Engraftment and albumin production of intrasplenically transplanted rat hepatocytes (Sprague-Dawley), freshly belated versus cryopreserved, into nagase analbuminemic rats (NAR)

Banking of cryopreserved hepatocytes is a prerequisite for large-scale hepa tocyte transplantation in the clinic. We compared the efficacy of intrasple nic transplantation into Nagase analbuminemic rats (NAR) of freshly isolate d (FIH) and cryopreserved (CH) hepatocytes. Hepatocytes were cryopreserved using a controlled rate freezing protocol. Albumin production of thawed CH and FIH was measured in vitro in culture by ELISA and by Western blot. Afte r in vivo intrasplenic transplantation of NAR with either FIH or CH we asse ssed 1) albumin in the serum of recipients by ELISA and by Western blotting analysis at different time intervals, and 2) hepatocyte engraftment by alb umin immunohistochemical staining into spleens and livers at euthanasia. In vitro, albumin was produced up to day 3 of culture in both CH and FIH. In vivo, no intrasplenic engraftment of hepatocytes occurred. Intrahepatic eng raftment of CH (cell number/mm(2)) was significantly (twofold) lower than t hat of FIH and appeared only as isolated cells and small (<10 cells) cluste rs, while bigger clusters (>10 cells) were observed with FIH. In the FIH gr oup, serum albumin production was observed up to 32-39 days posttransplanta tion while in the CH group no serum albumin production was detected. Our re sults emphasize the need to improve 1) hepatocyte transplantation procedure s either by repeated hepatocytes injections and/or by transplantation under a regeneration response, and 2) the freeze/thaw protocols of hepatocytes.