Liver repopulation after cell transplantation in mice treated with retrorsine and carbon tetrachloride

Background. Efficiency of engraftment after liver cell transplantation is less than 1% under conventional conditions. Our aim was to develop a high-efficiency, nonsurgical, no-genetic-advantage mouse model of liver repopulation with transplanted cells. Methods. Mice were conditioned with nonlethal doses of a cell cycle inhibitor, retrorsine, 70 mg/kg, to irreversibly block proliferation of native hepatocytes. After the drug was eliminated, 2 million freshly isolated beta-galactosidase-labeled liver cells were transplanted into the spleens of C57BL/6J recipient mice. To stimulate donor cell proliferation, three doses of carbon tetrachloride (CCl4), 0.5 ml/kg, were given. Several control groups were studied to evaluate the contribution of each treatment to liver repopulation. Results. Repopulation, as measured by cell isolation from recipient livers 1-7 months after transplantation, was on average 20%. Repopulation was 10% if CCl4 was given only once, between 0.5% and 1% if only retrorsine or CCl4 were used, and 0.05% if no conditioning was used. Phenotypically, whole livers turned blue on exposure to X-gal staining, whereas negative (control) livers remained pale brown. More than 55% of liver repopulation resulted from clusters containing 21 or more cells, some of which contained more than 200 cells, suggesting seven or more rounds of cell division in a subset of transplanted cells. Conclusion. This murine study demonstrates high levels of repopulation after liver cell transplantation into nongenetically modified livers, using a cell cycle inhibitor and chemical liver injury to provide transplanted cells a proliferative advantage. Liver repopulation was effected mostly by a small fraction of transplanted cells. Analogous nonsurgical liver cell transplantation strategies, but with clinically applicable drugs, could be devised for the treatment of liver-based metabolic diseases.