportion of thymus cells forming such rosettes decreases gradually . The thymus of a patient receiving prednisone treatment was found to contain only a few cells that formed stable E-rosettes . The minor population of thymus cells that fails to form stable E-rosettes (non-rosetting or NR cells) was isolated and tested for its cell surface markers and immunologic reactivity in vitro . Most of the NR-cells were capable of forming regular E-rosettes with SRBC at room temperature . Like the majority of human thymus cells they were sensitive to the cytotoxic effect of normal constituted less than 0.2% of the original thymus cell suspensions, but about 1 to 3% of the NR-population . Thymus cells from donors over the age of 36 and from a prednisone-treated child responded in vitro to stimulation with either phytohemagglutinin (PHA) or concanavalin A (Con A) . Unfractionated thymus cells from children up to the age of 14 failed to react to either PHA or Con A, but their NR-population responded vigorously to both lectins . In contrast to unfractionated thymus cell suspensions from children, the NR fraction showed a significant reactivity in mixed lymphocyte cultures with mitomycin-C treated allogeneic lymphocytes . It is concluded that like the thymus of other species, the human thymus contains a minor population of cortisone-resistant cells endowed with many of the immunologic properties characteristic for periperal T lymphocytes.

J Immunol, 1975 Sep, 115(3), 811 - 6
Modulation of HL-A antigens by anti-HLA antiserum: effects on the cytotoxicity assay and mixed leukocyte reaction; Sadeghee S et al.; Modulation of human leukocyte antigen (HLA) was attempted by treating leukocytes with specific anti-HLA antiserum or by their passage through columns coated with anti-HLA or a double layer of HLA-anti-HLA . The modulated cells were resistant to the cytotoxic effects of the anti-HLA, and they were poor stimulators and good responders to allogeneic cells in the unidirectional mixed leukocyte reaction . Modulated cells regained their HLA 16 hr after modulation if kept in cell suspension alone . The proliferative responses of modulated cells to mitogens were as good as non-modulated cells, indicating that modulation was probably not caused by depletion of lymphoid cells . Supernatants of modulated cells that were incubated overnight or preformed HLA-anti-HLA complexes were capable of suppressing and enhancing the MLR of specific cells depending on the dose used . The similarities of modulation of HLA to other lymphocyte receptors and the limitation of application of the modulation phenomenon to transplantation of allogeneic cells are discussed.

Transfusion, 1975 Sep-Oct, 15(5), 439 - 48
Efficacy of leucocyte-poor red blood cell suspensions prepared by sedimentation in hydroxyethyl starch; Dorner I et al.; Hydroxyethyl starch (HES), which is licensed for human use as a plasma volume expander, has red blood cell sedimenting properties similar to high molecular weight dextran (HMWD) . The present studies document the effectiveness of commercially available HES in preparing leukocyte-poor red blood cell suspensions (L-PRBCs) from donor whole blood and packed red blood cells stored in CPD up to 120 hours prior to processing . The effect of HES sedimentation on final white blood cell count, on erythrocyte p50 and 2,3-DPG concentration, and on short- and long-term in vivo survival of 72-hour stored donor red cells are described, along with favorable clinical experience with over 300 transfusions of HES-sedimented L-PRBCs . Since the method requires no special equipment, it is immediately available for widespread use.

Z Naturforsch {C}, 1975 Sep-Oct, 30(5), 650 - 8
Oxidative decarboxylation of para-hydroxybenzoic acids by peroxidases under in vivo and in vitro conditions; Berlin J et al.; Oxidative decarbyxylation of p-hydroxybenzoic acids in plant cell suspension cultures is catalyzed by peroxidases . This reaction has been characterized in vivo and in vitro . Decarboxylation of substituted benzoic acids yields monomeric, dimeric and oligomeric benzoquinones . All peroxidases obtained from soybean (Glycine max) cell suspension cultures by gel electrophoresis are equally capable to decarboxylate p-hydroxygenzoic acids as indicated by their rather low differences in specific activity for various benzoic acids.

Vopr Virusol, 1975 Sep-Oct, (5), 581 - 6
{Comparative study of Aujeszky's disease virus reproduction in a suspension of poultry embryonic cells and tissues (author's transl)}; Osidze DF et al.; Reproduction of Aueski disease virus in suspension cultures of trysinized cells and mechanically minced tissue of chick, duck and quail embryos was compared . The optimal conditions for cultivation of vaccine and virulent strains of virus in these systems were determined . The advantages and prospects of using suspension cultures of minced avian embryo tissue for preparation of virus materials with high biological activity and in large volumes in comparison with trypsinized cell suspensions were demonstrated.

Biofizika, 1975 Sep-Oct, 20(5), 853 - 8
{Light scattering by cell suspensions in normal conditions and exposed to external factors}; Ostashevskii IIa et al.; The characteristics of light scattering of cell suspensions in norm (pH 7,2, t=20degreesC) and upon external influences (change of pH and increase of tdegree) . The turbidity tauapproximatelylambda-n and n=0,2--0,3 for cells in norm . After cell damage n increases . Dependence of n correlates with the increase of some injured cells determined by eozin test . Alterations of light scattering after cell damage were connected with the increase of deposit of intercellular structure in general scattering.

J Biol Chem, 1975 Aug 10, 250(15), 5791 - 800
Changes in nuclear proteins of rat testis cells separated by velocity sedimentation; Platz RD et al.; The technique of velocity sedimentation at unit gravity has been used to separate rat testis cell suspensions into fractions enriched in particular cell types . Changes in the nuclear proteins from the various fractions have been characterized by polyacrylamide gel electrophoresis, and correlated with the changing morphology of the nucleus during spermatogenesis . The most striking alterations in both protein composition and nuclear morphology occur during spermatid maturation as both histone and non-histone proteins are replaced by highly basic, low molecular weight, spermatidal proteins . This replacement process is accompanied by a quantitative reduction in both histone and non-histone proteins . The synthesis of at least three basic proteins has been identified with late stage spermatids . One of these proteins is a highly basic sperm-specific protein containing high levels of cyst(e)ine and arginine . A second protein synthesized in late stage spermatids is lysine rich, while the third protein contains cyst(e)ine and co-migrates with histone F2a1 on acid-urea polyacrylamide gels . The changes in protein composition of rat testis nuclei after irradiation or hypophysectomy reflect the resulting changes in the cellular composition of the testis . After selective elimination of the germinal cells by irradiation, the electrophoretic pattern of acid-soluble proteins from the testis is very similar to that of somatic tissue . Thus, the cellular specificity of nuclear proteins demonstrated here using cell separation techniques is also apparent following treatments which selectively alter the cellular composition of the testis.

J Microsc, 1975 Aug, 104(3), 235 - 44
A freeze-fracture replication apparatus for biological specimens; Stolinski C; A freeze-fracture apparatus of original design has been constructed which can be fitted onto a standard vacuum evaporator unit . In it, cell suspensions and organized tissue may be processed by inserting a sample into a cylindrical holder . By leaving a small part of the tissue protruding from the holder, pre-selected and aligned portions of the specimen can subsequently be revealed by fracture under vacuum . After rapid freezing, the specimen remains firmly attached to the inner wall of the sample holder, preventing its possible loss during fracturing . A mechanism, in the form of a double-sided converging wedge, which is operated from outside the vacuum chamber, is used to produce a fracture in the specimen . The device gently induces a fracture in the desired part of the tissue and lifts the protruding part of the specimen out of the way . In this way, reasonably flat fracture faces are produced for subsequent replication . As the fracturing mechanism comes into contact only with the outer edges of the specimen, damage and contamination liable to occur when the entire specimen is traversed by a blade, is avoided . In addition the specimen stage is surrounded by a cold metal shroud which acts as an efficient trap for contaminants . In this way, favourable vacuum conditions are produced in the vicinity of the specimen . Such effective enclosing of the specimen also facilitates controlled sublimation of the sample.

J Gen Physiol, 1975 Aug, 66(2), 251 - 65
Reflection coefficients of permeant molecules in human red cell suspensions; Owen JD et al.; The Staverman reflection coefficient, sigma for several permeant molecules was determined in human red cell suspensions with a Durrum stopped-flow spectrophotometer . This procedure was first used with dog, cat, and beef red cells and with human red cells . The stopped-flow technique used was similar to the rapid-flow method used by those who originally reported sigma measurements in human red cells for molecules which rapidly penetrate the red cell membrane . The sigma values we obtained agreed with those previously reported for most of the slow penetrants, except malonamide, but disagreed with all the sigma values previously reported for the rapid penetrants . We were unable to calculate an "equivalent pore radius" with our sigma data . The advantages of our equipment and our experimental procedure are discussed . Our sigma data suggest that sigma is indirectly proportional to the log of the nonelectrolyte permeability coefficient, omega . Since a similar trend has been previously shown for log omega and molar volume of the permeant molecules, a correlatioo was shown between sigma and molar volume suggesting the membrane acts as a sieve.

Am J Vet Res, 1975 Aug, 36(08), 1207 - 10
Transmission and attempted isolation of the etiologic agent associated with lymphofollicular hyperplasia of the canine species; Jackson JA et al.; From 18 donor dogs of different breeds and ages, follicular lesions of the third eyelid (plica senilunaris conjunctiva) and genitalia were surgically removed, trypsinized, and inoculated on monolayers of HeLa, rabbit kidney, and canine kidney cell cultures . Blind passages of the lesion material were made every 96 hours for 10 to 15 cell culture passages . Cellular suspensions prepared from the lesions were grown in test tubes and passaged 3 times at 10-day intervals between passages . All cultures were observed each day for cytopathic effect . Transmission studies were made by (1) inoculating normal pups with cellular suspensions of the lesions from an infected dog and an infected pup, (2) placing normal pups in contact with infected ones for contact transmission, and (3) inoculating normal animals with cell suspensions prepared from inoculated monolayers . Cytopathic changes were not seen in any of the cell culture monolayers . All transmission attempts were successful, in that characteristic lesions comparable in appearance to those seen in natural infections were produced in susceptible pups . The lesion material from an infected pup was found to be infective for a normal pup after 6 passages in tissue culture (primary rabbit kidney cells) despite absence of cytopathic effect.

J Lab Clin Med, 1975 Aug, 86(2), 326 - 43
Blood cell structure-function studies: light transmission and attenuation coefficients of suspensions of blood cells and model particles at rest and with stirring; Frojmovic MM et al.; The Beer-Lambert law of light transmission was found valid for suspensions of blood platelets, erythrocytes, and leukocytes, as well as similarly sized model particles (latex spheres and flat guanine crystals), for a DB-G double-bean photometer at 605 and 710 mg, and a Payton aggregometer with red filter (660 to 750 mu) . Attenuation coefficients (K) showed a similar dependence on a particle "equivalent sphere" radius as data reported for a "zero-degree photometer," for particles at rest . Size distribution, degree of aggregation, state of contamination, and effects of aldehyde fixation of the cell suspensions, well as optical geometry, were related to the measured K values . Particle optical efficiencies (K) for asymmetric particles were similar whether calculated from cross-sectional areas derived from "equivalent spheres" or from geometric cross-sections accounting for orientation distributions at rest . Appropriate cell and latex models were identified for the erythrocytes (E) and platelets (P) at rest on the basis of similar K,k values . The biphasic change in K occurring with stirring of these asymmetric blood cells (E, HP) was partly explainable by the known variations of K with shear-induced particle alignment to the optic axis . No such effects were observed with spherical particles or "sphered" cells, even when polydisperse as to size an aggregates . This investigation begins to quantitate ongoing light transmission studies of blood cell shape/aggregation changes.

Cancer Res, 1975 Aug, 35(8), 2145 - 53
Sequential irreversible, actinomycin D-sensitive, and cycloheximide-sensitive steps prior to cortisol inhibition of uridine utilization by P1798 tumor lymphocytes; Stevens J et al.; Events preceeding the cortisol inhibition of uridine utilization by corticoid-sensitive P1798 lymphocytes have been investigated . When tumor cells were incubated with 1 muM cortisol for 15 min and then washed free of steroid and reincubated in the absence of hormone, the expected decrease of uridine uptake failed to appear 1.5 hr later . In contrast, the removal of cortisol after 30 or 60 min did not prevent subsequent development of the steroid effect . Addition of actinomycin D with cortisol, or 15 min after hormone treatment was started, blocked steroid action . However, when actinomycin D was added 30 or 60 min after the initial exposure to cortisol, hormone-induced depression of uridine uptake was no longer prevented . To study the role of protein synthesis, cycloheximide was added to the tumor cell suspensions at various times after cortisol treatment was started . Cortisol suppression of uridine utilization was blocked when cycloheximide was added with the hormone or 30 min after the start of hormone treatment . Cycloheximide added together with cortisol and washed out with the steroid after 30 min did not prevent subsequent appearance of decreased nucleoside uptake . Hydroxyurea, an inhibitor of DNA synthesis, did not prevent cortisol action, even when present throughout a 2 hr exposure to the steroid . Hormone removal or actinomycin D addition after 1.5 to 2 hr (when uridine uptake was already inhibited about 25%) did not prevent intensification of the steroid effect during a subsequent 1.5- to 2-hr incubation period, while addition of cycloheximide at this time completely prevented its progression . These results suggest aht: (a) cortisol inhibition of uridine uptake by P1798 lymphocytes involves an early irreversible step and appears to require continuing RNA but not protein synthesis during the first 15 to 30 min of hormone action; (b) protein synthesis but not RNA synthesis is required between 30 and 60 min; and (c) continuing protein synthesis but not RNA synthesis or hormone presence is necessary for the preestablished cortisol effect to progress.

Cancer Res, 1975 Aug, 35(8), 2098 - 103
Potentiation of hamster tumors by normal cells or charcoal; El Mishad AM et al.; Admixed spleen cells from normal animals or from animals given injections of Syrian hamster type C virus significantly potentiated the growth of the transplanted D9 lymphoma of random-bred hamsters . Potentiation was measured by an increase in incidence of tumors, a shortened latent period, and a decreased 50% tumor-producing dose of tumor cells . Intermediate doses of spleen cells (10 to 100 spleen cells per tumor cell) produced the greatest potentiation . Preincubation of admixed spleen and tumor cell suspensions in vitro was unnecessary . Immunization to isoantigens was not responsible for potentiation, since growth of a transplantable carcinoma of inbred hamsters was also facilitated by normal spleen cells . In addition, normal kidney or liver cells increased the incidence of tumors transplanted by a small number of tumor cells . Potentiation did not occur when spleen cells were injected at a site remote from the tumor cells . Since the potentiating cells might act eigher as a physical barrier to host response, or by blocking normal macrophage function, we injected charcoal with tumor cells . Simultaneous treatment with charcoal facilitated the growth of the lymphoma but not that of the carcinoma . Treatment with some doses of charcoal was also effective at distant sites . Although potentiation of tumor growth by cells or charcoal may operate through different mechanisms, these phenomena should be explored in regard to outgrowth of primary tumors, tumor immunity, or enhancement of tumor growth.

J Microsc, 1975 Aug, 104(3), 321 - 3
A container for processing small volumes of cell suspensions for critical point drying; Newell DG et al.; The attachment of lymphocytes to glass or filters, in order to facilitate handling for processing prior to scanning electron microscopy, may introduce artefacts in surface topography . A container has therefore been adapted, from an embedding capsule, for the preparation of small volumes of cell suspensions.

J Immunol, 1975 Aug, 115(2), 513 - 8
T cell products activating stem cells: further studies on the origin and action of the factor(s); Cerny J et al.; We studied the capacity of bone marrow hemopoietic stem cells to form colonies (colony forming units, CFU) in the spleens of irradiated recipient mice following incubation of the cells with T cell-derived mediators in short-term culture in vitro . The mediators were a) stem cell-activating factor (SAF) present in crude form in the supernatant from phytohemagglutinin (PHA); stimulated lymphocytes, and b) immunoenhancing factor (IEF) obtained in partially purified form from the supernatant of antigen-stimulated lymphocytes and characterized by its ability to enhance antibody formation in vitro . Both SAF and IEF increased the number of CFU in the cultured bone marrow cell suspension . However, only SAF, and not IEF, significantly stimulated DNA synthesis in the cultured cells, Furhter more, SAF appears to activate CFU in vitro, rather than merely to promote the survival of active CFU stem cells . Experiments with SAF and bone marrow from different strains of mice indicated that the biologic activity was not restricted by K and Ia regions of H complex.

Eur J Immunol, 1975 Aug, 5(8), 579 - 84
Antibody-dependent human lymphocytotoxicity: a micro assay system; Zeijlemaker WP et al.; A micro assay system is described for measuring antibody-dependent lymphocytotoxicity with human effector cells . The target cells were mouse mastocytoma cells sensitized with rabbit antibody . The tests were performed in microtiter plates, each well containing 5 x 10(3) 51Cr-labeled target cells, together with varying numbers of effector cells . Whereas highly purified lymphocyte suspensions induced lysis, purified monocyte and granulocyte suspensions had virtually no effect . In parallel experiments it was shown that the latter cell suspensions were very active in lysing sensitized human erythrocytes, which in turn were not lysed by purified lymphocytes . Finally, it was shown that the lytic capacity of lymphocytes was not affected by preservation of the cells at liquid nitrogen temperature . It is concluded that the system described can be used for the clinical analysis of antibody-dependent lymphocytotoxicity in lymphocyte suspensions.

Eur J Immunol, 1975 Aug, 5(8), 554 - 9
The nature of active and passive thyroglobulin binding lymphoid cells in Obese strain (OS) chickens; Richter E et al.; Thyroglobulin-binding lymphoid cells were identified in the spleen of Obese strain (DS) chickens by their capacity to form rosettes with thyroglobulin-coated chicken red blood cells . The nature of these cells was studied in inhibition experiments using turkey anti-chicken bursa or thymus cell sera and rabbit antisera specific for chicken Ig, gamma, mu, alpha, Fabgamma or Fcgamma . Spleen cells actively synthesizing surface receptors for thyroglobulin were identified as B cells and the receptors found to be complete IgM molecules . Normal T cells became thyroglobulin-rosette-forming cells via passive adsorption of thyroglobulin antibodies, a phenomenon which could be inhibited competitively by the addition of normal chicken serum to the incubation medium . Thyroglobulin antibodies passively adsorbed onto the surface of normal T cells also belong to the IgM class as verified both by inhibition experiments and studies employing IgM and IgG fractions of a high titered OS serum for the preincubation of the cell suspensions . Only preincubation with the IgM fraction of the anti-thyroglobulin antibodies resulted in the formation of significant numbers of passive rosette-forming cells.

Am J Physiol, 1975 Aug, 229(2), 334 - 9
Oxygen-linked CO2 transport in sheep blood; Bauman R et al.; We have analyzed oxygen-linked carbamate formation in sheep hemoglobin B by measuring a) the effect of CO2 on oxygen affinity and Bohr effect in red cell suspensions and dilute (1.3 mM Hb4) and concentrated (5 mM Hb4) hemoglobin solutions at 37 degrees C and b) CO2 binding curves of deoxygenated and oxygenated whole blood and hemoglobin solutions, respectively, at the same temperature . In the presence of CO2 both the Bohr effect and oxygen affinity were significantly lower in 1.3-mM Hb4 solutions than in either red cell suspensions or 5-mM Hb4 solutions, while in the absence of CO2 Bohr effect and oxygen affinity did not differ significantly in those preparations . Likewise, the fraction of oxygen-linked carbamate obtained from CO2 binding curves was found to be higher in 1.3-mM Hb4 (0.156 M HbCO2/M HbO2) solutions than in 5-mM Hb4 solutions (0.12 M HbCO2/M HbO2) at pH 7.2 . We conclude that hemoglobin concentration affects formation of oxygen-linked carbamate . Total oxygen-linked CO2 in sheep whole blood amounted to 0.18 M CO2/M O2 of which 70% is oxygen-linked carbamate . Assuming a respiratory quotient of 0.85, the contribution of oxygen-linked CO2 to carbon dioxide exchange in sheep blood was computed to be 21%.

Can J Physiol Pharmacol, 1975 Aug, 53(4), 592 - 602
Adrenocorticotropin secretion rates following histamine injection in adult and newborn dogs; Cowan JS; The metabolic clearance rate (MCR) for ACTH in adult dogs was previously shown not to vary significantly with varying plasma ACTH concentrations or among dogs . This is confirmed here for pups aged 1-7 days . Hence, ACTH secretion rates can be continuously calculated from a continuous function of plasma ACTH vs . time . Each of seven adult dogs under Nembutal anesthesia received two or three intravenous (i.v.) injections of histamine with increasing doses . The first injections in each dog ranged from 7 to 50 mug/kg, while the last dose was 62-108 mug/kg . A total of 16 injections were given . Twelve pups (two litters of six) aged 1-7 days each received one injection of histamine of 76-116 mug/kg (i.v.) . ACTH concentrations in plasma were determined by an adrenal cell suspension bioassay before, and 6 times after each injection . Nine pups also underwent determinations of their MCR for ACTH, with plateau concentrations determined at three times during an ACTH infusion . Continuous curves of ACTH secretion rates were calculated for all 28 histamine injections, showing that all except the 1-day-old pups secrete considerable ACTH when stressed . Compared to adult dogs, the pups show lower secretion rate peaks and shorter periods of rapid secretion . Changes in plasma glucocorticoids also suggest that the adrenal cortex of newborn dogs can respond to ACTH by increased glucocorticoid secretion.

Can J Physiol Pharmacol, 1975 Aug, 53(4), 531 - 9
An adrenal cell system suitable for long term study of factors affecting steroidogenesis; Price CS et al.; Cell suspensions of normal adult rad adrenals have been prepared by trypsinization and incubated in Ham's nutrient mixture F10 containing horse serum, fetal calf serum, and lima bean trypsin inhibitor . In most experiments culture medium was not changed during incubation . In this system the number of cells fell to 50% after 2 days, then slowly declined to 20% after 1 month of incubation . A corticosterone (B) response was seen to as little as 5 muU of ACTH per millilitre, a concentration which is within the range found in normal rat serum . With maximal stimulation (100 mU ACTH/ml) the rate of accumulation was highest during the first 24 h then slowly decreased over the following 9 days . When in separate experiments ACTH was added after various times of incubation up to 3 weeks, there was a B response which continued for as long as 1 week after the ACTH was added; the later the time at which ACTH was added the lesser was the initial B response and the longer the lag period before a substantial response occurred . In cell suspensions in medium containing 5.0 mequiv . of K+ per litre, aldosterone content increased for approximately 24 h, then showed little or no change over the next 9 days . With increased K+ concentration, aldosterone was found in greater amounts and accumulation continued for longer periods, both without and with ACTH . This adrenal cell system appears suitable for long term study of factors affecting steroidogenesis.

J Membr Biol, 1975 Jul 24, 22(3-4), 357 - 68
Effect of cell concentration on the uptake of amino acids by rat liver parenchymal cells in suspension; Bhargava PM et al.; The accumulation of several amino acids in the acid-soluble fraction and their incorporation into protein in rat liver parenchymal cell suspensions, has been shown to depend on the concentration of cells in the incubation medium; the uptake, both in the acid-soluble and the acid-insoluble fractions, decreased as the cell concentration increased from 0.03 X 10(6) cells/ml upwards, reaching a plateau at high cell concentrations (3-5 X 10(6) cells/ml) . The uptake values at high cell concentrations were the same as those obtained in liver slices in which a similar effect was not observed . Evidence is presented which suggests that this phenomenon is mediated by a material released from the cells in suspension, which is inhibitory to enhancement of the uptake of amino acids by these cells over and above the value obtained in normal, adult liver slices.

J Membr Biol, 1975 Jul 24, 22(3-4), 329 - 40
Comparison of some permeability properties of rat liver slices, liver cells in suspension, and in vivo-produced aggregates of dispersed liver cells; Kumar GK et al.; It has been reported earlier that when rat liver is dispersed to a single cell suspension, the parenchymal cells lose the ability to take up pyrimidine bases but acquire the ability to take up RNAase and macromolecular nucleic acids . It is now shown that these changes are largely reversed on intraperitoneal reaggregation of the parenchymal cells and that, in these respects, the aggregates behave more like the organized tissue than like the dispersed cells.

J Membr Biol, 1975 Jul 24, 22(3-4), 341 - 56
Uptake in vitro of nucleic acid precursors and nucleic acids by Zajdela ascitic hepatoma cells; Bhargava PM et al.; Zajdela ascitic hepatoma cells are shown to take up pyrimidine bases at much lower rates than obtained in slices from normal rat liver . The rates of uptake of adenine and uridine by the Zajdela cells are, however, as high as in the slices . Like the slices, again, the Zajdela cells take up E . coli RNA and DNA at very low rates but, unlike the slices, thses cells degrade rapidly the RNA taken up . The Zajdela cells resemble parenchymal cell suspensions derived from normal rat liver in regard to the uptake of pyrimidine bases and the ability to degrade heterologous RNA.

Int J Cancer, 1975 Jul 15, 16(1), 113 - 24
Human tumor--lymphocyte interaction in vitro: blastogenesis correlated to detectable immunoglobulin in the biopsy; Vanky F et al.; Binding of radioiodinated anti-immunoglobulin (Ig) reagent assayed by acid elutable radioactivity was shown in 18 of 44 cell suspensions (41%) prepared from surgical specimens of human tumors . Aliquots of these biopsies were admixed to autologous lymphocytes and in 13 cases they induced stimulation of DNA synthesis . In only one of these 13 cases was the anti-Ig reagent bound, while among the 26 biopsies with low or no binding capacity 12 (46%) were stimulatory, indicating that immunoglobulin-containing biopsies are not stimulatory . Experiments on 32 lymphocyte preparations from different lymphoid organs suggest that the immunoglobulin detected in the tumor-cell suspension is not derived from the infiltrating lymphoid cells.

Biochim Biophys Acta, 1975 Jul 3, 394(3), 463 - 9
Osmotic behaviour of Acholeplasma laidlawii B cells with membrane lipids in liquid-crystalline and gel state; van Zoelen EJ et al.; The osmotic behaviour of Acholeplasma laidlawii B cells was investigated with combined spectrophotometric and enzymatic measurements . The conclusion could be drawn that this osmotic behaviour depends largely on the physical state of the membrane lipids . When part of the membrane lipids is in the liquid-crystalline phase the cell is able to swell and behaves as a good osmometer . However, when the membrane lipid is in the gel phase, the cell is unable to swell and the change in absorbance of the cell suspension is then completely due to lysis.

Endokrinologie, 1975 Jul, 65(2), 105 - 8
The effects of CBG on the in vitro reduction of 3H-progesterone by liver homogenates and cell suspensions; Gromadzka J et al.; The effects of CBG on 3H-progesterone metabolism by ovine, bovine and rat liver homogenates and bovine and rat liver cell suspensions were studied . As a source of CBG served plasma from "estrogenized" women . Same plasma, pretreated at 60 degree for 20 min . served as control . It was found that the amounts of 3H-progesterone remaining after incubation were higher in the presence of CBG, as compared with control incubations, when using the homogenates . No such effect was observed in case of cell suspensions.

Tohoku J Exp Med, 1975 Jul, 116(3), 241 - 52
Separation of gastric mucosal cells of rat with proteolytic enzymes, pronase and trypsin, with special reference to the collection, morphology and viability of the generative cells; Kurokawa Y et al.; Methods to separate and collect gastric mucosal cells of the rat using proteolytic enzymes were devised . Pronase (1.0%) achieved better results than did trypsin (2.0%) in collecting single isolated cells with higher cell yields and viability . The cells dissociated with trypsin retained glandular structures as in situ . The measurement of radioactivity revealed that the incorporation of 3H-thymidine into generative cells was highest in the cell suspension collected by the second 15 min dissociation . It was concluded that the most effective method to obtain dissociated cells from the generative zone of the mucosa is to collect the cells dissociated with 1.0% pronase continuously for a period from 15 to 45 min after the start of dissociation . On autogradiographic analysis with 3H-thymidine, the ratio of generative cells was 10%, approximately 3 X 10(5) cells, in the specimens.

Mol Biol Rep, 1975 Jul, 2(2), 119 - 27
Poly(A)-containing RNA from Petroselinum hortense: isolation, properties and messenger function in vitro; Ragg H et al.; Cell suspension cultures from parsley (Petroselinum hortense Hoffm.) were labelled in vivo with {2-3H} adenosine . The RNA isolated from the ribosomal pellet was fractionated on an oligo(dT)-cellulose column . Approximately 1.5% of the RNA, representing about 15% of the total radioactivity, was retained at high salt concentrations and eluted at low ionic strength . As determined by two independent methods, this fraction contained poly(A) segments with an average length of about 80 nucleotides . It was active as template in a cell-free system from wheat germ, directing the synthesis of peptides ranging in molecular weight from about 4000-40000 daltons.

J Bacteriol, 1975 Jul, 123(1), 36 - 40
Are thiosulfate and trithionate intermediates in dissimilatory sulfate reduction?
Chambers LA, Trudinger PA.
The fate of 35-S during anaerobic metabolism of {35-S}sulfate, {35-S}thiosulfate, and {35-S}sulfate plus unlabeled thiosulfate by washed cell suspensions of Desulfovibrio spp, and of {35-S}thiosulfate by growing D . desulfuricans was examined . The results appear to be inconsistent with the hypothesis that thiosulfate is an intermediate in sulfate reduction . Since thiosulfate was produced from trithionate, the latter is also unlikely to be an intermediate in the reduction pathway . Extracts of D . desulfuricans catalysed exchange between sulfite and the sulfonate group of thiosulfate.

Endocrinology, 1975 Jul, 97(1), 39 - 45
Calcitonin secretion by monolayer cultures of human C-cells derived from medullary thyroid carcinoma; Roos BA et al.; Monolayer cultures of human calcitonin-secreting cells (C-cells) have been derived from medullary thyroid carcinomas . The cultures were prepared from cell suspensions obtained by enzymatic digestion of surgical specimens of the tumor . Human calcitonin (hCT) secretion by these cells was studied using a specific radioimmunoassay for the hormone . The cultures could be used for reproducible secretion studies up to 40 days after their initiation; they demonstrated a linear rate of hormone secretion in the basal state for at least 4 h . Calcium, pentagastrin, and prostaglandin E2 (PGE2) each produced a marked increase (up to 7-fold) in hormone secretion . Magnesium had no apparent secretory effect; and compared to PGE2, PGF2alpha had only a small secretory effect . In addition to responding to specific secretagogues shown to regulate calcitonin secretion in vivo, the secretory effects of each of the secretagogues could be raipdly reversed . Therefore, these cultures of human C-cells exhibit secretory responses which are quantitatively and qualitatively similar to C-cells in vivo . Accordingly, such cultures provide a useful model to study the regulation of calcitonin secretion in human C-cells.

J Microsc, 1975 Jul, 104(2), 121 - 5
Immersion fixation of rapidly frozen, untreated tissues and suspensions of cells including spermatozoa; Cummins JM et al.; Small pieces of tissue, and cell suspensions in plastic artificial insemination (AI) 'straws', were frozen rapidly in Freon-12 at -155 degrees C, without pre-treatment . Peripheral fragments were thawed directly in 2% glutaraldehyde at 0 degrees C, and processed for transmission electron microscopy (TEM) studies . Preservation of ultrastructure was satisfactory, and freezing artifacts were minimal.

J Clin Invest, 1975 Jul, 56(1), 98 - 110
Human bone marrow lymphocytes . I . Distribution of lymphocyte subpopulations in the bone marrow of normal individuals; Fauci AS; This study was undertaken to determine the proportions and in vitro immune capacities of lymphocyte populations in the bone marrows of normal humans . Relatively pure mononuclear cell suspensions were obtained from bone marrow aspirates by linear sucrose gradient centrifugations . Simultaneous peripheral blood and bone marrow specimens from each individual were assayed for lymphocyte surface markers and mitogen responsiveness . Maximal possible contamination of bone marrow aspirates by peripheral blood was determined by performing aspirates on individuals who had received 51chromium-labeled autologous erythrocytes . Rhymus-derived (T) lymphocytes, as determined by the sheep red blood cell (E) rosette assay, comprised 8.6-(plus or minus 1.6)% of the total bone marrow lymphocyte pool . Bone marrow-derived (B) lymphocytes, as determined by the presence of a complement receptor, made up 15.4-(plus or minus 1.9)% of the lymphocyte pool whereas 74.6 (plus or minus 2.4)% of mononuclear cells lacked easily detectable surface markers . These findings could not be explained by contamination with peripheral blood lymphocytes since contamination was corrected for in the calculations . Lymphocyte-enriched suspensions of bone marrow cells responded to stimulation with phytohemagglutinin, concanalin A, and particularly pokeweed mitogen . In vitro incubations of bone marrow and peripheral blood lymphocytes with tritiated thymidine followed by determinations of E and erythrocyte antibody complement (EAC) rosettes were performed . Simultaneous rosetteradioautographs demonstrated that the proliferative potential of bone marrow B lymphocytes was greater than peripheral blood B lymphocytes (P less than 0.01) . On the other hand, the proliferative potential of bone marrow T lymphocytes was the same as that of peripheral blood T lymphocytes . These findings demonstrate that in addition to containing B lymphocytes the normal bone marrow contains a small fraction of T lymphocytes similar to the mature T lymphocyte pool found in the peripheral blood . These T cells most probably enter the bone marrow parenchyma as part of the normal recirculating lymphocyte pool.

Am J Pathol, 1975 Jul, 80(1), 79 - 90
Separation of cells with histochemically demonstrable acid phosphatase activity from suspensions of human prostatic cells in an isokinetic gradient of Ficoll in tissue culture medium; Helms SR et al.; Collagenase, trypsin, and pronase were used in separate, parallel experiments to obtain cell suspensions from human prostates obtained from surgical resections and autopsies . All of the examined prostates demonstrated benign hyperplasia . The dissociation using pronase gave both the largest number of nucleated cells and the largest proportion of viable cells from prostates . Surgically resected prostates gave a larger number of cells per gram of tissue than prostates obtained from autopsy . Cells obtained from surgically resected prostates were separated both by isopycnic sedimentation and by velocity sedimentation in a previously described isokinetic gradient . We studied fifteen prostates obtained at surgery; using pronase, we obtained 2.1 plus or minus 3.5 times 10(6) cells/g . Of these cells, 34.0 plus or minus 14.7% contained histochemically demonstrable acid phosphatase . Cells from six prostates were separated by velocity sedimentation in the isokinetic gradients; in the purest fractions from the isokinetic gradients, 81.0 plus or minus 12.2% of nucleated cells had histochemically demonstrable acid phosphatase . The cells in the purest fractions appeared to be epithelial cells . More than 99% of these separated cells excluded trypan blue . Isopycnic sedimentation was not an effective means of purifying epithelial cells from human prostates.

Clin Sci Mol Med, 1975 Jul, 49(1), 13 - 26
Relation of intracellular K+ and steroidogenesis in isolated adrenal zona glomerulosa and fasciculata cells; Mendelsohn FA et al.; 1 . Intracellular K+ content, water spaces and corticosterone output were measured in isolated zona glomerulosa and zona fasciculata-reticularis cell suspensions of rat adrenal cortex, after incubation in vitro under conditions designed to alter steroidogenesis . 2 . Intracellular K+ of unpurified zona glomerulosa cells was not altered after stimulation of corticosterone output with serotonin . Similarly, with zona glomerulosa cells purified by unit gravity sedimentation, no change in intracellular K+ was detected after stimulation of steroidogenesis with serotonin or angiotensin II . 3 . In high-potassium medium (final concentration 8.4 mmol/1), parallel increases in intracellular K+ and corticosterone output were observed with both purified zona glomerulosa cells . However, a similar increase in intracellular K+ also occurred in high-potassium medium with zona fasciculata cells, whose steroid output is unresponsive to external potassium concentration ({K+}) . 4 . Ouabain at 10(-5) mol/1 depressed the intracellular {K+} of glomerulosa cells but did not alter basal or stimulated corticosterone output . Similar results were obtained with fasciculata cells . 5 . Ouabain at 5 times 10(-4) mol/1 further depressed intracellular {K-+} of glomerulosa cells and inhibited basal and stimulated corticosterone output . However, this concentration of ouabain also inhibited steroidogenesis in fasciculata cells . 6 . These results demonstrate a variety of situations where changes in intracellular {K+} are dissociated from those in corticosterone output and indicate that intracellular {K+} cannot be the sole mechanism regulating steroidogenesis under these conditions.

Arzneimittelforschung, 1975 Jul, 25(7), 1042 - 8
{Sensitivity tests of malignant tumours against cytostatic agents in vitro and in vivo/studies on the mouse sarcoma 180 (author's transl)}; Volm M et al.; The effects of ten different substances on the mouse sarcoma 180 have been compared using in vivo and in vitro test systems . The size of the tumours was taken as a measure of the success of the therapy in animal experiments . The in vitro effects were estimated by measuring the incorporation of radioactively labelled uridine in tumour cell suspensions from solid tumours, in ascites tumours and in tissue culture . Similar results were obtained using all three in vitro test systems . Four substances (daunomycin, fluoruracil, actinomycin D, adriamycin) exhibited activity both in vivo and in vitro, whereas five substances (cytosinarabinoside, methotrexate, ibenzmethyzin, triaziquone, bleomycin) showed no activity in any of the test systems used . With podophyllic acid ethyl hydrazide, however, no correlation between in vivo and in vitro effects was observed . Studies on the influence of the cytostatic agents on the rate of transport of uridine into the cells showed that podophyllic acid ethyl hydrazide strongly reduced the permeability of the cells to radioactive uridine.

Ann N Y Acad Sci, 1975 Jun 30, 254, 243 - 53
Comparative study of membrane and intracytoplasmic immunoglobulin classes in human lymphoid cells; Ferrarini M et al.; Immunofluorescent and "in vitro" biosynthetic techniques have been used to study the connections between the maturation process of human lymphocytes that leads to the appearance of actively secreting cells and a possible switchover from IgD to IgM production, both in homogeneous CLL cell populations and in heterogeneous cell suspensions from tonsils . The results obtained are compatible with the hypothesis that the switchover from IgD to IgM productions can be a clonal maturation phenomenon.

Nippon Naibunpi Gakkai Zasshi, 1975 Jun 20, 51(6), 561 - 72
{Steroidogenesis in isolated adrenal cells of rat . -Effects of cyanoketone on pregnenolone and corticosterone production- (author's transl)}; Bannai C; In order to study the effect of cyanoketone on steroidogenesis of rat adrenal, the assay technique for corticosteroids released into the incubated media of the rat adrenal cells treated with collagenase was basically investigated . Corticosterone was measured by fluorometric method and pregnenolone by radioimmunoassay . Reliability of radioimmunoassay was satisfactory . About 400,000 cells were obtained from one adrenal gland of male or female rats and sex-dependent difference in pregnenolone and corticosterone production in response to ACTH was not found . Net corticosterone production by isolated adrenal cells was related to the log of the concentration of ACTH by a sigmoid curve over the range 1 to 1000 muU/ml . The half-maximum response was observed at an ACTH concentration of 10 muU/ml, and maximum corticosterone production responding to ACTH (100-1000 muU/ml) was about 5 mug/adrenal/120 min . When cell suspensions were incubated with 1000 muU/ml of ACTH, the conversion from pregnenolone to corticosterone was inhibited 50% by cyanoketone at a concentration of 2 times 10(-8) M . The conversion was completely inhibited at a concentration of more than 10(-7) M . Cyanoketone up to a concentration of 10(-5) M seemed to have no inhibitory effect on cholesterol side-chain cleavage . In the absence of ACTH significant amount of pregnenolone was formed (about 60 ng/adrenal) by isolated adrenal cells obtained from normal adult female rats during incubation with 10(-7) M of cyanoketone for 60 min . To eliminate the possibility of the effect of endogenous ACTH which might be present in incubation medium, cell suspensions were obtained from hypophysectomized female rats . Incubations were carried out in the same condition as mentioned above and significant amount of pregnenolone was formed by cell suspension, which was about 35 ng/adrenal.

Eur J Biochem, 1975 Jun 16, 55(1), 315 - 21
Stimulation of tumor-cell respiration by inhibitors of pyruvate kinase; Gosalvez M et al.; In a model system consisting of highly coupled rat liver mitochondria respiring in the presence of substrate, pyruvate kinase, phosphoenolpyruvate, ATP, hexokinase and glucose, the increase in the mitochondrial concentration results in a progressive decrease in the activity of pyruvate kinase . These results are in accord with a role of pyruvate kinase as a determinant of glycolytic activity by competing with mitochondrial oxidative phosphorylation for the available ADP . The addition of adequate amounts of the amino acids, cysteine, alanine and phenylalanine, known as inhibitors of pyruvate kinase, to living Ehrlich ascites tumor cell suspensions results in a stimulation of the respiratory rate and in a decrease of the glycolytic rate of the cells . Concomitant with these changes, there is an accumulation of intracellular phosphoenolpyruvate and ADP, and a decrease in pyruvate and ATP . These results provide additional evidence for paying attention to pyruvate kinase as another key enzyme whose properties and activities may be major determinants for the control of glycolysis and the Crabtree and Pasteur effects of tumor cells.

Experientia, 1975 Jun 15, 31(6), 699 - 700
Simple method for a medium-cell separation; Jurovcik M; The present method makes possible a quick separation of cells from the medium with the aid of strips of filter paper and physiological solution . Cell suspension is put on the strip and all water soluble components are washed away by soaking in saline solution, while cells remain on the spot . The experiment on 14-C-uridine uptake proved the suitability of the method for membrane-transport studies.

C R Acad Sci Hebd Seances Acad Sci D, 1975 Jun 2, 280(21), 2485 - 8
{Kinetic, radiographic, and histologic demonstration of the curing of congenital osteopetrosis in rats}; Milhaud G et al.; In the "op" rat congenital osteopetrosis is a lethal condition . The management of this disease can be achieved by a single injection of a marrow cell suspension from a normal donor or by temporary parabiosis with a normal Rat, the latter procedure being less efficient than the former according to kinetic, roentgenographical and histological examinations.

Am J Physiol, 1975 Jun, 228(6), 1835 - 9
Renin secretion from rat renal cortical cell suspensions; Lyons HJ et al.; Renin secretion of rat renal cortical cell suspensions was studied as a functionof variations in the composition of the suspension media . Renin secretion was determined by incubating samples of the suspension media with rat renin substrate and measuringthe angiotensin I generated by radioimmunoassay . Increasing the medium sodium concentration (50-144 meq/liter) linearly decreased the rate of renin secretion . This relationship was unaffected by adding ouabain (10 minus 3 M) to the suspensions . The addition of furosemide (from 10 minus 5 to 10 minus 3 M) stimulated renin secretion . Considered with previous observations, these results suggest that renin secretionis controlled by some function of sodium on the lumenal boder of the macula densa cells.

Eur J Biochem, 1975 Jun, 54(2), 493 - 8
Spectroscopic evidence for the formation of a 4-keto intermediate in the UDP-apiose/UDP-xylose synthase reaction; Gebb C et al.; Uridine diphospho-D-glucose (UDP-Glc) and UDP-methyl-D-glucuronate (UDP-GlcUAMe) have been shown to be competitive inhibitors for the UDP-apiose/udp-xylose synthase from cell suspension cultures of parsley . The apparent Ki values for these substrate analogues were of the same order of magnitude as the apparent Km value for the substrate UDP-D-glucuronic acid (UDP-GlcUA) . The difference spectrum of the incubation mixture containing UDP-GlcUA, NAD+ and the highly purified enzyme showed a transient absorption with a maximum at 292 nm which disappeared upon addition of sodium hydroxide . The incubation with UDP-Glc or UDP-GlcUAMe also showed and NAD+-dependent absorption at 292 nm . However, in these cases a strong enhancement of the absorption at alkaline pH and a shift of the absorption maximum to longer wavelength were observed . Upon addition of 0-phenylenediamine to the enzyme incubation with UDP-Glc or UDP-GlcUAe a strong absorption with a maximum at respectively 335 and 315 nm appeared . The results show the transient formation of a 4-keto derivatives in the synthase reaction with the normal substrate UDP-GlcUA . The substrate analogues UDP-Glc and UDP-GlcUAMe can also be oxidized at C-4 by the enzyme in the presence of NAD+ to stable 4-keto derivatives which give rise to a strong absorption at alkaline pH or after reaction with o-phenylenediamine.

J Bacteriol, 1975 Jun, 122(3), 1351 - 63
Oxidation of C1 Compounds by Particulate fractions from Methylococcus capsulatus: distribution and properties of methane-dependent reduced nicotinamide adenine dinucleotide oxidase (methane hydroxylase); Ribbons DW; Cell-free particulate fractions of extracts from the obligate methylotroph Methylococcus capsulatus catalyze the reduced nicotinamide adenine dinucleotide (NADH) and O2-dependent oxidation of methane (methane hydroxylase) . The only oxidation product detected was formate . These preparations also catalyze the oxidation of methanol and formaldehyde to formate in the presence or absence of phenazine methosulphate with oxygen as the terminal electron acceptor . Methane hydroxylase activity cannot be reproducibly obtained from disintegrated cell suspensions even though the whole cells actively respired when methane was presented as a substrate . Varying the disintegration method or extraction medium had no significant effect on the activities obtained . When active particles were obtained, hydroxylase activity was stable at 0 C for days . Methane hydroxylase assays were made by measuring the methane-dependent oxidation of NADH by O2 . In separate experiments, methane consumption and the accumulation of formate were also demonstrated . Formate is not oxidized by these particulate fractions . The effects of particle concentration, temperature, pH, and phosphate concentration on enzymic activity are described . Ethane is utilized in the presence of NADH and O2 . The stoichiometric relationships of the reaction(s) with methane as substrate were not established since (i) the presumed initial product, methanol, is also oxidized to formate, and (ii) the contribution that NADH oxidase activity makes to the observed consumption of reactants could not be assessed in the presence of methane . Studies with known inhibitors of electron transport systems indicate that the path of electron flow from NADH to oxygen is different for the NADH oxidase, methane hydroxylase, and methanol oxidase activities.

J Immunol Methods, 1975 Jun, 7(2-3), 291 - 300
A procedure for removing red cells and dead cells from lymphoid cell suspensions; Davidson WF et al.; A procedure is described for simultaneously removing red cells and dead cells from lymphoid cell suspensions, based on the observation that when populations of lymphoid cells are centrifuged on a mixture of Isopaque/Ficoll, dead cells and red cells sediment whereas viable cells float . The technique very efficiently removed red cells from a wide range of lymphoid cell suspensions and eliminated lymphocytes killed by mechanical stress, by antibody and complement and by prolonged tissue culture . The depletion of red cells was greater than 99% and the recovery of viable lymphocytes usually greater than 90%, the resulting cell suspensions being around 95-100% viable . The immunological activity of B cells, helper T cells and cytotoxic T cells virtually unimpaired by the separation procedure.

Transplantation, 1975 Jun, 19(6), 495 - 504
The human mixed lymphocyte-endothelium culture interaction; Hirschberg H et al.; Cells separated from the wall of the umbilical cord vein by collagenase digestion could be identified as endothelial by their characteristic ultrastructure, their growth pattern in culture, and their microscopical morphology . These cells, both freshly explanted and after long-term culturing, were capable of stimulating allogeneic lymphocytes in vitro . Control experiments indicated that this stimulation was not attributable to contamination of the endothelial cell suspensions by foetal fibroblasts or passenger lymphocytes . The dose response characteristics and kinetics of the lymphoproliferative response using endothelial stimulating cells was similar to mixed lymphocyte cultures . Sera which were capable of inhibiting the mixed lymphocyte culture response were relatively ineffective in inhibiting the stimulation caused by endothelial cells.

Metabolism, 1975 Jun, 24(6), 681 - 89
The mechanism of the potentiating effect of glucocorticoids on catecholamine-induced lipolysis; Lamberts SW et al.; Preincubation of isolated epididymal fat cells with dexamethasone or treating rats with cortisol enhances the epinephrine-stimulated lipolysis of the cells as well as cAMP-dependent protein kinase activity in homogenates of these fat cell suspensions . During maximal inhibition of phosphodiesterase activity by theophylline or dibutyryl cAMP, this potentiating effect of glucocorticoids on the fat cells was also present . There was no lowering of the total phosphodiesterase activity in homogenates of fat cell suspensions of rats that were treated with cortisol, but there appeared to be a lower activity of the low KM phosphodiesterase activity . It is concluded that induction of protein kinase by glucocorticoid hormone is responsible for its special type of stimulative action on lipolysis.

J Natl Cancer Inst, 1975 Jun, 54(6), 1361 - 7
Alpha fetoprotein: effect of heterologous antiserum on hepatoma cells in vitro; Mizejewski GJ et al.; Hepatoma cells derived from The Jackson Laboratory mouse hepatoma BW7756 synthesized alpha fetoprotein (AFP) in vitro . The AFP was immunologically identical to that circulating in the sera of hepatoma-bearing mice . An in vitro cytotoxic effect of rabbit antiserum to AFP was studied in hepatoma cells obtained both from fresh cell suspensions and short-term cell culture . The use of intact and/or inactivated anti-AFP serum inhibited the growth of the AFP-producing cells . The cytotoxic effects of the antiserum depended on exposure time and serum concentration . The cytotoxicity was complement independent, as demonstrated by studies with heat-deactivated serum devoid of extrinsic complement . The control target cells included fresh cell suspensions of normal mouse liver and mouse muscle fibroblasts grown in short-term culture . Specificity of the antisera for the target cells was demonstrated by absorption with purified mouse AFP . The results could be explained by the presence of AFP on the hepatoma cell surface.

Immunology . 1975 Jun;28(6):1067.
Release of histamine from rat mast cells by the complement peptides C3a and C5a; Johnson AR et al.; Suspensions of rat mast cells were used to study the histamine-releasing actions of anaphylatoxins C3A and C5a in vitro . The peptides, derived from human or porcine complement proteins C3 and C5, were less potent than 48/80 but more potent than bradykinin in stimulating release of histamine from mast cells . The pattern of release resembled that of the anaphylactic release action, e.g . release was limited to less than 30 per cent of the cell histamine, the reaction was calcium-dependent and was potentiated by phosphatidyl serine . When C3a and C5a were added together to mast cell suspensions, the amount of histamine released was additive . Similarly, release by either peptide combined with bradykinin was additive . Histamine-releasing activity (as well as smooth muscle-stimulating activity) was abolished when the peptides were treated with pancreatic carboxy-peptidase B . Active or inactive peptides were bound by mast cells and addition of active C3a in combination with the inactive, des-arginine derivativ, C3ai, resulted in partial inhibition of histamine release.

Biochim Biophys Acta, 1975 May 5, 392(1), 131 - 40
A quantitative assay for concanavalin A- and Ricinus communis agglutinin-mediated agglutinations of rat ascites hepatoma cells . Relationship between concanavalin A binding and cell agglutination; Kaneko I et al.; A simple quantitative assay method was developed for the agglutination of rat ascites hepatoma cells mediated by Concanavalin A or Ricinus communis agglutinin . This method was based on the principle that the turbidity of a cell suspension is proportional to the sum of the cross-sectional area of cells and aggregatesmas predicted by the theoretical consideration, the turbidity decreased when cells were aggregated and the decrease was a function of the average number of the cells in aggregates . The agglutinability of the cells, judged by this method, showed a maximum value at a certain concentration of the agglutinin . By further addition of the agglutinin, the agglutinability slightly decreased from the maximum . These phenomena were observed both for Concanavalin A and Ricinus communis agglutinin . The binding and the agglutination experiments using {3-H}concanavalin A revealed that the binding to approx;0% of the total receptors caused a maximal agglutination . This suggested that the receptors responsible for the agglutination constitute only a small part of the total receptors on the surface.

Acta Cytol, 1975 May-Jun, 19(3), 306 - 12
Preparing cell suspensions from cervical smears with pepsine and ultrasonic treatment; Freni SC et al.; Since flow-through fluorometry seemed to be a workable method for prescreening cytological material, it became important to have available a reliable method of preparing suspensions of single cells with naked nuclei . An experiment was performed with cervical smears and tissue cultures exposed to varying degrees of pepsination and ultrasonication . The results were disappointing as it appeared impossible to digest cytoplasm completely without damaging the nucleus . Ultrasonic treatment appeared to have an effect only in combination with pepsin; sonication therefore is not a useful technique for dispersion of cell clusters . Moreover, a proposal found in the literature to apply sonication to effect selective damage to leukocyte nuclei was assessed but found to be unsuccessful . Pepsin treatment and ultrasonic treatment appeared conclusively to be unreliable methods for preparing suspensions of single and naked nuclei.

J Immunol, 1975 May, 114(5), 1631 - 7
Characterization of mouse cells releasing or responding to mitogenic factor induced by phytomitogens in vitro; Jacobsson H et al.; Mouse lymphocyte populations exposed to mitogen were tested for their capacity to release factors that stimulate other lymphocytes to synthesize DNA or enhance their response to mitogens in vitro . The target lymphocyte for this mitogenic factor(s) (MF) was also characterized . The results showed that lymph node and spleen cells release more MF than thymocytes upon exposure to Con A, PHA or PWM in vitro . The T cells were found to be largely responsible for MF production, since cell suspensions depleted of phagocytic cells did not exhibit any decreased ability to produce MF and spleen cells from congenitally athymic (nude) mice produced no detectable MF activity . The MF stimulated thymocytes, lymph node cells, and spleen cells to synthesize DNA . Spleen cells from nude mice were also stimulated . The MF released by lymphocytes in response to Con A was found to induce DNA synthesis of lymphocytes in itself and did not require the presence of mitogen . It is concluded that phytomitogen lectins stimulate T cells to synthesize DNA and to release soluble factor(s) which are mitogenic for both T and B cells . The latter cells may thus be unresponsive to the phytomitogen, but still undergo blast transformation.

Rev Rhum Mal Osteoartic, 1975 May, 42(5), 309 - 16
{Lymphocytic subpopulations implicated in the rheumatoid rosette phenomenon}; Sany J et al.; The significance of the phenomenon of rheumatoid rosettes (RR) was studied in 21 subjects suffering from rheumatoid polyarthritis (RP) and in 16 controls . Certain technical factors influenced the prevalence of RR . Thus the number of RR increased in the RP patients and the controls with increase in the number of sensitized 0 Rhesus - red corpuscules; nevertheless there was a significant difference between the results obtained with the RP patients and the controls . The length of incubation and the temperature played an important role: prolonged contact times (30 min to one night), particularly at 4C considerably increased the numbers of RR, which reached 10 per cent both, in the controls and in the RP patients . The separation of lymphocytes by filtration through nylon according to Greaves' technique was followed by a study of the T cells (E rosettes), the B cells (surface immunoglobulins), and the RR . In all cases (RP and controls), the numbers of RR increased after filtration through nylon which retained the B cells . Separation of the lymphocytes by the centrifuging of the E rosettes showed that the proportion of RR was clearly higher in a population rich in B lymphocytes and lower in a population rich in T lymphocytes . Futhermore, filtration through nylon of cell suspensions rich in B lymphocytes showed a considerable rise in the RR and parallel drop in the number of B cells . Reduction of the RR by centrifuging did not affect the numbers of T or B cells . In the light of these results, it seems that the cell responsible for the formation of the RR does not have the characteristics of either the T or the B lymphocytes: a lymphocyte carring a receptor for the fragment Fc of the immunoglobulins (K cells) could be involved . In this case, the RR should be related to the EA rosettes of which they perhaps represent and active fraction.

Cell, 1975 May, 5(1), 11 - 7
D-valine as a selective agent for normal human and rodent epithelial cells in culture; Gilbert SF et al.; A nutrient medium has been developed to enable the growth of normal epithelial cells while selectively inhibiting fibroblast proliferation . In this medium, D-valine is substituted for L-valine; and only those cells containing D-amino acid oxidase can convert the D-amino acid into its essential L-enantiomer . The ability to select for cells with this enzyme has enabled us to maintain epithelial cell populations free from fibroblast overgrowth . The presence of D-amino acid oxidase has been histochemically confirmed in the epithelial cells selected from renal cell suspensions and explants . The ability to proliferate in the selective medium is transmitted to the clonal progeny of these cells . Moreover, epithelial cell proliferation of this medium indicates the presence of D-amino acid oxidase, which we have detected in tissues where it had not previously been reported-fetal human kidney, lung, and cord . Fibroblasts will not grow in the selective medium, but will proliferate normally if the product of the D-amino acid oxidase reaction is supplied.

Am J Physiol, 1975 May, 228(5), 1589 - 96
Effect of temperature on rate of CO2 uptake by human red cell suspensions; Holland RA et al.; The time course of carbon dioxide uptake by oxygenated supensions of human red cells was followed using a CO2 electrode in a Hartridge-Roughton continuous-flow rapid-reaction apparatus . Measurements were made at several temperatures from 1i to 42 degrees C, with the initial PCO2 in the reacting mixture from 40 to 60 mmHg . The initial part of the uptake curve is presumably rate limited by the intracellular hydration of CO2 with reaction-velocity constants in cell water from 280 to 960 S-1 at 42 degrees C and an activation energy of 2.4 kcal mol-1 . The later stages of CO2 uptake were much slower, with half-times from greater than 1.5 s at 12 degrees C to 0.7 s at 42 degrees C, and were presumably rate limited by the chloride-bicarbonate shift and H+ interchanges . The results indicate that despite the acceleration of the hydration reaction in cell water by a factor of 5,000 at 37 degrees C and 3,800 at 42 degrees C, the later part of the exchange is too slow to permit blood to come intoC02, equilibrium with actively exercising muscles during its passage through the capillary bed.

Mayo Clin Proc, 1975 May, 50(5), 271 - 8
Gluconeogenesis in isolated hepatic parenchymal cells . VII . Effects of monobutyryl cyclic adenosine monophosphate on gluconeogenic intermediates, phosphofructokinase, and fructose diphosphatase; Veneziale CM et al.; Isolated parenchymal hepatocytes prepared from fasted (24 hours) rats convert added dihydroxyacetone and xylitol to glucose . Monobutyryl cyclic adenosine-3',5'-mono-phosphate (mb-cAMP) stimulated the rate from dihydroxyacetone (plus 32%; N equals 28) but not from xylitol (plus 4%; N equals 27) . Iodoacetate (0.15 to 0.20 mM) was an effective inhibitor of gluconeogenesis from lactate-pyruvate mixtures (72% inhibition); quinolinate (2.5 to 3.0 mM) was relatively ineffective (26% inhibition) . Measurements of glucogenic intermediates formed from added dihydroxyacetone in hepatocytes (20% cell suspension) preincubated with iodoacetate provided evidence that phosphoglyceraldehyde dehydrogenase was inhibited . Inhibition of additional glycolytic-gluconeogenic enzyme(s) involved in dihydroxyacetone metabolism was not disclosed by the metabolite concentration data . Because iodoacetate partially inhibited gluconeogenesis from dihydroxyacetone in 5% cell suspensions but did not prevent stimulation of gluconeogenesis by mb-cAMP (plus 31%), additional inhibition, probably nonspecific, occurred . With dihydroxyacetone as substrate, mb-cAMP had only a slight effect on the concentration of fructose diphosphate (decrease) relative to control experiments (no mb-cAMP), but preinculation of the cells with iodoacetate made the mb-cAMP-induced decrease much greater . mb-cAMP caused a 47% decrease (SE, 8;N equals 9) in the assayable activity of phosphofructokinase in liver cells incubated with dihydroxyacetone but did not alter fructose diphosphatase activity significantly . Cyclic GMP and cIMP were shown to stimulate gluconeogenesis from dihydroxyacetone; both nucleotides also cause a decrease in the assayable activity of phosphofructokinase . With xylitol as substrate, mb-cAMP did not cause stimulation of gluconeogenesis, decrease in fructose diphosphate concentration, or decrease in phosphofructokinase activity . Our results indicate that phosphofructokinase might be an important controlling enzyme in gluconeogenesis and subject to regulation by cAMP.

J Immunol, 1975 May, 114(5), 1473 - 9
Modulation of cyclic AMP in purified rat mast cells . I . Responses to pharmacologic, metabolic, and physical stimuli; Sullivan TJ et al.; The cyclic adenosine 3', 5'-monophosphate (cAMP) content of isolated unstimulated mast cells and the changes induced by a variety of pharmacologic, metabolic, and physical stimuli were studied . A modified bovine serum albumin density gradient purification method consistently provided mast cell preparations which were 95% or more pure, without apparent damage, and a 73% recovery of the mast cells applied to the gradients . The measured cAMP in unstimulated mast cells was high, a mean of 16 picomoles per million cells . Moderate agitation or contact with glass increased cAMP content about 2-fold . When calcium was omitted from the medium mast cell cAMP was markedly elevated; incremental increases in added calcium ion concentration from 1 muM to 1 mM caused a linear decrease in cAMP content . Theophylline (3 to 20 mM) caused a dose-related increase in mast cell cAMP content, approximately 2-fold at 20 mM theophylline . Epinephrine (0.01 to 1 mM) caused a modest, dose-related increase in cAMP . In the presence of theophylline, epinephrine increased cAMP levels equal to or greater than the sum of the effects of the agents used individually . The increase in cAMP induced by epinephrine was completely inhibited by 100 muM propranolol and partially inhibited by 10 muM propranolol, thus suggesting that a beta adrenergic receptor is involved . Prostaglandin E1 (PGE1) and histamine (in the presence of theophylline) also raised cAMP . Carbamylcholine (1 nM) lowered cAMP 38%; Atropine (0.1 mM) inhibited the decrease in cAMP induced by 1 nM carbamylcholine by 83% indicating that a muscarinic receptor is involved . In these homogeneous single cell suspensions, therefore, cholinergic and beta adrenergic agents have opposing effects on cAMP levels . Diazoxide (10 muM) and adenine (1 muM) caused 37 and 32% decreases in cAMP, respectively . The availability of highly purified mast cells and the identification of agents which either decrease or increase cAMP content allows a direct examination of the role of cAMP in histamine release.

J Biol Chem, 1975 Apr 25, 250(8), 2769 - 77
The hormonal control of gluconeogenesis by regulation of mitochondrial pyruvate carboxylation in isolated rat liver cells; Garrison JC et al.; The possibility that hormones control hepatic gluconeogenesis via the regulation of the rate of mitochondrial pyruvate carboxylation was investigated with the use of suspensions of liver cells isolated from fasted rats . The mitochondria prepared from liver cells were judged in good condition as they exhibited satisfactory phosphorus-oxygen and respiratory control ratios and transported Ca2+ and K+ ions in an energy-dependent manner . Addition of glucagon, epinephrine, or cyclic adenosine 3':5'-monophosphate to liver cells caused a 50 to 80% increase in the rate of glucose synthesis from lactate . When mitochondria were isolated from the cells after treatment with these agonists, they displayed 2- to 3-fold increases in the rate of pyruvate carboxylation, pyruvate decarboxylation, and pyruvate uptake . These mitochondrial changes are similar to those obtained in hepatic mitochondria prepared from intact, hormone-treated rats . The mitochondrial responses were specific for agents that stimulated gluconeogenesis; no response occurred with 5'-AMP or cyclic adenosine 2':3'-monophosphate . In the cell suspensions, the dose response curves for the activation of mitochondrial pyruvate metabolism and for increased glucose synthesis from L-lactate were coincident with four different agonists . The mitochondrial changes resulting from stimulation with glucagon developed in 1 to 2 min after the rise in cyclic adenosine 3':5'-monophosphate and occurred at least as early as the increase in the rate of gluconeogenesis . When the intracellular level of cyclic adenosine 3':5'-monophosphate returned to basal values, the rates of mitochondrial pyruvate carboxylation and glucose synthesis also declined to control levels . It is concluded that the rate of mitochondrial pyruvate metabolisms can be increased by hormones and cyclic nucleotides and that control of mitochondrial pyruvate carboxylation is an important regulatory site of hepatic gluconeogenesis.

J Biol Chem, 1975 Apr 10, 250(7), 2750 - 5
Induction of delta-aminolevulinic acid synthetase in isolated rat liver cells by steroids; Edwards AM et al.; The role of steroids in regulation of delta-aminolevulinic acid synthetase has been studied in isolated rat liver cell suspensions under conditions previously shown to support inducation of the enzyme by drugs . Addition of a variety of C-19 and C-21 steroids to cell suspensions resulted, after 4 to 6 hours of incubation, in 2- to 5-fold increase in the activity of delta-aminolevulinic acid synthetase as measured in liver cell homogenates . The increase was prevented by cycloheximide . The most active steroid inducers tested were pregnene or pregnane derivatives with keto or hydroxyl groups at C-3 and C-20; in particular a beta-hydroxyl group at C-20 enhanced activity . These C-21 steroids at optimal initial concentrations caused 3- to 5-fold induction over 4 hours . A number of C-19 androstene and androstane compounds caused 2- to 3-fold inducation over the same period . Hydrocortisone had no effect . For a variety of androstane and pregnane derivatives, inducation by 5alphaH steroids was as great as or greater than that by 5betaH compounds, in contrast to previous findings in chick embryo liver . Induction of delta-aminolevulinic acid synthetase by steroids in isolated liver cells was shown to be subject to feedback repression by hemin.

Biochim Biophys Acta, 1975 Apr 7, 385(2), 207 - 20
The effect of p-aminosalicyclic acid on iron transport and assimilation in mycobacteria; Brown KA et al.; p-Aminosalicylic acid inhibits growth of Mycobacterium bovis BCG and Mycobacterium smegmatis more effectively if cells are growing with a sufficiency of iron (more than 1 mu g Fe/ml) in the medium than if cells are deficient in iron (smaller than 0.1 mu g Fe/ml) . In iron-deficient cultures formation of mycobactin, an ionophore for iron transport, is strongly inhibited by p-aminosalicylic acid . Uptake of iron into cell suspensions is also inhibited and the activity of several iron-containing enzymes declines in cells exposed to p-aminosalicylic acid during their growth . p-Aminosalicylic acid is about 50 times more effective towards a mutant of M . smegmatis which required mycobactin under iron-deficient growth conditions than towards the wild-type parent . p-Aminosalicylate is taken up into cells by an active process independent of the salicylate uptake system, possibly by the route used for assimilation of p-aminobenzoate . (This could account for why p-aminobenzoic acid, but not salicylic acid, antagonizes the action of p-aminosalicylic acid.) With iron-deficient cells, salicylate assimilation is about 50 times greater than either p-aminosalicylate or p-aminobenzoate but with iron-sufficient cells and with the mycobactin mutant salicylate uptake is negligible whereas p-aminobenzoate and p-aminosalicylate uptakes are unaffected . p-Aminosalicylic acid at 3.3 mM (500 mu g/ml) partially inhibits the uptake of both p-aminobenzoate and, if it is occurring, that of salicylate as well . As p-aminosalicylic acid is always more effective when the intracellular concentration of salicylic acid is low, it probably acts as an anti-metabolite of salicylic acid, not, however, by inhibiting the conversion of salicylic acid to mycobactin, but probably somewhere along the metabolic pathway of iron uptake.

Biochim Biophys Acta, 1975 Apr 7, 385(2), 421 - 8
A plausible role for a membrane-bound cyclic AMP phosphodiesterase in cellular slime mold chemotaxis; Malchow D et al.; 1 . Kinetics of membrane-bound cyclic AMP phosphodiesterase of the cellular slime mold, Dictyostelium discoideum, were studied under two conditions: in the 27 000 times g sediment of cell homogenates (particle-bound phosphodiesterase) and in cell suspensions using external cyclic AMP as a substrate (cell-bound phosphodiesterase) . Both methods revealed non-Michaelian kinetics with interaction coefficients less than 1 . 2 . The membrane-bound phosphodiesterase has a specificity different from that of the cyclic AMP receptor, also present at the cell surface . 3 . The membrane-bound enzyme was solubilized by lithium 3, 5-diiodosalicylate and partially purified . In this state the non-linear kinetics were still retained; however, the enzyme was not inhibited by the D . discoideum inhibitor, unlike the cell-bound phosphodiesterase in vivo . This indicates that both enzymes share an inhibitor binding site and that this site is cryptic in the cell-bound state . 4 . Production of periodic cyclic AMP pulses by centers, and their relay by other cells, is believed to occur during aggregation . It is suggested that the cell-bound enzyme determines a "time window" significantly smaller than the period of pulsing, and optimizes stimulation of the cyclic AMP receptors in chemotaxis and signal relaying.

Genetika, 1975 Apr, 11(4), 77 - 82
{Development of 0-methylhydroxylamine induced mutations causing resistance to 8-azaguanine in Chinese hamster cells}; Reznik LG; The dependence of the yield of mutations of the resistance to 8-azaguanine induced with 0-methylhydroxylamine on the number of cell generations that have passed by the time of the creation of selective conditions and the existence of a phenotypic lag period is established . The highest rate of mutations is found after 8-azaguanine treatment in two generations . As the number of generations increased, the number of detectable mutations decreased . A dependence is observed between the induced mutation frequency and death of cells as the latter are transferred to selective conditions after two generations . In experiments on induced mutagenesis, one can obtain more exact quantitative characteristics by introducing a selective agent into the cell suspension but not by treating growing cells in colonies.

Antibiotiki, 1975 Apr, 20(4), 342 - 5
{Some characteristics of using polyalcohols as cryoprotectors in preserving Actinomyces noursei LIA-0471}; Konev IuE et al.; Freezing of Act . noursei cell suspensions at a rate of 0.5 degrees per minute in the presence of various substances showed that 5 per cent concentrations of glycerol, ethylen-, diethylen-, propylenglycol and polyethylenglycols had a cryoprotective effect . Polyethylenglycols with a molecular weight of 1500-20 000 had the highest protective effect on the actinomycetous cells from the damaging action of repeated freezing and thawing . The method of repeated freezing and thawing is recommended for studying the cryoprotective effect of various compounds.

Cancer Res, 1975 Apr, 35(4), 932 - 8
Repair of radiation damage in Lewis lung carcinoma cells following in situ treatment with fast neutrons and gamma-rays; Shipley WU et al.; Lewis lung tumor cells were irradiated with 60Co gamma-rays or cyclotron-produced neutrons in situ as solid s.c . tumors or in vitro as single cell suspensions . Cell survival was assayed by colony formation both in vitro in soft agar and in the lungs of isogeneic recipient mice . Survival curve characteristics measured in vitro were: Do = 111 rads, Dq = 342 rads, n = 22 for gamma-rays, and Do = 61 rads, Dq = 46 rads, n = 2 for neutrons . In situ, the hypoxic fraction was 0.36 . Irradiation in situ gave, for the hypoxic subpopulation, Do = 315 rads for gamma-rays and Do = 91 rads for neutrons . The oxygen-enhancement ratio for gamma-rays was 2.8 and for neutrons was 1.5 . Using the split-dose technique, in which two equal doses were administered, separated by 4 hr chronically hypoxic tumor cells repaired sublethal damage, assayed by leaving tumor cells in situ up to 24 hr posttreatment, could not be detected after neutrons, but after gamma-rays it was observed as a 3- to 6-fold increase in survival . The repair of potentially lethal damage increased the relative biological effectiveness of neutrons from 3.7 at a survival level of 5% when assayed immediately after treatment to 4.7 when assayed 6 to 24 hr after treatment . These observations, primarily limited to the chronically hypoxic subpopulation of tumor cells, suggest that decreased repair of potentially lethal damage as well as sublethal damage may be an important radiobiological difference between the effects of high and low linear energy transfer radiation.

Ann Immunol (Paris), 1975 Apr, 126(3), 281 - 9
{Immunological activities of rat lymphocytes . III . --Isolation of differing subpopulations of lymphocytes by two techniques of T cell separation}; Tardieu M et al.; In previous studies we have isolated by density gradient separation a population of relatively dense T lymphocytes capable of inhibiting the mitogenic response to concanavalin A (ConA) and phytohaemagglutinin . In this study we compared two current methods of cell separation for their relative yield in T lymphocytes able or not to respond to Con A . The present results show that cells passed through nylon wool columns--in contrast to those obtained by removal of lymphocytes which bind erythrocyte-antibody-complement complex-partially lose their ability to respond to ConA . The response lost by passage through nylon wool columns can be restored by centrifugation of the T cell suspension on a gradient of "Ficoll-Hypaque" . This suggests that methods of T cells purification using density gradient separation may remove the more dense suppressor cells.

Int J Radiat Biol Relat Stud Phys Chem Med, 1975 Apr, 27(4), 377 - 87
The lung-colony assay: extension to the Lewis lung tumour and the B16 melanoma--radiosensitivity of B16 melanoma cells; Hill RP et al.; Experiments are described which demonstrate that a lung-colony assay can be used to study the viability of unknown cell populations from the B16 Melanoma or the Lewis Lung Tumour . It is shown that the number of lung colonies formed can be increased by the addition of plastic microspheres to the injected cell suspension or by pre-irradiating the lungs of the recipient mice . The colony technique has been used to isolate melanotic and amelanotic cell-lines from the B16 Melanoma which were found to have different growth-rates . In vitro radiation survival curves for B16 Melanoma cells have also been established, and these have parameters in the usual range for mammalian cells.

Exp Hematol, 1975 Apr, 3(2), 94 - 100
Augmentation of marrow growth by thymocytes separated by discontinuous albumin density-gradient centrifugation; Pritchard LL et al.; Mouse thymocytes were separated by discontinuous albumin density-gradient centrifugation . The ability of B6 thymocytes from the gradient fractions to increase the number of spleen nodules formed by B6 marrow cells in B6D2F1 recipients was compared with that of similar numbers of B6 thymocytes from an unfractionated cell suspension . Thymocytes from any of the gradient fractions were not more effective than thymocytes from an unfractionated suspension in augmenting marrow-cell growth, indicating that the presumptive 'effective cell type' in this system cannot be separated from the total thymocyte population by its buoyant density properties.

Tissue Antigens, 1975 Apr, 5(2), 89 - 98
Reactions of kidney cells with cytotoxic antisera: possible evidence of kidney-specific antigens; Perkins HA et al.; HL-A typing of cadaver kidney cell suspension by fluorochromasia cytotoxicity was successful in 76 out of 124 cases . In the 44 cases with confirmed phenotypes, 24 had identical results for lymphocytes and kidney cells . Twenty kidney cells had HL-A antigens not detected on that donor's lymphocytes, most commonly HL-A7 and HL-A8 . In eight of these 20 cases, the additional kidney antigens brought the total to more than two per segregant sereis, in disagreement with an earlier report from this laboratory . The discrepancy was traced to a change in the method of complement preparation . The complement was successfully deprived of the resulting non-specific cytotoxicity for kidney cells by absorption with human red blood cells . Prior to absorption, the complement had rendered kidney cells susceptible to the lytic effect of anti-A and B red cell antibodies . Using the absorbed complement, a patient who had hyperacutely rejected two cadaver kidneys provided sera with an antibody reacting with, and absorbed by, the kidney cells but not the lymphocytes of the donors.

J Immunol, 1975 Apr, 114(4), 1230 - 6
Characterization of immunoglobulin-bearing and other small lymphocytes in mouse bone marrow by sedimentation and electrophoresis; Osmond DG et al.; 125I-antiglobulin binding and radioautography have been used to define the sedimentation and electrophoretic properties of immunoglobulin-bearing and other small lymphocytes in bone marrow . Bone marrow cell suspensions from CBA mice were exposed to 125I-labeled rabbit anti-mouse immunoglobulin for 30 min at 0 degrees C . Cell fractions were collected after either sedimentation at unit gravity or continuous free-buffer film preparative electrophoresis . The sedimentation profiles of labeled, antiglobulin-binding small lymphocytes and of unlabeled small lymphocytes were identical, peaking at 2.6 mm/hr, except for 5% of the labeled small lymphocytes which sedimented at 3.8 to 4.1 mm/hr . In electrophoretic fractions most labeled small lymphocytes were contained in a peak of low mobility and unlabeled small lymphocytes were of intermediate mobility, but high mobility fractions also contained small numbers of labeled and unlabeled small lymphocytes . Small lymphocytes comprised 80 to 85% of all nucleated bone marrow cells in fractions taken at the peak of the small lymphocyte distribution profile after either sedimentation or electrophoresis, accompanied mainly by either erythroid or granulocytic precursor cells, respectively . The results demonstrate that small lymphocytes with readily detectable surface immunoglobulin in the bone marrow closely resemble those in peripheral lymphoid tissues with respect to their sedimentation and electrophoretic properties . The possibility is raised that the rapidly sedimenting immunoglobulin-bearing small lymphocytes in bone marrow are a functionally distinct group of potential precursor cells . Non-antiglobulin-binding small lymphocytes in bone marrow resemble electrophoretically the double negative (immunoglobulin -and theta -negative) small lymphocytes in other lymphoid tissues . Small lymphocytes of high electrophoretic mobility, prominent in the theta- bearing lymphocyte populations of peripheral lymphoid tissues, are few in bone marrow . Lymphocyte-rich suspensions, containing either high or low proportions of immunoglobulin-bearing small lymphocytes, can be separated from bone marrow by electrophoresis

Proc Natl Acad Sci U S A, 1975 Apr, 72(4), 1579 - 83
Modulation of lymphocyte receptor mobility by locally bound concanavalin A; Yahara I et al.; Binding of concanavalin A (con A) to the lymphocyte surface at room temperature leads to restriction of the mobility of a variety of cell surface receptors including those from immunoglobulin (Ig), O, H-2, beta2-microglobulin, Fc receptors results in "co-capping" of Ig, H-2, theta, Fc receptors, as well as receptors for other lectins . Addition of colchicine to the cell suspensions reverses this effect and allows Con A receptors as well as other receptors to form patches and caps . Capping of Con A receptors, and beta2-microglobulin in the absence of ligands specific for these receptors . Receptors binding Limulus hemagglutinin and wax bean agglutinin, as well as those interacting with carbohydrate-specific antibodies, were partially co-capped with Con A but receptors for wheat germ agglutinin were not co-capped, excluding the possibility that restriction of receptor mobility by Con A resulted simply from cross-linkage of mobile receptors to immobilized glycoproteins (Con A receptors) . Latex beads and platelets coupled to Con A were bound to lymphocytes under the same conditions as free Con A . Binding of these particles to local regions of the cell surface resulted in restriction of the mobility of those receptors that could be co-capped with free Con A . In contrast to the findings with free Con A, however, addition of colchicine resulted in capping of the bound particles but did not cause co-capping of either the unbound Con A receptors or other receptors . These findings support the hypothesis that modulation occurs via a submembranous assembly containing microtubules, and they further suggest that the transitions of this assembly induced locally by Con A may be propagated via cooperative processes.

Pediatr Res, 1975 Apr, 9(4), 167 - 71
Studies on tissue factor activity and production by leukocytes of human umbilical cord and adult origin; Rivers RP et al.; Human adult and umbilical cord-derived leukocytes were shown to be capable of generating tissue factor activity on exposure to endotoxin and to reduced pH . Blood for leukocyte separation was collected from normal adults and from newly delivered sections of umbilical cord and mixed leukocyte preparations were obtained by separation over methyl cellulose Hypaque . The coagulant activity of the cell suspension was assayed using a one-stage or two-stage method . Cord-derived leukocytes were shown to develop greater coagulant activity than adult-derived leukocytes when stimulated by endotoxin in vitro at 2,000 cells/mm-3 . This response to endotoxin was partially inhibited by prior exposure of the cells to prostaglandin (PG) E1 an to L-epinephrine . Acetylcholine stimulated the production of coagulant activity in the absence of endotoxin . Both cord and adult-derived leukocytes (20,000/mm-3) developed coagulant activity when exposed to pH reduction by lactic or hydrochloric acids and this activity was shown to be tissue factor.

J Appl Physiol, 1975 Apr, 38(4), 710 - 8
Time course of exchanges between red cells and extracellular fluid during CO2 uptake; Forster RE et al.; A stopped-flow rapid-reaction apparatus was used to follow the time course of extracellular pH in a human red cell suspension following a sudden increase in PCO2 . The extracellular pH change was slow (t1/2 similar to 3.5 s) considering the presence of carbonic anhydrase in the cells . When carbonic anhydrase was added to the extracellular fluid, the half-time was reduced to less than 20 ms . The explanation for these phenomena is that the equilibration of H+ across the red cell membrane is rate-limited by the uncatalyzed reaction CO2 plus H2O formed from H2CO3 outside the cells . A theoretical model was developed which successfully reproduced the experimental results . When the model was used to simulate CO2 exchange in vivo, it was determined that blood PCO2 and pH require long times (greater than 50 s) to approach equilibrium between cells and plasma after leaving an exchange capillary . We conclude that cell-plasma equilibrium may never be reached in vivo, and that in vitro measurements of these quantities may not represent their true values at the site of sampling.

Acta Med Okayama, 1975 Apr, 29(2), 147 - 50
Preparation of single cell suspensions from hepatoma cells in culture; Tokiwa T et al.; Fundamental examination was carried out on the liberation of single cells from hepatoma cells in culture . dRLa-74 cells derived from rat hepatoma, which hardly disperse as single cell suspensions with several proteolytic enzymes or EDTA alone, dispersed with a high yield of single cells by the combination of trypsin and EDTA . HUH-6 cells derived from human hepatoblastoma also showed similar results . The degree of cell dissociation by the combination was dependent on the incubation temperature or pH.

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