Synchronous Parenchymal and Non-Parenchymal Cell Proliferation Contributes to the Regenerative Growth of Engineered Livers

BACKGROUND: Recent success in hepatocyte transplantation toward the treatment of liver diseases has encouraged further investigation into bioengineering liver tissues as a novel cell transplantation therapy. Our group has developed several innovative experimental approaches to create liver tissues at ectopic sites by transplanting purified hepatocytes. We have also succeeeded in engineering biologically functional liver tissues that have an ability to perform regenerative growth at similar levels as naïve livers (Hepatology 41: 132, 2005, Nat Med 13: 880, 2007). To investigate further mechanism of the ectopic liver regenerative events, this study explores daily profile of the growth activity as well as parenchymal and non-parenchymal cell participations. METHODS: Isolated and purified mouse hepatocytes were transplanted under the kidney capsule of naïve mice for enigneering liver tissues as previously described. Functional volume of the engineered tissues were assessed by measuring serum marker proteins. At day 70, we then conducted 2/3 hepatectomy (PH) for inducing liver regeneraive stimulus. BrdU was then provided for each 24-hr period to the mice after the PH. By performing BrdU-staining, BrdU labeling index (LI) of hepatocytes and non-parenchymal cells were assessed. RESULTS: The functional volume of the engineered liver tissues stably maintained for 70 days period in all recipient mice. During the 7-day period after the PH, the serum marker protein levels gradually increased and reached to >200% of the prehepatectomy staus at day7, indicating that the engineered liver performed rapid regenerative growth. BrdU LI assessment revealed that BrdU LI of hepatocytes reached the peak at day 2 (2nd 24-hr period) and that of non-parenchymal cells reached the peak at day 3 (3rd 24-hr period). CONCLUSION: The present studies established that hepatocytes and surrounding non-parenchyaml cells cooperate together to accomplish the regenerative events of the engineered livers. This data suggests that ectopially transplanted hepatocytes have an ability to recruit liver-specific non-parencymal cell types to form complex liver tissues.