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D H Pluznik

Publications and source records attributed to D H Pluznik.

At least 73 records · Page 4Linked to original sources
PubMed1981

Generation of lipopolysaccharide-induced interferon in spleen cell cultures. I. Genetic analysis and cellular requirements.

Incubation of murine spleen-cell cultures with lipopolysaccharide (LPS) induces interferon (IF) production. Maximal IF levels are obtained after incubation with 100 microgram/ml for 10 h. Two inbred mouse strains differing in their ability to generate LPS-induced IF in spleen-cell cultures were used: C3H/eB, which generates high levels of IF (about 60 units/ml), and C3H/HeJ, which fails to generate detectable quantities of IF. In a genetic analysis these strains were hybridized and IF production was determined in spleen-cell cultures from F1 and F2 generations, and from backcrosses of F1 hybrids to parent strains. The results indicate that, in parent strains, a single dominant autosomal gene is responsible for differences in IF production in spleen cultures. LPS-induced IF in spleen-cell cultures resists pH 2 for as long as 48 h, but is labile to heating at 56 degrees C for 30 min. Both macrophages and lymphocytes must be present in cultures for generation of LPS-induced IF. By using mixed cultures of macrophages and lymphocytes from C3H/eB and C3H/HeJ mice, it was shown that macrophages have to interact directly with LPS to enable IF production in the cultures.

Animals↗
PubMed1981

Scanning electron microscope observations on surface changes occurring in murine granulocytes and macrophages during maturation in soft agar cultures.

The present work has been undertaken to study by scanning electron microscopy (SEM) the cell surface changes occurring in granulocytes and macrophages during the process of maturation in soft agar cultures. All stages of maturation of these cells can be seen in a single colony, with immature cells at the center and mature cells toward the periphery. Special emphasis was made, therefore, to maintain the integrity of the colonies during preparation of the specimens for the SEM. Colonies of granulocytes appeared as tight aggregates of cells. The surfaces of cells in such colonies in 3 to 4-day-old cultures contained many microvilli. Cells in colonies of 5 to 7-day-old cultures contained fewer microvilli on their surfaces. Colonies of macrophages appeared as loose aggregates of cells. The surfaces of cells in such colonies in 3 to 4-day-old cultures was slightly ruffled, while the surfaces of cells in 5 to 7-day-old colonies had well developed ruffles. Granulocyte and macrophage colonies showed the same internal organization of the colony: In granulocytes, cells with many microvilli were located in the center of the colony and cells with less microvilli were at the periphery of the colonies. In macrophages, cells slightly ruffled were located in the center of the colony, while cells highly ruffled were at the periphery of the colonies. These observations have been confirmed by transmission electron and light microscopy preparations.

Animals↗
PubMed1979

Growth enhancement and serum replacement in cloning of murine mastocytoma and granulocyte/macrophage precursor cells: two distinct activities present in hemolysates.

In the present study we investigated the serum replacing and enhancing activities of erythrocyte lysates, obtained from different animal species, on the growth of murine mastocytoma cells and normal granulocyte/macrophage precursor cells (CFU-C) present in bone marrow. The soft agar technique for cloning hemopoietic cells in vitro was used to quantitate the growth of these cells in culture. Rat, rabbit, guinea pig, mouse and sheep hemolysates were tested for their capacity to replace or to enhance serum as a growth promoter. All hemolysates except mouse were able to replace serum efficiently when mastocytoma cells were cloned. On the other hand, only rat hemolysate proved to be efficient for cloning normal murine CFU-C. Since increasing the concentration of hemolysates in the soft agar medium did not change these results, the differences in activity are unlikely to reflect quantitative variations. However, hemolysates with little or no serum replacing capacity enhanced the clonal growth of normal CFU-C when added to small amounts (2.5%) of horse serum. Trypsin treatment of rat hemolysate destroyed its serum replacing capacity but not its enhancing activity. No correlation was found between the amount of hemoglobin and the replacing activity of each hemolysate. The data obtained in the present study point to the presence of at least two distinct activities in hemolysate: serum replacing and growth enhancing capacities.

Animals↗
PubMed1978

F20C, a new fluorescent membrane probe, moves more slowly in malignant and mitogen-transformed cell membranes than in normal cell membranes.

New fluorescent probes of membrane mobility can be introduced into cell membranes at single points with particles of a membrane mobility agent, A2C. The initial entry of fluorescence from the particle into the cell membrane and the subsequent lateral spread of fluorescence have been observed for cells in suspension. A dramatic difference between the behavior of normal lymphocytes and that of mitogen-transformed and mastocytoma cells is found. Both the initial entry and the spreading of fluorescence are much slower in the transformed and tumor cells than in the normal cells at 18 degrees C. Entry and spread of fluorescence in normal cells become slow enough to be observed only at 12 degrees C or below.

Cell Line↗
PubMed1978

Differences in the buoyant density characteristics of murine granulocyte-macrophage progenitor cells cloned in the presence of serum and/or hemolysate.

Bone marrow-derived progenitor cells of granulocytes and macrophages undergo proliferation and maturation in vitro in the presence of colony-stimulating factors and suitable sera. It has recently been found that rat hemolysate can replace serum as a source of growth-supporting activities for cloning these progenitor cells. It is unclear, however, whether serum or hemolysate allows the growth of separate or identical precursor cell subpopulations. Granulocyte-macrophage progenitor cell populations cloned in either serum or hemolysate were identical in their buoyant density characteristics. However, an additional population of granulocyte-macrophage progenitors was preferentially detected when both substances were added to the cell cultures. Different responding cells may, therefore, differ in their nutritional requirements for in vitro cloning. Variability in the respective amounts of these growth factors in the stimulator source would influence the number of CFU-C cloned. The actual amount of stimulator in unpurified sources may, therefore, be grossly overestimated.

Animals↗
PubMed1977

Inhibition of cytokinesis in Friend leukemia cells by membrane mobility agents.

Treatment of a line of Friend leukemia cells with a dispersion of the membrane mobility agent, A2C, yields cells that undergo successive nuclear divisions without cytokinesis, resulting eventually in cells with as many as 30 nuclei. Neither the DNA replication rate of the cells nor the generation time is different after treatment; in addition, the multiple nuclei divide synchronously, and the chromosome number corresponds to the number of nuclei in the cell. Inhibition of cytokinesis is not observed if the cells are washed with reagent-free medium within 1 hr of treatment, but is observed if washing is delayed for 24 hr. Membrane mobility agent loaded with the fluorescent probe, Flomol F20C, leads to fluorescent membrane; fluorescence disappears from the membrane after a change of medium within 1 hr, but not after a change of medium within 24 hr. Some stages in the overall development resemble those seen for cytochalasin B inhibition of cytokinesis, although the mechanisms may well be different for the inhibition promoted by membrane mobility agent. The inhibition of cytokinesis by A2C provides a potentially interesting means of studying cytokinesis and the regulation of differentiation.

Animals↗
PubMed1977

Correlation between movement of concanavalin A membrane receptors and cytolysis. A scanning electron microscopy study.

The present study was undertaken to test whether cytolysis induced by Concanavalin A (Con A) requires lateral mobility of membranal lectin receptor sites into caps. Treatment of interphase murine mastocytoma cells with 10(-4) M colchicine promoted cap formation by Con A in about 30% of the cells, followed by cytolysis. Pretreatment of the cells with NaN3, low temperature, or glutaraldehyde decreased the degree of capping and, to the same extent, the degree of cytolysis. The addition of antibodies to cells bound with Con A increased the appearance of capping and cytolysis. A linear relationship with a high correlation coefficient exists between the degree of capping and cytolysis, suggesting that lateral mobility of membrane Con A receptors is required for cytolysis by the lectin. The process of cap formation by Con A up to the stage of cytolysis was followed by scanning electron microscopy.

Animals↗
PubMed1977

Regulation of lipopolysaccharide-induced granulopoiesis and macrophage formation by spleen cells. I. Relationship between colony-stimulating factor release and lymphocyte activation in vitro.

Addition of bacterial lipopolysaccharide (LPS), a B cell mitogen, to mouse spleen cultures strongly stimulated production of colony-stimulating factor (CSF), the humoral regulator of granulopoiesis, and macrophage formation in vitro. Secretion of CSF from LPS-stimulated spleen cells coincided with cellualr DNA synthesis and cell transformation and both activities could be attributed to the lipid A moiety of the molecule. Different experimental approaches were used to study the relationship of CSF release and lymphocyte activation in response to LPS: a) modification of LPS with polymyxin B, an antibiotic bactericidal for most Gram-negative bacteria, caused a marked reduction in mitogenic activity, although the ability to induce CSF was not significantly altered; b)spleen cells from CBA/N mice, a mutant strain with an x-linked genetic defect in immunologic and mitogenic responses to polyclonal activators including LPS, showed diminished mitogeinc responses; however, high levels of CSF were produced; c) mitotic and DNA inhibitors (colchicine and cytosine arabinoside) did not affect CSF release although they completely inhibited mitogenicity. Thus, the spleen cell population participating in the process of LPS-induced CSF generation is probably a nondividing, terminally differentiated one without need for DNA synthesis. In addition, it was also shown that active RNA and protein synthesis are needed in this process.

Animals↗
PubMed1977

Replacement of serum by hemolysate as growth promoter for murine leukemic and normal hemopoietic progenitor cells in culture.

The present study was undertaken to test whether lysates prepared from rat erythrocytes can replace serum as in vitro growth promoters for murine leukemic and normal hemopoietic progenitor cells (CFU-C). Normal bone marrow and three leukemic cell lines (P-1081, L-1210 and P-815) were used in all the experiments. The soft agar technique for cloning hemopoietic cells was used to quantitate cell proliferation. Addition of hemolysate to the agar medium at a final concentration of 4% promoted the growth of a maximal number of leukemic colonies, similar to the number of colonies obtained when 20% serum was added to the soft agar. As for normal CFU-C, addition of 10% hemolysate or 40% horse serum promoted the growth of comparable numbers of colonies. Rat hemolysate can therefore replace horse serum as a growth promoter of leukemic and normal CFU-C present in culture. It was also shown that rat hemolysate cannot substitute for the colony stimulating factor (CSF) needed for the cloning of CFU-C. However, addition of hemolysate, CSF and serum to the soft agar cultures promoted a potentiated rather than an additive growth of CFU-C.

Animals↗
PubMed1977

Genetic analysis of generation of serum interferon by bacterial lipopolysaccharide.

Administration of bacterial lipopolysaccharides (LPS) (LPS) to mice causes interferon (IF) to appear in their blood stream. The generation of this serum IF is "early" and its maximum concentration appears 2 hr after an inoculation of a single optimal dose of LPS (100 microgram). Two inbred strains of mice differing in their ability to generate LPS-induced serum IF were used: a high responder strain (C3H/eB), which generates high levels of IF (about 100 IF units/ml serum), and a low responder strain (C3H/HeJ), which fails to generate any detectable amounts of IF in the serum. A genetic analysis was carried out and circulating IF production was determined in the F1 hybrid, in the F2 generation, and in the progeny of backcrosses of F1 hybrids to either parent strain. The results indicate that a single dominant autosomal gene is responsible for the difference in circulating IF production between both parent strains. In addition it was shown that this type of IF is stable to treatment at pH 2 for periods of time as long as 48 hr, but is labile to heating at 56 degrees C for 30 min.

Animals↗
PubMed1976

Lipid A, the active part of bacterial endotoxins in inducing serum colony stimulating activity and proliferation of splenic granulocyte/macrophage progenitor cells.

An analysis of which component of lipopolysaccharides (LPS), the lipid or the polysaccharide (PS), is active in stimulating the murine granulopoietic system has been performed. LPS with different structures, isolated from different mutant strains of Salmonella and chemical degradation products of lipopolysaccharides have been used. Lipid A obtained by acid hydrolysys of the LPS and complexed to bovine serum albumin (BSA) (lipid A-BSA) was shown to be active in generating serum colony stimulating factor (CSF) and in increasing the splenic colony forming cells (CFC) levels, although it was less active than the parent LPS. The polysaccharide (PS) showed no significant activity at the concentrations used. LPS (glycolipids) from R mutants of Salmonella minnesota were active to the same extent as the LPS. The fact that even the most defective LPS from the R mutant R595 which contains lipid A and KDO only is a potent endotoxin, points unequivocally, to lipid A, as the active principle in stimulating the granulopoietic system.

Animals↗
PubMed1976

Genetic control of lipopolysaccharide induced generation of serum colony stimulating factor and proliferation of splenic granulocyte/macrophage precursor cells.

Administration of bacterial lipopolysaccharides (LPS) to mice causes a rise in tissue and serum colony stimulating factor (CSF) levels and in bone marrow and splenic colony forming cells (CFC). Two inbred strains of mice differing in their response to LPS were used to study the genetic control of LPS induced granulopoietic responses: a high responder strain (C3H/eB) which reacts to LPS by an elevation in serum CSF and by an increase in splenic CFC levels, and a low responder strain (C3H/HeJ) which fails to show these responses. The ability to generate serum CSF after administration of LPS is controlled by a single autosomal dominant gene, while the splenic CFC response to LPS follows the characteristic patterns of a polygenic inheritance control. The associated relationships of CSF and CFC responsiveness have been investigated in backcross (F1 X C3H/Hej) and F2 mice. Most mice which generated high levels of CSF showed a high or intermediate CFC response and most mice which did not generate any detectable levels of serum CSF showed a low splenic CFC response. The results suggest that CSF may play a physiologic role in vivo as a granulopoietin. In addition it was shown that the genetic control mechanisms governing the CSF/CFC responses are determined by the lipid A-KDO portion of the LPS molecule, suggesting that lipid A is the active part of the LPS molecule in stimulating granulopoiesis.

Animals↗