| Introduction Acute pancreatitis is a common acute abdomen, which is one of the risks of clinical manifestations, complications, and mortality high. Its pathogenesis is not clear now .It is possible that trypsin was abnormal activated and activated leukocyte in the process of pancreatic tissue damagement. Leukocyte activation and excessive cytokine cascade theory was such important causes and pathogenesis that the SAP caused systemic inflammatory, multi-organ failure and death. Most existing treatments are gastrointestinal decompression, sedation, spasm, inhibit secretion of trypsin, acid and maintain water and electrolyte balance, hemodialysis, peritoneal dialysis, antibiotics, application, medicine, nutrition support. The other part of SAP need surgical drainage when secondary infection of pancreatic necrotic tissue was taken place. But it is difficult to completely block the inflammatory cascade and the development of the course because the current lack of specific treatment was effectiveness.The pancreatic self-repair mechanism is unclear after injury happened. At present, adult stem cells of the pancreas and bone marrow-derived stem cells are important factors. Adult multipotent stem cells or similar cells would be fundamentally when the function and structure of the pancreas were to an ideal repair.Because separation and purification of pancreatic adult stem cells were more difficult,it is restricted in their therapeutic applications. Unfortunately, the current research in this area have not yet broken through.MSCs (marrow-derived mesenchymal stem cells) are adult stem cells in bone marrow. Under certain conditions, MSCs could differentiate into more than three sources of mesoderm cells, such as vascular, muscle, lung or pancreas, including nerve cells . A number of animal experiments and clinical trials showed that BMSc can reach the other organs to involve in physiological and pathological damage repair with the systemic blood circulation. Recent studies have shown that MSCs could exert the protection of damaged tissue and repair functions through the secretion of cytokines and regulatory proteins, inhibition apoptosis, alleviation the organ fibrosis and the role of anti-inflammatory response in addition to differentiation . Because of the pancreatic apoptosis and necrosis as well as the systemic inflammatory response syndrome (SIRS) are the important aspect about the occurrence of the pathophysiological process of SAP, MSCs may be new ideas for the treatment of SAP.Based on these study and the characteristics of pancreatic microcirculation, as well as the sensitive to hypoxia-ischemia. In this study, a rat model of acute pancreatic injury was successlly established by ligation of pancreatic tissue of pancreatic tail. Allogeneic MSCs was injected into the rat pancreatic necrosis model by the rat tail vein and its repairment of pancreatic injury was investigated and its possible mechanism to repair pancreatic injury was explored. The purpose of this study was to provide a reliable, objective and scientific basis on MSCs to treat pancreatic injury, pancreatitis. The value of research and clinical applications of the future is immeasurable.Method:1.We collect tibia and femur bone marrow of SD rat and MSCs was separated and purificated by adherent culture.Then we make primary culture and serial subcultivation in vitro and observe the cell morphology and cell growth characteristics by inverted phase contrast microscope.â‘ MSCs were induced to differentiate into osteoblasts and fat cells in vitro. The results was confirmed by alkalinephosphatase staining and oil red O staining .That was confirmed that MSCs possessed multipotency.â‘¡The expression of CD34 was determined by flow cytometry the molecular phenotype of MSCs was confirmed.â‘¢Immunological characteristics of MSCs was verificated through the expression of MHC II molecule by Immunohistochemistry.2.The rat ischemic model of acute pancreatitis was set up through ligating pancreatic tail end .3.Allogeneic MSCs was labeled DAPI and transplanted into animal models. The pancreatic tissue was taken and taken into frozen section.The migration and foregone conclusion of MSCs was observed through confocal microscope.4.Histopathology on pancreatic repairment of MSCs was observated and its mechanism was preliminary studied.Results1.The purified and steadily passaged MSCs were gotten by adherent culture with digestion after MSCs were isolated from mouse bone marrow.The result of cell cycle showed that more MSCs were G1/G0 phrase,it suggested that MSCs possessed multipotency. Based on the following: â‘ MSCs of rat didn't express CD34 by flow cytometry.â‘¡The expression of surface MHCâ…¡molecules on MSCs of rat was negative by immunocyto-chemistry.â‘¢It showed that alkaline phosphatase staining was positive that the dark grey sedimentation appeared in the osteoblasts differentiated from MSCs.That confirmed MSCs Obtained by this method could differentiated into osteoblast-like cellsâ‘£It showed that Fat-like cells was induced in vitro and oil red O staining was positive,which proved that MSCs could differentiated into fat-like cells.2.The pancreatic tissue was observed 6 hours after ligation in general and pathology.Liquid exudation in the abdominal cavity can be seen clear, and the pancreatic tail department became dark. The inflammatory cell infiltration, cell structure unclear, islet structure collapse,large necrotic tissue, hemorrhage and necrosis, in adipose tissue were observed in pancreatic tissue pathology.All these showed a rat model of pancreas with severe injury was successfully set up.3.The results about repairment of MSCs on injury pancreatitis were following: allogeneic MSCs began to move and settle in the damaged microvascular and organizations of pancreas 24 hours after it was implanted the model rats by the tail vein of rats and involve in pancreatic tissue repair. The organizational structure of the pancreas regenerated step by step, islets built, exocrine pancreas function gradually recoveried. The enzyme levels of blood amylase, lipase in the treatment group were obviously lower than that in the model group .The expression of glucagon and insulin by immunohistochemical detection showed that exogenous MSCs in the body can be transformed into islet-like cells and begin to secret glucagon and insulin.4.We Preliminary studied the repair mechanism of MSCs on rat pancreatic tissue.DAPI-labeling MSCs were found in pancreatic tissue by laser scanning confocal microscope.Sry gene were identificated by PCR and it appeared in the treatment group.All showed that MSCs could migrate to injured tissues and involve in the repairment of acinar, islet and mesenchymal structures in the pancreas. Apoptosis genes of Bcl-2 and Bax in pancreatic cells were detected by immunohistochemistry.The positive for Bcl-2 staining in MSCs treated group was significantly higher than that in the model group, while Bax-positive staining was significantly lower than the model group,which suggested that MSCs may inhibit apoptosis and promot tissue repair through the increase of Bc1-2 and inhibition of Bax.Conclusion:1.In this study, bone marrow mesenchymal stem cells of rats were successfully isolated and cultured through screening adherent and the whole bone marrow culture. Biological characteristics of the cells and the detection of surface antigens through flow cytometry confirmed that adherent cells were bone marrow-derived mesenchymal stem cells.So MSCs can be isolated, purificated, amplificated in vitro and obtained the higher purity of MSCs with multi-differentiation potential.This would provide sufficient MSCs products and a good experimental basis for further study of BMSCs used as seed cells for the treatment of diseases.2.Allogeneic MSCs began to move and settle in the damaged microvascular and organizations of pancreas 24 hours after it was implanted the model rats by the tail vein of rats .They could gradually repair and regenerate pancreatic acinar and blood vessels,as well as islet structure, so that the pancreas function of internal and external secretory has been gradually improved. The results of this study, not only laid a good foundation for human clinical application research, but also provided a road for the problems we encountered problems-the repair and regeneration of acute pancreatic injury. Its clinical significance and clinical application prospect is immeasurable. In addition, the results suggest that allogeneic MSCs transplantation can successfully repair and regenerate islet structure and glucagon and insulin secretion tends to improve the perfect level.Would it bring new hope to the therapy on diabetes which is high worldwide incidence and is not possible to cure atpresent? It remains to be more in-depth research.
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