| Induced pluripotent stem cells (iPSCs) are a type of embryonic stem (ES) cell–like cells derived fromadult somatic cells via enforced expression of a specific group of transcription factors or combined withsome small chemical molecules. These cells exhibit the essential characteristics of ES cells and are capableof generating cells of all three germ layers. Studies of iPSCs not only promoted many basic fields of lifescience, such as the regulation of cell differentiation, regenerative medicine and cell development, but alsohave many potential clinical applications.In this study, we transfected four transcription factors of Oct4, Sox2, c-Myc and Klf4into rat primarycultured hepatocytes by recombinant lentivirus technology and attempted to generate induced pluripotentstem cells. After7days of infection, some hepatocytes exhibited a morphological change, and thentransferred them onto MEF feeder cells. For about11day's culture, rat ES-like clones appeared from someof hepatocyte-derived cells, which have a cutting edege, big nucleus and a high nucleus/cytoplasm ratio.Moreover, they expressed a high alkaline phosphatase activity as well.Reverse transcription PCR analysis results obtained from selected hepatocytes-derived clones, ratprimary cultured cells and rat embryos of4.5-days show that neither clones nor4.5-days embryos expresshepatocyte specific gene albumin, and exogenerous transcription factors of Oct4, Sox2, c-Myc and Klf4only appeared in hepatocytes-derived clones, while endogenerous Oct4, Sox2and some other embryospecific genes like Nanog, Fgf4, Lin28and Ncam were expressed in both clones and rat embryos.Immunofluorescence staining indicated that these clones expresse rat embryonic stem cell marker proteins,SSEA-1and Nanog.All above mentioned results show that we have successfully generated induced pluripotent stem cellsfrom rat adult hepatocytes. |
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