Previously, our laboratory generated a panel of seven monoclonal antibodies (MAb) recognizing oval cell/bile duct surface antigens. These antibodies have been found to be valuable tools for studying liver ontogeny and carcinogenesis. As a result of successful completion of the specific aims of the current grant, we have identified and partially characterized the proteins recognized by two of these MAb. Using LC/MS, we have identified a surface-localized form of heat shock cognate 70 (Hsc70) recognized by MAb OC.10 and by primary amino acid sequence analysis, a rat member of the nectin/polio/herpes virus receptor family of Ig-like surface proteins recognized by MAb 324.5 In addition, we have discovered that in neutrophils, OC.2 resides in annexin 1+ gelatinase granules and that rat oval, prostate, and hepatoma cells express myeloperoxidase. Another major finding was the ability of MAb OC. 10 to enhance the growth and/or survival of bile duct epithelial cells in vitro and the morphogenesis of bile ducts in vivo. The three specific aims of the second revision of this competing renewal application are aimed at testing hypotheses regarding the expression and function of Hsc70, MPO and OC.2 that integrate our findings into the current understanding of liver development and carcinogenesis.
Specific Aim 1 will test the hypothesis that 1) the cell surface localization of OC.10/Hsc70 results from structural features not shared by its cytoplasmic counterpart and 2) that MAb OC. 10 binding to Hsc70 initiates signaling that increases proliferation and/or plays a central role in protecting bile duct cells against anoikis or apoptosis.
In specific Aim 2, we will test the hypothesis that MPO expression by oval cells provides a CYP450 independent mechanism for the initiation of bipotent ductal progenitors. This hypothesis is based on previous studies showing that MPO can activate procarcinogens and cause direct damage to DNA and cellular proteins by production of strong oxidants such as HOCl. The expression of MPO rat hepatocellular carcinomas with mixed oval/cell-hepatocyte phenotypes supports a role in liver development and carcinogenesis.
In specific aim 3, we will continue with our efforts to identify OC.2 and to characterize the OC.2+ granules found in bile duct and oval cells. The research proposed in this revised competing renewal application will not only offer new insights into pathways regulating the growth, survival and differentiation of ductal cells but will also provide increased knowledge of carcinogenesis pathways involving ductal progenitors.
