Study Of Hepatocyte Transplantation By Splenic Artery In SD Rats

Object:To research the improvement of method and the influencingfactors of hepatocytes isolation technology in SD rats；To establish theanimal model of hepatocyte transplantation via splenic artery in SD rats; toevaluate the treatment effects of hepatocyte transplantation by splenicartery and intrasplenic injection in rats with acute hepatic failure.Methods: Hepatocyte was isolated by the two step collagenesedigestion, and the method was improved in various conditions such asperfusion in the rats. Acute hepatic failure (AHF) was induced by D–gal inSD rats. After24hours, a cather was inserted into the splenic artery in65rats with AHF. Then the60rats were randomly divided into3groups.GroupⅠreceived2×107hepatocytes through intraplenic injection and0.4ml Hank’s solution was infused through splenic artery; GroupⅡ received0.4ml Hank’s solution through intraplenic injection and2×107hepatocyteswas infused through splenic artery; Group Ⅲ received0.4ml Hank’ssolution through intraplenic injection and0.4ml Hank’s solution throughsplenic artery. Surviving rate was observed at14days and liver functionwas measured at different time points. The distribution of transplanted CFDA–SE–labeled hepatocytes through splenic artery was observed.Secretion of albumin and liver histopathological change were observed inspleen by immunofluorescence and HE staining.Results: Highly viable (85-95%)hepatocyte suspension were stained.At14days, the survival of Group Ⅰrats was significantly higher than thatof Group Ⅱ rats （P=0.031,<0.05）; the survival of Group Ⅱwassignificantly higher than that of Group Ⅲ (P=0.048,<0.05). Hepaticfunction of GroupsⅠand Ⅱhad improved, especially of GroupⅠ. In GroupⅡ,24h after transplantation of CFDA–SE–labeled hepatocytes, scatteredgreen fluorescence region was found under fluorescent microscopy in thespleen and liver. In GroupⅠandⅡ, at14days post–transplatation,immunofluorescent staining of albumin demonstrated some positive cells inspleen; at7days post–transplatation, transplanted hepatocytes was found inspleen.Conclusion: There were many influence factors in the success ofhepatocyte separation, such as collagen enzyme, PH value, perfusion fluidand perfusion method, etc. Hepatocyte transplantation throughcatheterization splenic artery via carotid route can improve the survival ofrats with AHF and ameliorate hepatic function, but intrasplenic injection issignificantly superior to it.