胰岛细胞联合骨髓间充质干细胞移植对大鼠胰岛细胞再血管化的实验研究

Experimental study on revascularization of rat islet cells combined with bone marrow mesenchymal stem cells transplantation

目的探讨大鼠胰腺被膜下胰岛细胞联合骨髓间充质干细胞(bone marrow mesenchymal stem cell,BMSCs)移植能否促进胰岛的再血管化,进而减少移植后胰岛细胞的丢失,提高胰岛细胞移植的成功率。方法40只Sprague-Dawley大鼠建立糖尿病模型。待BMSCs及胰岛细胞培养鉴定后开腹,对照组于胰腺被膜下注射等量生理盐水,胰岛细胞组和BMSCs组于相同部位分别移植胰岛细胞和BMSCs,胰岛细胞+BMSCs联合组(联合移植组)同时于相同部位移植胰岛细胞和BMSCs,每组10只大鼠。检测各组大鼠不同时间点血清胰岛素及血糖水平的变化,实时定量PCR方法检测胰腺组织中血管生成相关因子缺氧诱导因子-1α(hypoxia inducible factor-1α,HIF-1α)、基质细胞衍生因子1α(stromal derived factor 1α,SDF1α)、血管内皮生长因子(vascular endothelial growth factor,VEGF)、碱性成纤维细胞生长因子2(basic fibroblast growth factor 2,FGF2)的mRNA表达量。结果(1)胰岛细胞组和联合移植组在术后第15天时血糖水平明显低于对照组和BMSCs组(P<0.05)且降至正常水平,联合移植组一直至第29天时仍能维持正常水平,而胰岛细胞组于第15天后开始升高且与BMSCs组接近(P>0.05)。(2)相较于对照组和BMSCs组,胰岛细胞组和联合移植组在术后第1、3、7、15、29天均表现为较高水平胰岛素释放(P<0.05),尤其是联合移植组更明显。(3)联合移植组胰腺组织中HIF-1α、SDF1α、VEGF、FGF2 mRNA相对表达量均分别高于其他3组且差异有统计学意义(P<0.05)。结论从本研究初步研究结果看,胰腺被膜下胰岛细胞移植术后可降低糖尿病大鼠血糖水平,但单纯胰岛细胞移植第15天后降糖效果逐渐减弱,而胰岛细胞联合BMSCs移植术后大鼠血糖控制平稳且作用时间更长,而且大鼠血管生成相关因子mRNA相对高表达,结合病理切片提示BMSCs有促进移植胰岛血管再通可能。

Objective To investigate whether transplantation of islet cells combined with bone marrow mesenchymal stem cell(BMSCs)of the pancreatic subcapsular promoting revascularization of pancreatic islets in rats,so as to reduce the loss of islet cells after transplantation and improve the success rate of islet cell transplantation.Methods The model of diabetic rat was established.The BMSCs and islet cells were cultured and identified,then the simple islet cells,simple BMSCs,and combination of islet cells and BMSCs were injected into the pancreatic subcapsular of the islet cell group,BMSCs group,and combination group,respectively.In addition,the same amount of normal saline was injected into the same site as the control group.There were 10 rats in each group.The changes of blood glucose and serum insulin in different time point were detected in each group.The mRNA expressions of angiogenesis factors such as hypoxia inducible factor-1α(HIF-1α),stromal cell derived factor 1α(SDF1α),vascular endothelial growth factor(VEGF),and basic fibroblast growth factor 2(FGF2)were determined by real-time quantitative PCR.Results(1)The blood glucose levels of the islet cell group and combination group were lower than those of the control group and the BMSCs group within 15 d after surgery(P<0.05)and decreased to the normal level,which of the combination group could still maintain the normal level until on day 29(P<0.05),but which of the islet cell group began to increase on day 15 after surgery and was similar to that in the BMSCs group(P>0.05).(2)Compared with the control group and the BMSCs group,the insulin levels were higher in the islet cell group and combination group on day 1,3,7,15,and 29 after surgery(P<0.05),especially in the combination group.(3)The expression levels of HIF-1α,SDF1α,VEGF,and FGF2 mRNAs in the combination group were higher than those the other three groups,and the differences were statistically significant(P<0.05).Conclusions Islet cell transplantation of pancreatic subcapsular could decrease blood glucose level in diabetic rats.Hypoglycemic effect of single islet cell transplantation gradually weakens on day 15 d after surgery.After BMSCs combined with islet cells transplantation,the glycemic effect of rat is stable for a longer time.Expressions of angiogenesis factors of BMSCs combined with islet cells transplantation rat are high,which combined with pathological sections suggests that BMSCs could promote vascular recanalization of islet transplantation.