Different Transplant Bone Marrow Mesenchymal Stem Cells For Treating Diabetes Mellitus

Objective:To study the effects of different transplantation of bone marrow mesenchymal stem cells(BMSCs)on the Type 1 diabetes,BMSCs labeled by Enhanced greenfluorescent protein(EGFP- BMSCs) were transplatated into streoptozotocin-induced diabetic rats by pancreatic peplos and intravenous injection,Methods:Four male Sprague Dawley rats aged 1 week were used for establishing BMSCs.The Cells were isolated and culture by the whole bone marrow method,and then purified and amplified by adherence.We transfered EGFP gene into P3 BMSC,The expression ratio of EGFP gene and the proliferation ability was detected using FACS, BMSCs were successfully labed by EGFP.Thirty-two female Sprague Dawley rats abdominally injected with streptozotocin to establish diabetes models.Diabetes rats were equally and randomly divided into pancreas peplos transplantation group(group A),vein transplantation group(group B),diabetes control group(group C),and normal control group(group D).of EGFP-BMSCs(5×10~6/mL) from the third passage was suspended into the 0.2mL phsphate buffered saline,and multiply injected into pancreas peplos in the pancreas peplos transplantation group.In the vein transplantation group,rats were subjected to an equal volume of BMSC suspension via the vein.Rats in the diabetes control group and normal control group were left intact.Following transplantation,Blood glucose,insulin, C-peptide level changes,and glucose tolerance were observed dynamically after transplantation.Blood glucose level was monitored by Accutrend sensor comfort blood glucose meter.ELISAs were used to assay insulin and C-peptide in sera.42 days after transplantation,the pancreatic tissue of rats from each group was harvested to prepare paraffin sections.For histology of pancreas,cryo-sections were stained with hematoxylin and eosin.Fluorescent immunohistochemistry were used to analyze recipients' pancreas tissues for assessment the protein expression of insulin and EGFP.Pdx-1Mrna and SRY gene was detected by fluorescence in situ hybridization(FISH).Results:Compared with diabetic control group,blood glucose value significantly decreased in the pancreas peplos transplantation group and vein transplantation group within 30 days following transplantation(P＜0.05),and the decrease range in the pancreas peplos transplantation group was mostly significant(P＜0.05).At 2,4 and 6 days following transplantation,insulin and C-peptide levels significantly increased in the pancreas peplos transplantation group and vein transplantation group(P＜0.05),and the increase range was significant in the pancreas peplos transplantation group(P＜0.05),but still lower than in the normal control group(P＜0.05).Following intraperitoneal injection of glucose,blood glucose levels peaked at 30 minutes,decreased to fasting leveling at 90 minutes,and the glucose treatment ability was nearly to normal control group in the pancreas peplos transplantation group.The treatment ability to glucose in the vein transplantation group was slightly stronger compared with the diabetes control group(P＜0.05),but the levels did not reach a normal blood glucose level within 180 minutes in this two groups.HE-stained sections demonstrated increased islets in group A and B,and few lymphocyte present.HE stained sections demonstrated the number of islets was markedly increased in group A than that of diabetic controls(P＜0.05) and similar to that of normal controls(P＞0.05). EGFP-BMSCs were not detected in group B,C and D.Analysis of pancreatic sections for EGFP in pancreas peplos transplantation group indicated the transplanted BBMSCs survived within pancreas.Fluorescent immunohistochemistry showed insulin protein and EGFP are coexpressed in pancreas peplos transplantation group.Fluorescence in situ hybridization (FISH) showed some cells expressed SRY gene in Y chrosome are coexpressed with Pdx-1 mRNA in pancreas peplos transplantation groupConclusion:Whether BMSCs transplantation by femoral vein injection or by pancreatic peplos injection,the two methods both have therapeutic effect for diabetes.While the effect of MSCs transplantation by pancreatic punctiform injection is better in alleviating diabete symptom and decreasing blood glucose level than BMSCs transplantation by femoral vein rejection,and MSCs transplantation by pancreatic punctiform injection can promote MSCs trans-differentiate into pancreatic islet-like cells that can secrete insulin.So MSCs transplantation by pancreas peplos injection may become new pathway of cell transplantation to treatment diabetes.