Increased production of collagen in Vivo by hepatocytes and nonparenchymal cells in rats with carbon tetrachloride-induced hepatic fibrosis

Abstract

We have shown, using the proline:ornithine dual label method, that in normal rats, hepatocytes contribute in vivo about 80 to 90% of the newly synthesized hepatic collagen. In order to quantify the contribution of hepatocytes and nonparenchymal cells to collagen synthesis in vivo in hepatic fibrogenesis, rats with CCl4-induced liver fibrosis were given [5(3H)]proline and [14C]ornithine intraperitoneally. About 80% of the 14C in albumin and transferrin was present as arginine, following conversion of [14C]ornithine via the urea cycle. In contrast to hepatocyte proteins, in nonparenchymal cells and serum a negligible percentage of the radioactivity was present as [14C]arginine. These combined findings indicate that, in spite of the hepatocellular damage, the labeling of hepatocyte proteins was efficient and specific, validating the use of the proline:ornithine method in this experimental model of hepatic fibrosis. We calculated the [3H]proline/[14C]arginine ratio in hepatic collagen (after correcting for the relative frequencies of amino acids) as a percentage of the same ratio in either albumin or transferrin, the index hepatocyte proteins. In this experimental model, during active fibrogenesis, both hepatocytes and nonparenchymal cells increase their production of collagen 2-fold when compared to normal animals, and hepatocytes produce the majority of the newly synthesized hepatic collagen.

Expression of the ECM proteins fibulin-1 and -2 in acute and chronic liver disease and in cultured rat liver cells

“…As shown by the analysis of in vitro cultures, the fibulin-1 gene is expressed in a greater number of cell types than is fibulin-2, including both parenchymal and non-parenchymal cells. The synthesis of ECM proteins in hepatocytes has been previously reported (Chojkier et al 1988) and raises the question of the role of parenchymal cells in the process of fibrogenesis (Dooley et al 2008). Nevertheless, we should also take into consideration that primary cultures of isolated hepatocytes might be overgrown by mesenchymal cells, and the contaminating cell populations might express ECM proteins.…”

Stimulation and proliferation of primary rat hepatic stellate cells by cytochrome P450 2E1-derived reactive oxygen species

“…33 The interaction of cells with ECM regulates cell morphology, motility, proliferation, survival, differentiation, or gene expression via integrin-mediated signal transduction. 26,[34][35][36] The activation process in HSC is classically recognized by a significant modulation in their cytoskeletal filament phenotype, primarily the increased expression of ␣-sma filaments. 37 There was a time-dependent increase in the production of ␣-sma, with maximal differences in the E47 versus the C34 coculture after 3 days.…”

“…End-products of lipid peroxidation can stimulate collagen production by stellate cells. 26,27 Oxidative stress and lipid peroxidation are key features in alcoholinduced liver injury, 28,29 and CYP2E1 is a potent catalyst of lipid peroxidation. We determined the amount of lipid peroxidation end-products present in HSC alone and in both cocultures, and found an increase of about 80% in HSC coincubated with the E47 cells when compared with the other two cocultures (P Ͻ .005) (Fig.…”

Extracellular matrix gene expression increases preferentially in rat lipocytes and sinusoidal endothelial cells during hepatic fibrosis in vivo.

“…These observations suggest that parenchymal liver cells play a central role in the production of interstitial collagen that accompanies hepatic fibrosis. In vivo labeling studies also suggest that the majority of collagen extracted from liver after carbon tetrachloride administration derives from hepatocytes (10). Other experiments, however, have led to a different conclusion, that hepatic extracellular matrix synthesis is largely a function of mesenchymal liver cells.…”

“…Lipocytes, which remained at the top two interfaces, were collected and washed twice in culture medium followed by centrifugation at 500 g to remove hepatocyte debris. Endothelial cells, collected from the bottom two interfaces of the gradient, were purified further by centrifugal elutriation at 2,500 rpm and a flow rate of [10][11][12][13][14][15][16][17][18] Probe preparation. cDNAs encoding rat al(I) collagen (paIRI) (24), rat al(III) collagen (PRGR5) (Makela, J. K., and E. Vuorio, unpublished observations), mouse al(IV) collagen (pPE123) (25), and mouse laminin B2 chain (pPE9) (26) (27), which cross-hybridizes with rat fl-actin, was provided in SP64 by Dr. J. M. Friedman, The Rockefeller University, New York.…”