The elucidation of the cellular progenitors of neoplasia is an essential step in deciphering the mechanisms involved in chemically induced hepatocarcinogenesis. To this aim we will use several different transplantation models to examine the biological potential of highly purified preneoplastic cell populations isolated from the livers of LE and ACI rats following treatment with diethylnitrosamine by the Solt/Farber protocol, ethionine in a choline deficient (CD) diet or 2-acetylaminofluorene and partial hepatectomy. Immunoselection by flow cytometry, panning, adsorption with antibody coated magnetic beads or selective killing with indirect immunotoxins will be used to further purify nonparenchymal cell (NPC) and preneoplastic nodule (PN) cell fractions isolated from collagenase dissociated livers. Immunization schemes designed to restrict the immune response against normal hepatocyte antigens will be used in the production of subpopulation specific monoclonal antibodies (MAbs). Attempts will also be made to generate MAbs recognizing the heterotetrameric forms of aldolase A and B and pyruvate kinase L and K. An existing panel of MAbs will be used in combination with new MAbs generated in the proposed studies to define the antigenic phenotype and the biological potential of colonies resulting from the transplantation of purified cell populations into fetal (ACIxLE)F1 rats and into the liver, spleen, fat pads and pancreas of adult F1 rats. The donor origin of colonies and hepatic lesions in F1 host rats will be established using alloantisera. Biological potential will also be examined by effecting the complete transformation of preneoplastic cell populations with N-methyl-N-nitrosourea or by examining the differential progression of cell populations induced following treatment with 4,4'-diaminodiphenylmethane in CD or choline sufficient diets. Immunocytochemical and immunochemical techniques will be used to analyze the structure and function of MAb defined antigens. Of particular interest will be the oval cell antigens defined by MAbs 270.38 and 374.3 and the PN antigen recognized by MAb 374.8. These studies will contribute to our knowledge of the cellular lineages leading to hepatocellular carcinoma and will provide insight into the antigenic alterations associated with progression to malignancy.
