Biomedical subjects

M Odenthal

Publications and source records attributed to M Odenthal.

25 records · Page 2Linked to original sources

PubMed1993

Localization and mRNA steady-state level of cellular fibronectin in rat liver undergoing a CCl4-induced acute damage or fibrosis.

In an attempt to investigate cellular fibronectin synthesis and deposition during acute liver damage and fibrogenesis, we used the presence of the additional type III-related ED-A domain of cellular fibronectin as a characteristic for distinguishing it from the plasma form. Using site-specific antibodies, we localized cellular fibronectin deposition in the necrotic pericentral areas of acutely damaged liver tissue after a single CCl4-gavage, whereas in control liver only trace amounts of cellular fibronectin were detectable along the sinusoids. Upon several CCl4-administrations leading to liver fibrosis, cellular fibronectin deposits were accumulated in the fibrotic septa. Northern blot hybridization using a cDNA representing part of the ED-A domain revealed that in liver tissue, in response to an acute intoxication, cellular fibronectin synthesis was initiated within the first 48 h after CCl4-gavage. By in situ hybridisation transcripts for cellular fibronectin were identified in the necrotic areas of acutely damaged tissue restricted to single, pericentrally located cells, whereas no cellular fibronectin mRNA was detectable in control liver. During fibrogenesis cellular fibronectin transcripts were shown to be synthesized in the immediate vicinity of septa. We conclude that upon acute or chronic intoxication, cellular fibronectin is a member of the accumulating biomatrix and is produced locally by mesenchymal liver cells.

Animals↗

PubMed1992

Alternative splicing products of the tenascin gene distinguish rat liver fat storing cells from arterial smooth muscle cells and skin fibroblasts.

Fat storing-(Ito-)cells (FSC) transform into a myofibroblast-like cell type during liver fibrogenesis. A similar development can be observed in cell culture. At the moment, a definite marker to differentiate transformed FSC from smooth muscle cells (SMC) is not available. We recently found that FSC, SMC and skin fibroblasts (SF) synthesize tenascin, a novel matrix protein. As it is reported that various tissues express different tenascin forms by the mechanism of alternative pre-mRNA splicing, we analyzed the tenascin transcripts in these cell types. Total RNA extracted from cultured FSC, SMC and SF, analyzed by Northern blot hybridization, showed a 7.2 kb transcript in FSC, a 8.7 kb mRNA in SMC, whereas SF produced both messages. As the splicing pattern of FSC in primary culture did not change after passaging, this differential expression of tenascin mRNA might provide a tool to identify myofibroblast-like cells derived from FSC. The important fibrogenic mediator transforming growth factor-beta (TGF-beta) increased tenascin gene expression in each cell type. In SMC, TGF-beta additionally induced the production of the 7.2 kb transcript. Determination of tenascin transcripts will allow to examine the purity of FSC cultures and facilitate a better identification of the cells involved in liver fibrosis.

Adipose Tissue↗

PubMed1992

The gene of hepatocyte growth factor is expressed in fat-storing cells of rat liver and is downregulated during cell growth and by transforming growth factor-beta.

Hepatocyte growth factor (HGF) has been detected in non-parenchymal cells but not in hepatocytes. We performed Northern blot analysis of total RNA extracted from rat hepatocytes, Kupffer cells, endothelial cells and fat-storing (Ito-) cells. Total RNA was extracted from fat-storing cells at different times after isolation and from cells treated with different amounts of transforming growth factor beta. The RNA was hybridized with HGF, fibronectin-, and alpha-actin-specific cDNA probes, consecutively. We found an abundant amount of HGF mRNA in freshly isolated fat-storing cells, but not in other liver cells. The amount of the HGF transcripts decreases significantly in FSC during the time of culture, while fibronectin gene expression increases and alpha-actin gene expression as well. TGF-beta dramatically inhibits HGF gene expression, but causes an enhanced fibronectin mRNA level. Northern blot hybridisation of total RNA from CCl4-chronically damaged liver with HGF cDNA shows a significant increase of HGF mRNA during development of liver fibrosis. We suggest that in damaged liver either non-parenchymal cells, others than FSC, became able to express the HGF in vivo, or other mediators overcome the inhibitory effect of TGF-beta.

Actins↗

PubMed1992

Fat-storing cells of the rat liver synthesize and secrete C1-esterase inhibitor; modulation by cytokines.

During liver fibrogenesis, fat-storing cells transform into myofibroblast-like cells and produce increasing amounts of extracellular matrix proteins. Because fat-storing cells produce alpha 2-macroglobulin, an important serine protease inhibitor (serpin), we investigated whether fat-storing cells also synthesize C1-esterase inhibitor, another important serpin. C1-esterase inhibitor synthesis was studied in rat fat-storing cells at day 0, 3 and 7 after isolation by biosynthetic labeling, immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Messenger RNA was examined by Northern-blot analysis. C1-esterase inhibitor gene expression and synthesis were detectable in freshly isolated fat-storing cells and increased distinctly during the time in culture. The cellular source of C1-esterase inhibitor in fat-storing cell cultures was also identified by in situ hybridization of cells at different times after isolation. By inhibition of the N-glycosylation using tunicamycin, rat C1-esterase inhibitor was identified as a glycoprotein. The time course of C1-esterase inhibitor secretion was determined by pulse-chase experiments. C1-esterase inhibitor synthesis was increased 6-fold to 10-fold by interferon-gamma. Specific messenger RNA levels were also raised distinctly by this cytokine. In contrast, interferon-alpha and dexamethasone did not alter C1-esterase inhibitor gene expression. Because C1-esterase inhibitor synthesis is increased by advancing culture time and by the inflammatory mediator interferon-gamma, we suggest that fat-storing cells may enhance the deposition of extracellular matrix proteins by inhibiting their degradation.

Animals↗

PubMed1992

Rat hepatocytes in primary culture synthesize and secrete cellular fibronectin.

Fibronectins, involved in cell-matrix interactions and cell attachment, are glycoproteins which show a remarkable heterogeneity, due to alternative splicing. The type III-related domains, ED-A and ED-B, are present in cellular fibronectin in a variety of ratios whereas they are absent in circulating plasma fibronectin. Fibronectin synthesis by hepatocytes which are accepted as suppliers of plasma fibronectin was studied in primary cultures during a 6-day culture period. Using site-specific antibodies we demonstrate that rat hepatocytes are also able to synthesize and secrete fibronectin bearing the ED-A domain from Day 3 on after inoculation. By immunocytological characterization of the hepatocyte monolayer with antibodies directed against desmin, laminin, collagen IV, alpha-SM-actin, or ED-1 or factor VIII-related antigen, contaminating mesenchymal hepatic cell-types as a source for cellular fibronectin production could be ruled out. Dexamethasone treatment caused enhanced fibronectin synthesis and cellular fibronectin was already detectable at Day 1 after plating. Elevation of cellular fibronectin synthesis after prolonged culture-terms and by dexamethasone could also be demonstrated on mRNA steady-state level, using ED-A cDNA as a probe in hybridization analysis. Dot blot hybridisation proved a prominent response of cellular fibronectin mRNA level to dexamethasone at Day 1 when dexamethasone treatment resulted in an increased contribution of ED-A-positive fibronectin transcripts to total fibronectin mRNA level.

Animals↗

PubMed1992

Synthesis of undulin by rat liver fat-storing cells: comparison with fibronectin and tenascin.

Fat-storing cells (FSCs) are known to synthesize various components of the hepatic extracellular matrix and thereby play an important role during liver fibrogenesis. The aim of our study was to investigate the synthesis of undulin, a recently described connective tissue protein belonging to the fibronectin-tenascin superfamily of glycoproteins, by fat-storing cells in primary culture. SDS-PAGE analysis of immunoprecipitates from cell layer lysates or media pulse-labeled with radioactive methionine revealed undulin-specific bands A (270 kDa), B1 (190 kDa), and B2 (180 kDa) after reduction. A single undulin-specific transcript was detected at about 7 kb. Undulin synthesized by cell-free translation revealed two polypeptides migrating about 5000 Da below the B1 and B2 subunits. Treatment of FSCs with tunicamycin created two novel bands slightly below the B2 chain. Since the electrophoretic patterns of undulin chains recovered by cell-free translation and tunicamycin treatment of cells were very similar we suggest that N-glycosylation is the major post-translational processing event. Newly synthesized undulin was detected after 30 min of pulse labeling in the cell layer fraction and was secreted into the medium at a slower rate than fibronectin. In contrast to fibronectin and tenascin, undulin was already synthesized by freshly isolated FSCs and during the early stage of primary FSC culture ("resting" cells), supporting the hypothesis that undulin is associated with a differentiated mesenchyma. However, in analogy to fibronectin and tenascin, undulin was also synthesized by "activated" FSCs, indicating that undulin might also be of importance in dedifferentiated tissues.

Animals↗

PubMed1992

Synthesis of cellular fibronectin by rat liver fat-storing (Ito) cells: regulation by cytokines.

Fibronectins are multifunctional extracellular matrix glycoproteins that seem to play a pacemaker role in liver fibrogenesis. The expression of cellular fibronectin by rat liver fat-storing cells in primary culture and their modulation by cytokines was studied. Cellular fibronectin was localized in the cytoplasm and on the surface of cultured fat-storing cells as well as in extracellular matrix fibrils by indirect immunofluorescence. Immunoprecipitation of endogenously labeled fibronectin using type specific antibodies showed the synthesis and secretion of cellular fibronectin by fat-storing cells in vitro. ED1 positive sequences specific for cellular fibronectin and absent in plasma fibronectin were detected within the total RNA of fat-storing cells. Cellular fibronectin synthesis was severalfold enhanced in "activated" vs. "resting" fat-storing cells. Treatment of fat-storing cells with transforming growth factor beta 1 resulted in a dose dependent increase of fibronectin synthesis. In contrast, interferon gamma inhibited the synthesis of fibronectin. The stimulation of fibronectin synthesis by transforming growth factor beta 1 and the inhibition by interferon gamma might be of importance for pathophysiology and therapy of liver fibrogenesis.

Animals↗