The Roles of Different Multigene Combinations of Pdx1 , Ngn3 , Sox9 , Pax4 , and Nkx2.2 in the Reprogramming of Canine ADSCs Into IPCs.

Adipose-derived mesenchymal stem cells (ADSCs) are ideal sources for the treatment of diabetes, and the differentiation of ADSCs into insulin-producing cells (IPCs) through transfection of exogenous regulatory genes in vitro has been studied in depth. The differentiation of ADSCs is strictly regulated by a variety of transcription factors such as Pdx1 , Ngn3 , Pax4 , Nkx2.2 , and Sox9 . However, whether these genes can coordinately regulate the differentiation of ADSCs into IPCs is still unknown. In this study, five multigene coexpressing adenovirus vectors ( pAdTrack-Pdx1-Ngn3-AdEasy , pAdTrack-Pdx1-Ngn3-Sox9-AdEasy , pAdTrack-Pdx1-Ngn3-Pax4-Sox9-AdEasy , pAdTrack-Pdx1-Ngn3-Nkx2.2-Sox9-AdEasy , and pAdTrack-Pdx1-Ngn3-Nkx2.2-Pax4-AdEasy ) were constructed, and then the stocks of the packaged adenoviruses were used to infect the canine ADSCs (cADSCs). Based on results of morphological observation, dithizone staining, sugar-stimulated insulin secretion test, cellular insulin immunofluorescence assays, and the detection of pancreatic β-cell development-related genes in the induced cells, the best induction combination ( pAdTrack-Pdx1-Ngn3-Nkx2.2-Pax4-AdEasy ) was identified after comparative screening. This study provides a theoretical reference and an experimental basis for further research on stem cell replacement therapy for diabetes.