Induction of efficient differentiation and survival of porcine neonatal pancreatic cell clusters using an EBV-based plasmid expressing HGF.
Porcine neonatal pancreatic cell clusters (NPCCs) have been actively studied as a source of pancreatic stem cell transplantation for the treatment of diabetes. In this study, the hepatocyte growth factor (HGF) gene was cloned in an Epstein-Barr virus (EBV)-based plasmid vector (pEBVHGF) and the effects of the HGF expression on the survival and differentiation of NPCCs were analysed. For comparison, pHGF was constructed by deleting EBNA-1 and OriP from pEBVHGF. The expression of HGF, as measured by ELISA, lasted longer when pEBVHGF was used than when pHGF was used. C-Met phosphorylation co-related with the expression of HGF in the transfected NPCCs. Immunocytochemistry experiments showed that NPCCs showed a higher and longer expression of insulin when they were transfected with pEBVHGF than with pHGF. Moreover, a greater number of NPCCs survived for a longer period after they were transfected with pEBVHGF than when they were transfected with pHGF. Taken together, these results indicate that transfecting NPCCs with the HGF gene using an EBV-based plasmid is a more effective method of inducing differentiation to beta cells and enhancing survival than using a conventional plasmid. Therefore, it may be possible to use EBV-based plasmids to modify pancreatic stem cells for xenotransplantation.
Kim
Min Sun
MS
Research Institute of Immunobiology, Department of Biomedical Sciences, Catholic University of Korea, Seoul, Republic of Korea.
Kim
Ji-Won
JW
Sun
Chenglin
C
Oh
Sang Taek
ST
Yoon
Kun Ho
KH
Lee
Suk Kyeong
SK
D000818:Animals; D000831:Animals, Newborn; D002454:Cell Differentiation; D002470:Cell Survival; D004797:Enzyme-Linked Immunosorbent Assay; D017228:Hepatocyte Growth Factor / Q000235:genetics*; D004854:Herpesvirus 4, Human / Q000235:genetics*; D007150:Immunohistochemistry; D010179:Pancreas / Q000166:cytology* / Q000378:metabolism; D010766:Phosphorylation; D010957:Plasmids; D013552:Swine; D014162:Transfection
D017228:Hepatocyte Growth Factor