骨髓间充质干细胞对大鼠到小鼠胰岛移植的保护作用

Bone Mesenchymal Stem Cells Induced Immunotolerance on Rat to Mouse Islet Transplantation

目的探讨同种异基因骨髓间充质干细胞(bone mesenchamal stem cells,BMSC)静脉输注对大鼠到小鼠胰岛移植物的功能保护和小鼠糖尿病状态改善。方法全骨髓培养法获得C57BL/6小鼠BMSC。不连续梯度离心法分离纯化Sprague-Dawley(SD)大鼠胰岛,将300胰岛当量的胰岛单独或与BMSC联合移植入链脲菌素诱导的糖尿病BALB/c小鼠肾包膜下,并通过尾静脉在移植后0、3和5d注射CM-DiI标记的BMSC 5×105/只,对照组给于磷酸盐缓冲溶液。移植后监测血糖,第9天处死小鼠,取肝、脾、胸腺、淋巴结和移植胰岛的肾脏,冰冻切片,荧光显微镜观察CM-DiI标记细胞的组织分布;免疫荧光法观察移植物中胰岛素和胰高血糖素表达,评价胰岛的功能。结果 BMSC静脉输注后主要分布于胸腺,其次是脾脏和淋巴结,肾和肝组织中未观察到BMSC;BMSC联合胰岛移植组血糖控制水平优于其他组,且在第7天的口服糖耐量实验优于单纯胰岛移植组。结论与胰岛联合移植的BMSC对受者免疫器官和组织有明显的趋向性,且对胰岛细胞的体内存活有一定保护作用。

Objective To research on the protection function by the allogeneic rat bone mesenchymal stem cells（BMSC） on rat to mouse islet transplantation and the improvement of diabetic state in mouse.Methods BMSC were prepared from C57BL/6 mouse bone marrow cells and identified by flow cytometry（FCM）.Islets were isolated from Sprague-Dawley（SD） rats with Ficoll discontinuous centrifugation.CM-DiI labeled BMSC at 5×105 for one mouse were intravenously infused into STZ induced diabetic BALB/c mice after rat to mouse islet transplantation at day 0,3 and 5.Mice with PBS intravenously infused after islet transplantation were set as the negative controls.Blood glucose was monitored every day at the first 3 days after transplantation,and then monitored every two days.At day 9 after transplantation,spleen,thymus,lymph nods,liver and islets recipient kidney were harvested.Ice slices were prepared and CM-DiI labeled cells were investigated with fluorescence microscope.Results CM-DiI-labeled BMSC were mainly distributed in thymus followed by spleen and lymph nodes.In liver and kidney,there was no red fluorescence observed.The blood sugar control for combined BMSC infusion group was superior to other groups,and the control level of islet combined BMSC infusion group were better than single islet transplantation group in OGTT at day 7.Conclusion Allogeneic BMSC can sustain the insulin secretion of islets in vivo and tend to distribute in immune organs or adenoid tissues after infusion.