人胰腺干细胞诱导胰岛样细胞团及其治疗大鼠糖尿病的效果  

Differentiation of human pancreatic stem cells into islet-like cell clusters in the treatment of diabetic rats

作　　者：赵婷[1] 乔海[1] 王赟[1] 贾文文[1] 窦琳[1] 效梅[2] 窦忠英[1] 

机构地区：[1]西北农林科技大学干细胞研究中心,陕西省杨凌示范区712100 [2]广东海洋大学,广东省湛江市524088

出　　处：《中国组织工程研究与临床康复》2007年第7期1259-1262,I0003,共5页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：国家"八六三"计划资助项目(2005AA219050);教育部重大项目(03160);陕西省科技研究发展计划项目(2006KZ05G1)~~

摘　　要：目的:观察人胎儿胰腺干细体外诱导分化为胰岛样细胞团及治疗大鼠糖尿病的效果。方法:实验于2005-09/2006-09在西北农林科技大学陕西省干细胞工程技术研究中心完成。人胰腺干细胞为本实验室冻存的1株传至50代的人胎儿胰腺干细胞。采用DMEM/F12培养液,添加B27、1g/L胰岛素-转铁蛋白-亚硒酸钠、1g/L牛血清白蛋白、10mmol/L烟酰胺、10μg/L碱性成纤维细胞生长因子及10μg/L肝细胞生长因子,诱导胰腺干细胞分化为胰岛样细胞团,应用双硫腙染色、RT-PCR及葡萄糖刺激试验对该胰岛样细胞团进行鉴定。采用完全随机法将30只SD大鼠分为链脲菌素注射组24只,正常对照组6只。空腹12h后,两组大鼠分别腹腔注射70mg/kg体质量的链脲菌素和0.1mol/L柠檬酸-柠檬酸钠缓冲液。大鼠分别于注射前、注射后48h、第5天和第8天空腹断尾采血,测定全血血糖浓度。选取连续2次血糖浓度高于16.7mmol/L的大鼠作为糖尿病模型。采用完全随机法从已成模的糖尿病大鼠中挑出18只分为3组:胰岛移植组9只,进行肾被膜下胰岛样细胞团移植,每只大鼠植入胰岛数(3000±100)个;干细胞移植组3只,肾被膜下移植未进行诱导的胰腺干细胞,细胞数约2×106个;糖尿病对照组6只,肾被膜下移植不含细胞的培养液。3组大鼠均于移植前及移植后48h测定空腹血糖浓度,每周定点测空腹血糖及称体质量1次。结果:18只符合糖尿病模型标准的大鼠与正常对照组6只大鼠均进入结果分析。①成功将人胎儿胰腺干细胞体外诱导分化为胰岛样细胞团,双硫腙染色阳性,RT-PCR检测表达胰岛素,经葡萄糖刺激能释放胰岛素。②共有22只大鼠糖尿病成模,成模率达91.7%。③胰岛移植组大鼠移植后2周内,血糖浓度均有所降低,但仍高于正常血糖浓度范围;移植后第3周血糖浓度迅速上升,之后一直维持高血糖状态。干细胞移植组大鼠移植后血糖一直处�AIM： To induce human fetal pancreatic stem cells to differentiate into islet-like cell dusters and observe the effects in the treatment of diabetic rats by transplantaticn. METHODS： The experiment was conducted in Shaanxi Branch of National Stem Cell Engineering ＆ Technology Center, Northwest A ＆ F University from September 2005 to September 2006. Human fetal pancreatic stem cells were induced in DMEM/F12, B27, 1 g/L ITS, 1 g/L bovine serum albumin （BSA）, 10 mmol/L Nicotamide, 10 μg/L basic fibroblast growth factor （bFGF） and 10ng/mL hepatocyte growth factor （HGF） to differentiate into islet-like cell dusters. The islet-like cell dusters were identified by dithizone and RT-PCR, and measurement of insulin was conducted. Thirty rats were randomly divided into streptozotocin （STZ） group （n =24）, and normal control group （n =6）. Rats in both groups were intraperitoneally injected with STZ （70mg/kg of body weight） after 12-hour fasting, while rats in the normal control group were intraperitoneally injected with 0.1 mol/L solution of citric acid and sodium citrate （70 mg/kg of body weight）. Rats were cut off the tails to obtain fasting blood before injection, 48 hours after injection, 5 days after injection and 8 days after injection respectively to determine the level of glucose in whole blood. Rats with the level of blood glucose hiher than 16.7 mmol/L for two continuous times were taken as the models of diabetes. Eighteen diabetic rats were selected and randomly divided into 3 groups： Islet transplantation group （n =9）, rats in which group were implanted with islet-like cell dusters with （3 000±100） in each rat. Stem cell transplantation group （n =3）, rats were implanted with pancreatic stem cells （2×10^6）. Diabetic control group （n =6）, rats ware implanted with pancreatic stem cells without culture fuid of cells. The level of fasting blood glucose of rats in 3 groups were determined before transplantation and at 48 hours after the transplantation, and

关 键 词：胰腺 干细胞 移植 糖尿病 大鼠 

分 类 号：R394.2[医药卫生—医学遗传学]