中国组织工程研究与临床康复2008，Vol.12Issue(29)：5768-5772,5.
胰腺干细胞体外诱导分化为胰岛素分泌细胞及对1型糖尿病模型鼠的治疗作用
Pancreatic stem cells differentiating into insulin-producing cells in vitro and their curative effect on mouse models of type 1 diabetes mellitus
摘要
Abstract
BACKGROUND: Embryonic stem cells or adult stem cells, which are both derived from human or animals, have been proved to have the ability of differentiating into insulin-producing cells, but it is necessary to evaluate their function of secreting insulin and the effect on experimental diabetes.OBJECTIVE: To evaluate the insulin secretion function in vitro by rat pancreatic stem cells after differentiation and their potential application for experimental diabetes therapy in vivo.DESIGN: Randomized control observations.SETTING: Institute of Clinical Medical Sciences in China-Japan Friendship Hospital (Beijing, China).MATERIALS: The experiment was carried out in the Laboratory of Pathophysiology, Institute of Clinical Medical Sciences,China-Japan Friendship Hospital between August 2004 and December 2007. SD male rats and female nude mice were purchased from Beijing Vital River Lab Animal Technology Co., Ltd. All the processes were obeyed by the ethical standard of animals. Nicotinamide and streptozocin were purchased from Sigma Company; anti-nestin monoclonal antibody was purchased from BD Biosciences; radio-immune-detection kit for insulin was purchased from Linco Research Company; anti-insulin antibody and anti-glucagon antibody were purchased from Santa Cruz Company,METHODS: Rat stem cells were isolated and purified from pancreatic duct cells using anti-nestin monoclonal antibody conjugated magnetic beads. These stem cells were induced to proliferate, differentiate and form insulin-producing cells in vitro. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect the expression of insulin mRNA during the cell differentiation. The immunofluorescence staining was used to detect the endocrine cells in islet-like structure induced from stem cells. The radioimmunoassay was used to detect the insulin release from stem cell differentiated islets. The native and stem cell differentiated islets were respectively transplanted under the capsule of left kidney in diabetic nude mice and glycemia was measured to evaluate the function of grafted islets.MAIN OUTCOME MEASURES: Expression of insulin mRNA during the cell differentiation. Analysis of insulin-positive cells and glucagon-positive cells in islet-like structure. Insulin release in stem cell differentiated islets. Glycemia change in diabetic mice after islet transplantation.RESULTS: RT-PCR revealed that, the expression of insulin mRNA was significantly up-regulated in stem cells during differentiation, Immunofluorescence staining indicated that there were lots of insulin-positive cells in the middle of islet-like structure and a few of glucagon-positive cells around border of the islet-like structure. Insulin analysis found that fast phase-release of insulin was absent in the stem cell differentiated islets in response to dextrose stimulation and released insulin was 39% as compared with that in native islets. Glycemia was decreased to 10 mmol/L at the 3rd day in native islet group, and then maintained at a normalized level for 60 days after transplantation; while glycemia was achieved to 10 mmol/L at the 8th day in stem cell differentiated islet group after transplantation. However, glycemia was increased gradually from the 35th day after transplantation in stem cell differentiated islet group.CONCLUSION: The stem cells derived from rat pancreas can be induced to proliferate and differentiate into insulin-producing cells and to form islet-like structure. The stem cell differentiated islet could ameliorate the glycometabolic disorder in diabetic animals.关键词
胰腺干细胞/胰岛素/糖尿病1型/诱导分化/胰岛移植分类
医药卫生引用本文复制引用
蔡寒青,葛焕琦,门秀丽,许世清,娄晋宁..胰腺干细胞体外诱导分化为胰岛素分泌细胞及对1型糖尿病模型鼠的治疗作用[J].中国组织工程研究与临床康复,2008,12(29):5768-5772,5.基金项目
国家"863"基金(2006AA02A113) （2006AA02A113）
北京市科委基金(D307050701350705) （D307050701350705）
吉林省科委基金(200505124) the National High Technology Research and Development Program of China (863 Project), No. 2006AA02A113 （200505124）
a grant by Jilin Provincial Science Committee, No.D0705070135070 5 （）
a grant by Jilin Provincial Science Committee, No. 200505124 （）
