CELLULAR PATHOPHYSIOLOGY OF HEPATIC FIBROSIS

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Fibrosis is the liver's response to diverse injuries. Representing parenchymal "scar", it consists of collagens and other large extracellular matrix (ECM) proteins. The effects of fibrosis are both regional and local, involving disruption of lobular structure and also cellular function. Fibrosis in its advanced form, cirrhosis, is the most important cause of morbidity and mortality from liver disease. When this project began, in 1982, its goal was defining the cellular source(s) of fibrosis, as an essential first step towards understanding its regulation. This was accomplished through development of methods for mass isolation and primary culture of the individual cell populations of the liver. The lipocyte (Ito or fat-storing cell) emerged as the principal fibrogenic cell type. It was found, moreover, to undergo activation, defined as a pleiotypic response to inflammation. In this response, lipocytes convert from a normally quiescent, retinoid-storing state to one in which ECM production is sharply upregulated and features of smooth-muscle cells appear, including contraction. During the recent period of funding, our focus has been the regulation of lipocyte activation. While soluble mediators of inflammation (cytokines) are in part responsible, the role of the BCM itself is, if anything, more important than that of individual cytokines. In pursuing the lipocyte-activating effect of ECM, we have investigated fibronectin in detail, finding that a splice variant, one containing the EIIIA segment, plays a critical role. Production of this form increases sharply within 12 hours of an injury and is localized specifically to sinusoidal endothelial cells. In culture the EIIIA-containing form (A+-fibronectin) has direct lipocyte-activating effects, which are completely neutralized by a monoclonal antibody to the EIIIA segment. The-major goal of this continuation proposal is to extend these observations with studies of: (1) the impact of the inflammatory milieu on the lipocyte activating effect of the fibronectin EIIIA segment: elements to be examined include other variable segments in the -fibronectin molecule, soluble mediators of inflammation and the ECM context; (2) the minimum domain within EIIIA that is responsible for its activity; (3) the lipocyte receptor that recognizes the EIIIA segment; (4) the effect on fibrogenesis in vivo of anti-EIIIA antibodies or of peptides modeled on the active region of EIIIA; and finally (5) the regulation of endothelial production of A+-fibronectin by soluble mediators of inflammation, particularly transforming growth factor- B. The culmination of this work will be a new approach to therapy for fibrosis, based on competitive inhibition of the EIIIA segment for its lipocyte receptor.