The adaptation to postnatal life involves liver growth and functional differentiation. The underlying hypothesis for this subproject is that normal and perturbed hepatocyte differentiation are co-regulated with hepatocyte proliferation. Previous work on this project has shown that fetal rat hepatocytes have a proliferative phenotype which is independent of signaling by paracrine or autocrine mitogens. Furthermore, this """"""""mitogen-independent"""""""" growth shows a complex ontogenic pattern between the 17th day of gestation and the end of the first postnatal week. Most notably, there is a temporary cessation of proliferation for approximately 24 hr surrounding parturition. Beyond the fourth postnatal day, hepatocytes gradually attain their quiescent, adult state. The present proposal focuses on the cell cycle mechanisms which are responsible for this pattern. A culture system in which fetal rat hepatocytes are induced to quiesce in vitro over 96 hr has been developed. Thus, we propose to study regulation of hepatocyte proliferation in culture at the level of cell cycle-dependent protein kinases (CDKs), CDK positive regulators (Cyclins) and CDK inhibitors (CKIs). These results will be complemented by studies on CDK, Cyclin and CKI expression and regulation in vivo. A primary goal of the proposed studies is to determine the cell-to-cell relationship between negative cell cycle regulation by CKIs and hepatocyte differentiation. The latter will be assessed using immunocyto/histochemistry for two enzymes required for hepatic glucose production, glucose-6-phosphatase and PEPCK, as well as other hepatocyte- specific markers. Our observations will be extended to an animal model of impaired fetal growth, maternal starvation for 48 hr prior to delivery at term in the rat. In particular, the role of insulin in regulating these processes in cultured hepatocytes from IUGR fetuses will be investigated in order to elucidate the hepatic pathophysiology of intrauterine growth retardation.
