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Biomedical subjects

H Baumann

Publications and source records attributed to H Baumann.

At least 235 records · Page 13Linked to original sources
PubMed1983

UV micro-irradiation of the Chinese hamster cell nucleus and caffeine post-treatment. Immunocytochemical localization of DNA photolesions in cells with partial and generalized chromosome shattering.

UV micro-irradiation of a small part of the Chinese hamster nucleus and caffeine post-incubation often results in shattered chromosomes at the first post-irradiation mitosis. In some of these mitotic cells, chromosome shattering is restricted to a few chromosomes spatially related in a small area of the metaphase spread; in others, shattering includes the whole chromosome complement. These 2 types of damage have been called partial and generalized chromosome shattering (PCS and GCS). Using antisera that specifically react with UV-irradiated DNA, we identified micro-irradiated chromatin in interphase nuclei and in mitotic cells with PCS or GCS by indirect immunofluorescence microscopy. In PCS, immunofluorescence staining was found in the damaged area, while the surrounding intact chromosomes were not stained. In GCS, staining was also restricted to a small region of the shattered chromosome complement. In other experiments, cells synchronized in G1 were micro-irradiated in the nucleus, pulse-labelled with [3H]thymidine and post-incubated with caffeine. Autoradiographs of cells with GCS showed unscheduled DNA synthesis restricted to a small chromatin region. Our data present direct evidence that the distribution of DNA photolesions does not coincide with the sites of chromosomal damage in GCS. As a working, hypothesis, we propose that an indirect mechanism is involved in the induction of GCS by which DNA photolesions in a small nuclear segment induce shattering of both micro-irradiated and non-irradiated chromosomes.

Animals↗
PubMed1983

Preferential degradation of the terminal carbohydrate moiety of membrane glycoproteins in rat hepatoma cells and after transfer to the membranes of mouse fibroblasts.

Glycoproteins in the plasma membrane of rat hepatoma cells were labeled at their externally exposed tyrosine residues with 131I and at their galactose and sialic acid residues with 3H. The degradation of both isotopes in the total cell protein fraction, in glycoproteins purified by concanavalin A, and in glycoproteins separated on two-dimensional gels was determined. Similarly, the total cellular membrane glycoproteins were metabolically labeled with [35S]methionine and [3H]fucose. The fate of both incorporated labels was followed by lectin chromatography or by precipitation of the proteins with specific antibodies followed by electrophoretic gel separation. In both labeling experiments, the carbohydrate markers were lost from the ligand-recognized fraction with similar kinetics as from the total cell protein fraction. In some glycoprotein species which were separated by two-dimensional gel electrophoresis, the polypeptide portion exhibited up to a twofold slower rate of degradation relative to that of the carbohydrate moiety. This difference is most pronounced in carbohydrate-rich glycoproteins. To corroborate this finding, double-labeled membrane glycoproteins were incorporated into reconstituted phospholipid vesicles which were then transferred via fusion into the plasma membrane of mouse fibroblasts. Both the polypeptide and carbohydrate moieties of the transferred membrane glycoproteins were degraded with the same relative kinetics as in the original hepatoma cells. The rate of degradation is mostly a function of the structural properties of the membrane components as shown by the preservation of metabolically stable fucogangliosides of Reuber H-35 hepatoma cells transferred onto the fibroblasts. The technique of insertion of membrane components into the plasma membrane of another cell should assist in the elucidation of the exact route and mechanism of membrane protein destruction.

Animals↗
PubMed1983

Regulation of mouse haptoglobin synthesis.

A cloned line of mouse hepatoma cells (Hepa-1) responded to treatment with dexamethasone by a 30-80-fold increase in synthesis and secretion of functional haptoglobin. Under the same conditions, the production of albumin was only slightly elevated whereas that of alpha 1-fetoprotein was reduced by 50%. The hormone concentration for half-maximal stimulation of haptoglobin synthesis was between 1 and 2 X 10(-8) M. The time course of induction is characteristic for a glucocorticoid-regulated protein. Cell-free translation of RNA indicated an increase in the amount of functional haptoglobin mRNA that can account for the change in the protein production. To correlate our findings on Hepa-1 cells with those on nontransformed liver cells, we tested the hormonal response of isolated hepatocytes in tissue culture. Haptoglobin was first synthesized and secreted by hepatocytes from 17-19-d-old fetuses. But neither prenatal nor adult hepatocytes showed a dexamethasone-dependent increase in haptoglobin synthesis. However, when several independent clones of hybrid cells formed from adult mouse hepatocytes and rat hepatoma cells were treated with dexamethasone, the synthesis of mouse haptoglobin was in all cases elevated. It appears that haptoglobin expression in mouse liver cells is potentially sensitive to glucocorticoids, but this modulation is manifested only in transformed cells and their derivatives.

Animals↗
PubMed1983

Synthesis and regulation of acute phase plasma proteins in primary cultures of mouse hepatocytes.

Adult mouse hepatocytes respond in vivo to experimentally induced acute inflammation by an increased synthesis and secretion of alpha 1-acid glycoprotein, haptoglobin, hemopexin, and serum amyloid A. Concurrently, the production of albumin and apolipoprotein A-1 is reduced. To define possible mediators of this response and to study their action in tissue culture, we established primary cultures of hepatocytes. Various hormones and factors that have been proposed to regulate the hepatic acute phase reaction were tested for their ability to modulate the expression of plasma proteins in these cells. Acute phase plasma and conditioned medium from activated monocytes influenced the production of most acute phase plasma proteins, and the regulation appears to occur at the level of functional mRNA. Purified hormones produced a significant anabolic response in only a few cases: dexamethasone was found to be effective in maintaining differentiated expression of the cells; and glucagon produced a specific inhibition of haptoglobin synthesis. When cells were treated with a combination of conditioned monocyte medium and dexamethasone, secretion of proteins was markedly reduced. The carbohydrate moieties of all plasma glycoproteins were incompletely modified, apparently as a result of decreased intracellular transport of newly synthesized plasma proteins. Although primary hepatocytes were not phenotypically stable in tissue culture, the cells nevertheless retained a broad response spectrum to exogenous signals. We propose this as a useful system to study the production of plasma proteins and thereby pinpoint the nature and activity of effectors mediating the hepatic acute phase reaction.

Albumins↗
PubMed1983

Synthesis of oligosaccharides that form parts of the complex type of carbohydrate moieties of glycoproteins. Three glycosides prepared for conjugation to proteins.

Condensation of 3,6-di-O-acetyl-4-O-(2,3,4,6-tetra-O-acetyl-beta-D-galactopyranosyl)-2-deoxy-2- phthalimido-beta-D-glucopyranosyl bromide with p-nitrophenyl 3-O-benzoyl-4,6-di-O-benzylidene-alpha-D-mannopyranoside, p-nitrophenyl 3,6-di-O-benzyl-alpha-D-mannopyranoside and p-nitrophenyl 3,4-di-O-benzyl-alpha-D-mannopyranoside gave protected tri- and pentasaccharides. The oligosaccharide glycosides 1, 2 and 3 were obtained after de-protection of these condensation products. These oligosaccharides will, after suitable conversions, be conjugated to proteins for biological studies.

Glycoproteins↗
PubMed1982

Dexamethasone increases the synthesis and secretion of a partially active fibronectin in rat hepatoma cells.

After treatment with dexamethasone, rat hepatoma-tissue culture cells show a markedly enhanced adhesion to the substratum and increased cell-to-cell interaction. In addition, there is a profound change in the production of secretory glycoproteins. Although the relative synthesis and secretion of a gelatin-binding, fibronectinlike glycoprotein is increased threefold, we do not think this protein is responsible for the improved adhesion properties of the cells because the hepatoma cells do not bind normal fibronectin and because the HTC-produced fibronectin is neither bound by fibroblasts nor has it any affinity for ganglioside-containing phospholipid vesicles. Therefore, these hepatoma cells represent a unique system for studying the regulation of fibronectin synthesis by glucocorticoids. Furthermore, analyses of primary fetal rat hepatocytes have shown that these cells, unlike normal adult hepatocytes, synthesize and secrete fibronectin, which is structurally related to the HTC-cell protein. The comparison of this protein with fibronectin from normal cells will allow a structural characterization of the functional defect in the fibronectin synthesized by transformed cells.

Animals↗
PubMed1982

Influence of the liver on the profile of circulating antigens recognized by antiserum against hepatoma membrane glycoproteins.

Rat hepatoma tissue culture cells have retained only a few liver-specific membrane functions. To find out whether transformation of these cells led to the appearance of tumor-specific membrane glycoprotein structures, which might also be present in released form in the circulation of tumor-bearing animals, we compared immunologically the sera and ascites fluids of normal and of hepatoma-bearing rats. For structural identification, we used polyspecific antibodies against the membrane glycoproteins of the hepatoma cells. Crossed immunoelectrophoresis of the body fluids resulted in the separation of several antigens, one of which appears to be tumor specific. Immunoelectrophoretic analyses of the glycoproteins from the membranes of liver and hepatoma cells revealed many shared antigens; in one instance, we could assign the membrane associated form to major antigen detected in the serum. Characterization of the molecular size and charge of the two selected antigens by two-dimensional gel electrophoresis indicated that the hepatoma cells products were not completely sialylated. In the case of the circulating, hepatoma-specific antigen, this property appears to result in hepatic clearance of the more basic ("asialo") portion of the glycoprotein species via the receptor-mediated uptake of galactose-terminated glycoproteins. The result was the accumulation of the acidic forms of the antigen in the serum and ascites fluid of tumor-bearing rats.

Animals↗
PubMed1981

Biosynthesis and hormone-regulated expression of secretory glycoproteins in rat liver and hepatoma cells. Effect of glucocorticoids and inflammation.

Exposure of rat hepatoma tissue culture cells to dexamethasone results in appearance of a new glycoprotein, gp35-50 (Mr = 35,000 to 50,000) and increased synthesis of another glycoprotein, gp50 (Mr = 50,000). The glycoproteins synthesized in a fractionated cell-free translation system (containing dog pancreas microsomes) appear as a well separated series of spots on two-dimensional polyacrylamide gels which differ in size and charge. Glycoproteins synthesized by glucose-starved cells show similar size and charge heterogeneity. The size heterogeneity consists of a series of spots, each differing in molecular weights by about 3,000, which could be almost completely abolished by treatment with endo-beta-N-acetylglucosaminidase H. Our results indicate that unglycosylated gp50 (Mr = 42,000) typically acquires 3 N-glycan units, whereas gp35-50 (Mr = 22,000) possesses eight N-glycosylation sites. Analysis of the cell-free translation products directed by mRNA from hepatoma tissue culture cells grown in tissue culture, from hepatoma tissue, and from normal liver tissue indicated that administration of dexamethasone causes a pronounced increase in gp35-50 mRNA in all three tissues. A similar increase was observed in liver after inflammation which along with other biochemical properties suggests that gp35-50 may be alpha 1-acid protein. In contrast, mRNA coding for gp50 was not increased by dexamethasone in tumor tissue and no protein structurally related to gp50 was detected in the liver mRNA translation products. Thus, gp35-50 is expressed in normal liver, whereas gp50 is expressed in hepatoma cells and is differentially regulated by steroid hormones depending on whether the cells are grown in tissue culture or as a tumor in the rat.

Animals↗