Increased connective tissue protein deposition in the liver is the hallmark of cirrhosis. However, the relative contribution of hepatocytes and nonparenchymal cells to hepatic connective tissue protein production in vivo is unknown. To answer this question, we devised a method to determine the relative contribution of hepatocytes to the hepatic production of proteins in vivo (J Clin Invest, in press). We injected rats i.p. with (3H) proline and (14/C) ornithine. (3H) Proline labeled prolyl-t-RNA in both hepatocytes and nonparenchymal cells. In contrast, (14/C) ornithine was rapidly converted to (14/C) arginine via the urea cycle only in hepatocytes, labeling arginyl-t-RNA. About 60% of the 14/C in albumin and transferrin was present as arginine while the remainder is found in proline and related aminoacids. As expected for proteins that have the same proline:arginine and that are produced solely by the hepatocyte, the (3/H) proline:(14/C) arginine was very similar in albumin and transferrin. Conversely, in non parenchymal cells a negligible percentage of 14/C was present as arginine; given the greater proline (+hydroxyproline):arginine in hepatic collagen, our data indicate that in normal rats, hepatocyte contribute most of newly synthesized hepatic collagen. Using the proline:ornithine dual-label method, the specific aims of this research proposal are the following: i) to assess the relative contribution of hepatocytes to hepatic connective tissue macromolecule production in vivo, in cirrhotic animals. ii) to ascertain whether the hepatocyte produces laminin and fibronectin in vivo in normal animals. iii) to characterize the above described parameters in vivo in animals undergoing hepatic regeneration. iv) to evaluate the same parameters in vivo in animals with bile duct ligation. v) to determine the hepatocyte contribution to macromolecule production in co-cultures of hepatocytes and nonparenchymal cells. vi) to assess the hepartocyte contribution to alpha 2 macroglobulin production in vivo in normal, acute-phase response, and cirrhotic animals.
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