Effects of ischemia-reperfusion preconditioned hepatocytes on cocultured islet cells

Abstract

Introduction: Hepatic ischemia and insufficient neovascularization of transplanted islets are considered to be barriers to islet survival and function. Hepatocytes are well known to be regenerative. Additionally, hepatocytes have a protective mechanism against ischemia. Based on the effects of hepatocytes under ischemia-reperfusion preconditioning, I hypothesized that ischemia-reperfusion preconditioning of hepatocytes might beneficially affect islet cells cocultured with hepatocytes. Thus, a cell line coculture model of RIN-5F (insulin secreting cell line) and Hep-G2 (hepatoma cell line) cells was used, as well as a primary islet and hepatocyte coculture model of cells from Sprague-Dawley rats to confirm the beneficial effects of ischemia-reperfusion preconditioning.Methods: Hep-G2 cells and primary isolated hepatocytes were incubated in a hypoxic chamber under 0% or 1% O2 (hypoxic or hypoxia-normoxia) conditions. The RIN-5F cells were cocultured with Hep-G2 cells incubated under different O2 hypoxia conditions. Primary islets were cocultured with primary hepatocytes under hypoxia or hypoxia-normoxia-hypoxia single and double settings for 15 and 30 min, which were determined for the ischemia-reperfusion preconditioning (IRP) of hepatocytes.Results: Insulin secretion was increased in the RIN-5F cells and primary islets that were cocultured with the ischemic-preconditioned Hep-G2 cells and hepatocytes. No changes were observed in hepatocyte cell viability, indicating that IRP did not damage cell viability. IL-6-STAT3 pathway activity, which is known to be beneficial to hepatocytes after ischemic injury, was also increased by hypoxia and IRP. The levels of IGF1, HGF, TGF-α, and TGF-β were also increased by IRP in hepatocytes. Insulin secretion and the expression levels of the survival-related genes Bcl-2 and Reg-1α in the islets were increased by coculture with IRP hepatocytes.Conclusion: These results suggest that IRP of the liver or hepatocytes could improve islet survival and insulin secretion under conditions of intraportal transplantation.