MECHANISMS OF IMPAIRED REGENERATION IN CIRRHOTIC LIVER

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This research proposal will examine the fundamental mechanisms responsible for the critical clinical problem of hepatic growth. The authors have had extensive experience in the development of several animal models which are ideal for the in depth study of hepatic regeneration, and propose to use these models in a rigorous and complementary fashion to attempt to study the mechanisms, regarding control of hepatic regeneration in both normal and cirrhotic liver. The hypotheses to be tested include: 1) Administration of so-called hepatotropic factors, such as insulin, glucagon, epidermal growth factor (EGF), and transforming growth factor alpha (TGF-alpha) stimulate liver regeneration of cirrhotic rats in vivo. 2) Significant alterations in peptide binding occur in plasma membranes from regenerating and cirrhotic liver, and are responsible, in part for the decreased proliferative response seen in cirrhosis; 3) Antiproliferative factors, such as somatostatin-14 (SS-14) and transforming growth factor beta (TGF-beta) inhibit hepatic proliferation by direct action on second messenger pathways in rat hepatocytes, which can be mimicked by specific pharmacologic probes. Specifically dideoxyadenosine (DDA) (an inhibitor of the catalytic subunit of adenylyl cyclase) and other pharmacologic reagents may mimic the action of SS-14, and alter intracellular accumulation of cyclic AMP; 4) Signaling mechanisms of GI peptide receptors for hepatic proliferation are linked to G proteins, and alterations of G protein structure and function occur during cirrhosis; 5) Altered patterns of early gene expression exist in cirrhosis which are responsible for the attenuated proliferative response; The specific aims of this proposal will be: 1) To attempt to increase hepatic regeneration in cirrhotic rats by the administration of exogenous growth factors. 2) To study the effect of hepatic regeneration and cirrhosis on the binding of EGF, TGF-alpha, insulin, glucagon, SS-14, and TGF-beta to liver plasma membranes; 3) To compare the antiproliferative effects of pharmacologic probes - which may mimic exogenous peptides - of the adenylyl cyclase system on hepatocytes from quiescent and hepatectomized normal and cirrhotic rat liver; 4) To determine the role of G-proteins in signal transduction pathways leading to hepatocyte proliferation, and to compare G proteins in normal liver to those found in cirrhotic liver; 5) To study the effects of hepatectomy and cirrhosis on the expression of immediate early response genes which are known to play a role in hepatic regeneration. The models utilized to accomplish the specific aims include 1) hepatotropic factor stimulation of the sham hepatectomized rat with analysis of plasma membrane binding, second messenger analysis, structural G-protein studies, early gene regulation, and analysis of hepatic DNA synthesis; 2) 70% hepatectomy and biochemical and molecular analysis as described above; 30 the Wistar cirrhotic rat model; 4) isolated cultured hepatocyte preparations; 5) highly purified subcellular fractions, including isolated liver plasma membrane preparations.