Transfection of Peripheral Blood Monocytes withSOX2Enhances Multipotency, Proliferation, and Redifferentiation into Neohepatocytes and Insulin-Producing Cells

Abstract

Following a several-day incubation in medium containing IL-3 and M-CSF to generate a more plastic intermediate “reprogrammed multipotent cells of monocytic origin (RMCMO),” peripheral blood mononuclear cells (PBMCs) can be efficiently converted to hepatocyte-like cells (neohepatocytes) and insulin-producing cells. However, continuous efforts are devoted to enhancing the proliferative capacity of these multipotent cells while maintaining or further increasing their redifferentiation potential. In the present work, PBMCs were transfected with one pluripotency gene (SOX2) and the resulting RMCMO compared to standard RMCMO with respect to cell viability, proliferative activity, and redifferentiation potential. EctopicSOX2expression increased the number of viable RMCMO, activated cell cycle genes, and enhanced proliferation as shown by quantitative RT-PCR and Ki67 immunofluorescent staining, respectively. Redifferentiation of RMCMO derived fromSOX2-transfected PBMCs to neohepatocytes was more complete in comparison to control cells as revealed by higher urea and glucose secretion, increased activity of cytochrome P450 isoforms, and a phase II enzyme, while the same was true for insulin-producing cells as assessed by the expression ofINS,PDX1, andGLUT2and glucose-stimulated insulin secretion. Our results indicate thatSOX2transfection increases both multipotency and proliferation of RMCMO, eventually allowing production of neohepatocytes and insulin-producing cells of higher quality and quantity for transplantation purposes.