MafA Overexpression: A New Efficient Protocol for In Vitro Differentiation of Adipose-Derived Mesenchymal Stem Cells into Functional Insulin-Producing Cells.

ABSTRACT: Objective:We proposed a novel differentiation method for the efficient differentiation of adipose-derived mesenchymal stem cells (ADMSCs) into functional insulin-producing cells (IPCs) based on MafA overexpression. Materials and Methods:In this experimental study, a eukaryotic expression vector containing MafA [MafA/pcDNA3.1(+)] was constructed and purified. ADMSCs were differentiated into IPCs. ADMSCs were assigned in two groups including control (C), and the MafA overexpressed (MafA+) groups. The ADMSCs were transfected by MafA/pcDNA 3.1(+) at day 10 of the differentiation. Differentiated cells were analyzed for the expression of multiple ? cell specific genes (Nkx2.2, Ngn3, Isl-1, Pdx1, MafA, Nkx6.1, and Insulin) using real-time polymerase chain reaction (PCR). The insulin secretion potency of the differentiated cells in response to glucose exposure was also determined using an enzyme-linked immunosorbent assay (ELISA) method and Dithizone (DTZ) staining. The IPCs from the control manipulated group, and un-differentiated ADMSCs group were transplanted to streptozotocin (STZ)-diabetic rats. Rats were monitored for blood glucose and insulin concentration. Results:The results revealed that ADMSCs were successfully differentiated into IPCs through the 14 day differentiation protocol. The expression of ?-cell specific genes in MafA+ IPCs was higher than in control cells. Glucose-induced insulin secretion after the exposure of IPCs to glucose was higher in MafA+ group than the control group. The STZdiabetic rats showed an ability to secrete insulin and apparent hyperglycemic condition adjustment after transplantation of the control IPCs. The mean insulin concentration of diabetic rats that were transplanted by manipulated IPCs was significantly higher than ADMSCs-transplanted rats; however, no effect was observed in the concentration of bloodn glucose. Conclusion:The overexpression of MafA can be used as a novel promising approach for the efficient production of IPCs from ADMSCs in vitro. However, the future therapeutic use of the MafA+ IPCs in diabetic animals needs further investigations.

SUBMITTER: Dayer D 

PROVIDER: S-EPMC6397604 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature