The general goal of this proposal is to determine how specific growth factors and cell cycle associated genes regulate normal and neoplastic liver growth. We have shown that during liver regeneration after partial hepatectomy (PH) in rats there is a sequential and transient expression of """"""""competence"""""""" protooncogenes followed a few hours later by the activation of a positive autocrine growth control circuit involving TGF-alpha, and a negative paracrine circuit mediated by TGF-beta1. We now ask whether protooncogene activation and TGF-alpha expression are obligatory events in hepatocyte DNA synthesis and whether protooncogene activation makes hepatocytes capable of responding to TGF-alpha and other growth factors. To answer these questions we have devised methods to block TGF-alpha and c- myc expression in vivo and in vitro by antisense genes and oligodeoxynucleotides. Using a receptor mediated carrier system we will deliver TGF-alpha antisense genes to the liver in vivo and study the effects of these constructs on hepatocyte DNA synthesis complement the studies of TGF-alpha gene blockage, we will analyze the effects of TGF- alpha overexpression on hepatocyte replication in vivo using transgenic mice that express large amounts of TGF-alpha mRNA in the liver. As more than 50% of these animals develop spontaneous hepatocellular carcinomas at 12-14 months of age, the transgenic mouse model will be used to explore the relationships between TGF-alpha overexpression and hepatocarcinogenesis. Control of normal cell proliferation and malignant transformation involve, however, the interplay between activating and suppressor genes. TGF-beta1 is a potent inhibitor of hepatocyte DNA synthesis but the effects, expression and localization in the liver of other TGF-betas need to be determine. Further, we will investigate whether changes in expression of the retinoblastoma gene (Rb), a tumor suppressor gene, are associated with liver regeneration and if TGF-beta1 inhibition of hepatocyte DNA synthesis interferes with cell cycle associated phosphorylation of the Rb protein.
Specific aims are: a) investigate the role of TGF-alpha in hepatocyte proliferation using antisense constructs to block TGF-alpha in vivo and in vitro; b) determine the effect of TGF-alpha overexpression on liver regeneration and carcinogenesis; c) examine the mechanism of action and effects of TGF-betas 1-4 in the regenerating liver and their role in chronic human liver disease and d) determine if c-myc activation is an obligatory step in DNA synthesis and whether changes in Rb expression are associated with hepatocyte proliferation.
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