TRANSPLANTATION OF HIGHLY DIFFERENTIATED IMMORTALIZED HUMAN HEPATOCYTES TO TREAT ACUTE LIVER FAILURE

Abstract

26 Introduction: Because the liver has an enormous capacity for regeneration, temporary support of a damaged liver by a bioartficial liver (BAL) devise is a promising approach for the treatment of acute liver failure. Large quantities of viable and actively functioning hepatocytes are required for BAL treatment of ALF or as a bridge to liver transplantation. However, the shortage of human livers available for hepatocyte isolation severely limits the use of this modality. A clonal hepatocyte cell line that could be grown economically in tissue culture and would exhibit liver-specific function would be an attractive solution to this problem. Purpose: To test this alternative, primary human fetal hepatocytes were immortalized by transfection with a plasmid pSV3neo containing the genes encoding Simian virus 40 large T antigen (SV40Tag) and neomycin phosphotransferase. One of the immortalized cell lines, OUMS-29 secreted albumin, expressed mRNAs for CYP 1A1, 1A2 and 2E1 and showed an ammonia removal activity. OUMS-29 cells were not tumorigenic after transplantation into athymic nude mice. In order to investigate the potential of OUMS-29 cells, cells were transplanted into the spleen of rats followed by 90% hepatectomy. Methods: Rats were divided into the following groups: Group1 (G1:n=12), intrasplenic transplantation of 20×106 OUMS-29 cells; Group2 (G2:n=5), intrasplenic transplantation of 20×106 HepG2 cells (transformed human liver cells); Group3 (G3:n=10), intrasplenic injection of 0.5ml medium. All rats underwent 90% hepatectomy 12 hr after transplantation. Results: Rats in G1 showed a significantly lower increase in blood ammonia levels after 90% hepatectomy. Six out of 12 animals in G1 survived more than 48 hr after hepatectomy, whereas all animals in G2 and G3 died of acute liver failure within 28 hr. Survival of G1 rats was significantly prolonged compared to G1 and G2 rats. Conclusions: Intrasplenic transplantation of OUMS-29 cells protected animals from hyperammonemia and prolonged survival in 90% hepatectomized rats. The present studies show that primary human hepatocytes can be immortalized, expanded in culture with retaining a highly differentiated liver function, and are capable of providing metabolic support in acute liver insufficiency. OUMS-29 cells offer the advantages of unlimited availability, uniformity, sterility and freedom from infectious pathogens and these cells might be clinically used for hepatic function in BAL to improve the lives of a large number of patients with ALF.