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作 者:贾延劼[1] 钟乐[1] 宋建辉[1] 罗芳[1] 孙吉平[1] 杨于嘉[1]
出 处:《中国当代儿科杂志》2003年第5期393-397,F004,共6页Chinese Journal of Contemporary Pediatrics
基 金:National Natural Science Foundation (No. 30200128)
摘 要:目的探索大鼠骨髓间质干细胞体外诱导分化为胰岛素分泌细胞的方法。为解决胰岛移植物来源匮乏这一问题提供新的思路。方法 采用横向分化技术,将成年大鼠骨髓间质干细胞诱导成为胰岛素分泌细胞。间接免疫荧光法鉴定诱导前后细胞nestin、胰岛素、胰高血糖素、生长抑素及Pdx-1的表达,RT-PCR法检测诱导前后细胞nestin,胰岛素-1,葡萄糖转运子-2,葡萄糖激酶及其转录因子Isl-1,Pdx-1,Pax-4和Pax-6 mRNA的表达;测定24 h胰岛素分泌量和胰岛素刺激实验评价诱导前后细胞的功能。结果 诱导5 h,nestin阳性细胞为(44.6±7.3)%;诱导24 h,nestin阳性细胞增至(61.8±8.4)%;此后,nestin阳性细胞数目开始下降,诱导第14天后,nestin表达基本消失。诱导后的胰岛素分泌细胞可以表达胰岛素、胰高血糖素、生长抑素和Pdx-1等蛋白;表达胰岛素-1、葡萄糖转运子-2、葡萄糖激酶及其多种转录因子mRNA;胰岛素分泌量增加;胰岛素刺激实验反应敏感等。而诱导前MSCc不县备上述特点。结论 大鼠骨髓间质干细胞体外可以诱导成为胰岛素分泌细胞,为胰岛移植开辟新的研究思路。Objective To explore the protocol that induced marrow mesenchymal stem cells (MSCs) differentiating into islel-secreting cells in vitro and to provide new clues for the sources of islet transplantation. Methods Using a defined culture medium and technique for transdifferentiation, MSCs from adult SI) rats were guided into specific insulin-secreting cells. The expressions of nestin and islet-specific hormones and proteins, such as insulin, glucagon, somatoslatin and pancreatic duodenal homeohox 1 (Pdx-1) were analyzed by indirect immunofluorescence cytochemistry staining before and after induction. The expressions of pancreatic islet cell differentiation-related transcripts, such as neslin, insulin 1, glucose transporter 2 (GLUT 2), Isl-f, Pdx-1, Pax-4 mid Pax-6 were detected by reverse transcription-PCR (RT-PCR). In addition, the quantity of insulin secretion was examined using radioinimuna-issay. Results Five hours after induction, (44.6 ± 7. 3)% of differentiated MSCs expressed nestin and it increased to (61. 8 ± 8. 4)% 24 hs after induction, but the expression of nestin almust disappeared at day 14. In the meantime, islet-like cellular clusters appeared after day 14 and became more apparent by day 28. Differentiated cells were found to be immunoreactive to insulin, glucagon, somalostatin and Pdx-1, and expressed insulin 1, GLUT 2, GK, Isl-1, PDX-1, Pax-4, Pax-6 mRNA. In addition, the results of cumulative quantities of insulin of 24 hs and the stimulation index showed that differentiated cells were able to produce insulin at higher levels, and displayed glucose-dependent insulin release in vitro. Conclusions Adult rat MSCs can be differentiated into insulin-secreting cells in vitro. This approach might lead to widespread cell replacement therapy for Type 1 diabetes.
关 键 词:骨髓间质干细胞 胰岛素分泌细胞 细胞分化 胰岛移植
分 类 号:R329[医药卫生—人体解剖和组织胚胎学]
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