Effects Of Ngn3 Overexpression On The Differentiation Of Human Umbilical-cord Mesenchymal Stem Cells Into Insulin-producing Cells

Objective: To explore whether neurogenin3(Ngn3)overexpression can promote the differentiation of human umbilical cord mesenchymal stem cells(hUCMSCs)into insulin-producing cells(IPCs).Methods: HUc MSCs were isolated and cultured,identified by flow cytometry,and identified by lipid formation and osteogenesis induction.HUcMSCs were induced into IPCs in different stages,including a low glucose(containing EGF)induction stage and a high glucose(containing activin A and nicotinamide)induction stage.The experiment was divided into two groups: The control group was induced by the above scheme,while the experimental group was additionally infected the lentivirus overexpression vector carrying the target gene Ngn3 on the the optimal time of the same induction process.After induction,the changes of cell structure were observed by electron microscope;mRNA and protein were collected and Real-Time PCR and Western blot were used to compare the expressions of insulin and MafA in the two groups;medium supernatant was collected and C-peptide content was determined by ELISA.Results: HUcMSCs were successfully isolated with positive expression of CD105 and CD90 and negative expression of CD34 and CD45,which had adipogenic and osteogenic differentiation ability.Then,hUcMSCs were induced into IPCs by stage induction,and the cells expressed MafA and insulin positively.Ngn3 was overexpressed in the experimental group during the induction.After induction,electron microscopy showed that the cell structure was more mature in the experimental group.The expression levels of insulin and MafA in the experimental group were significantly higher than those in the control group.During the induction process,the amount of C-peptide secreted by the experimental group was higher than that of the control group.Conclusion: Lentivirus-mediated Ngn3 overexpression improved the differentiation efficiency of hUcMSCs into IPCs.