| Transplantation of islets is expected to be an effective and final method to treat diabetes. The procedure of islets transplantation is simple, with reduced use of exogenetic insulin, which can reduce patient’s risk of complications, and improve their live quality. In recent years, with the extensive development of the research on islets transplantation, the technology of transplantation also made great progress, but insufficient number of islets and postoperative immunologic rejection are still the biggest obstacles of islets transplantation for the final use to clinical.To establish an effective plan of immune tolerance of islets transplantation, we established an efficient islets separation and transplantation method, and explored the immune tolerance potential mechanisms and application value of the transplantation of islets with immature dendritic cells (imDCs) and mesenchymal stem cells (MSCs).The main research contents of this paper are summarized as follows:1. To develop animal model of the diabetic mouse. In this study, different doses of streptozocin (STZ) were used to intraperitoneally inject to the mice (200 mg/kg,150 mg/kg and 100 mg/kg). Research results showed that a dose of 150 mg/kg was better than a dose of 200 mg/kg. Though both of the doses are effective to elevating blood glucose in mice, the high-dose may cause greater damage to mice, resulting in the death of some mice, which is not conducive to the establishment of the model.2. To establish methods of isolation and purification of rat islets. Islets transplantation is an important reason for the effect of islets graft quality and insufficient quantity. For research, this study established a pancreatic collagenase perfusion and density gradient centrifugation method to purify islets. The results indicated that the purity and quality of islets were remarkably improved.3. To establish an efficient and stable method for simultaneous isolation and culture of immature dendritic cells (imDCs) and bone marrow mesenchymal stem cells (BM-MSCs). The cultured dendritic cells were identified by flow cytometry. The results showed that mouse bone marrow-derived dendritic cells with the expressions of CDllc, MHC-Ⅱ, CD80 and CD86 were 89.9%,36.61%,5.03%,1.5%, respectively, in line with the imDCs surface marker characteristics. Osteogenic and adipogenic differentiation induction were performed on mesenchymal stem cells in vitro. The results showed that mesenchymal stem cells induced from murine bone marrow still have multiple differentiation potential.4. Application of imDCs and MSCs in islet transplantation for the treatment of diabetic mice. From the results of co-transplantation groups, it was found that transplantation did not prolong graft survival (11±2.31 d) significantly, but significant decreased blood glucose and glycosylated hemoglobin concentration in the diabetes of mouse by co-transplantation of islets and imDCs. Co-transplantation of islets with MSCs could significantly decrease blood glucose level as well as prolong graft survival for days (12±2.65). While transplantation of islets with MSCs and imDCs could not only restore normal blood glucose level, but also remarkably prolong graft survival for more than 15 days (15.6±3.48). This results demonstrate that co-transplantation of islets with imDCs or MSCs can promote graft survival and reverse blood glucose level, and effectively control the glycosylated hemoglobin concentration as well, which is better than islets transplantation alone. |
