Monitoring engraftment of transplanted hepatocytes in recipient liver with 5-bromo-2'-deoxyuridine.

Abstract

For studies on the intrahepatic engraftment of transplanted hepatocytes, labeling of donor cells is necessary. Current labeling techniques enable only short-term monitoring of engraftment. In the present study, we describe the use of 5-bromo-2'-deoxyuridine (BrdU) for a more permanent hepatocyte labeling. BrdU is stably incorporated into replicating DNA; consequently, BrdU labeling was performed in regenerating livers. In 10 Lewis rats, a two-thirds partial hepatectomy was performed, followed by continuous, low-dose BrdU administration. This approach provided a fraction of 89+/-1.5% BrdU-labeled donor hepatocytes, without influencing the efficacy of the ensuing isolation of donor hepatocytes. Subsequently, +/-1 x 10(7) isolated hepatocytes were transplanted either intraportally or intrasplenically into syngeneic recipients, and the engraftment of transplanted cells was evaluated in liver lobes at successive time intervals after transplantation. BrdU-positive hepatocytes could be identified and quantitated in recipient livers up to 180 days after transplantation. Repetitive quantitative assessments over time revealed an initial, drastic loss of transplanted cells (<24 hr), followed by a stabilization at approximately 7% of the injected cells. Histological monitoring showed that during this period (<48 hr) the transplanted cells migrate from the portal venules to the liver parenchyma. In recipient livers a homogeneous lobe distribution of hepatocyte engraftment was found 30 days after both intraportal and intrasplenic transplantation. Moreover, no significant difference between the intrahepatic liver cell engraftment of the two transplantation routes was demonstrated. In conclusion, the BrdU-labeling technique of donor hepatocytes enables long-term histological monitoring and quantitative evaluation of the engraftment of transplanted liver cells in recipient livers.