Study On The Immune Response Reaction Impact Of HUCMSCs In Combination With GAD On PBMC In Type 1 Diabetes In The MLR

Objective:To investigate the immune response reactionimpact of umbilical cord-derived mesenchymal stem cells (hUCMSCs) as well as in combination with glutamic acid decarboxylase antigen peptide (GAD) with allogeneic mononuclear cells (PBMC) in type1diabetes in the mixed lymphocyte reaction (MLR) and explore its mechanism.Methods:Establishing different concentrations of hUCMSCs as well as in combination with GAD and allogeneic PBMC in type1diabetes in co-culture system. Taking hUCMSCs as stimulative cells and GAD as the stimulus, PBMC of patients with type1diabetes as the reactive cell.It was divided into4groups according to the proportion of different concentrations. Group Ⅰ (the control group):1×105PBMC; group Ⅱ (the low concentration group):1×104hUCMSC and1×105PBMC; group Ⅲ (the high concentration group):1×105hUCMSC and1×105PBMC; group Ⅳ(the combination group):1×105hUCMSC in combination with GAD (5μg) and1×105PBMC. During mixed culture of72h, lymphocyte proliferation inhibition rate was determined by CCK-8, TNF-a, IL-2, IL-4in the supernatant was determined by ELISA,and CD4+、CD8+and CD4+CD25+T lymphocyte were determind by flow cytometry.Results:CD4+T and CD4+/CD8+T-proportion of type1diabetes patients was high.After the application of umbilical cord mesenchymal stem cells, compared with the control group,they were declined to some extent,while the proportion of CD8+T was significantly increased in a dose-dependent manner (P<0.05). In addition, our studies have shown that mesenchymal stem cells can also increase the number of CD4+CD25+regulatory T cells. In combination with GAD, compared with the stem cell group of pure high concentration, the level of TNF-alpha wasdeclined and the level of IL-4was increased,the proportion of CD4+T was reduced and the proportion of CD8+T was risen.Stem cells The level of IL-2and TNF-alpha of the stem cell group level was significantly decreased than the control group,while the level of IL-4was significantly increased (P<0.05).Conclusion:hUCMSCs inhibit immune responses of allogeneic PBMC of patients with type1diabetes in vitro as the dose-dependent manner; Inhibitation rate is enhanced when hUCMSCs are mixed with GAD. It provides a new approach for the clinical treatment of type1diabetes.