PDX-1腺病毒载体构建及其在脂肪间充质干细胞中的表达

研究通过重组腺病毒表达系统包装重组腺病毒Ad—PDX-1，通过重组腺病毒将Pancreaticduodenalhomeoboxl（PDX-1）导入大鼠脂肪间充质干细胞（ratadiposemesenchymalstemcells，rASCs）中，评价PDX．1在rASCs中的表达情况，为进一步探讨ASCs．PDX-1在体内修复糖尿病的潜能提供了实验基础。研究通过构建pAdEasy-CMV—PDX-1腺病毒质粒，在重组腺病毒表达系统PadEasy-1中成功包装出重组腺病毒Ad-PDX-1。培养及鉴定大鼠脂肪间充质干细胞，将携带PDX-1的重组腺病毒感染rASCs，通过RT-PCR、免疫荧光及免疫印迹（Westernblotting）检测PDX-1在rASCs中的表达及产物定位。结果显示，构建的重组腺病毒包装成功，具有较高滴度，并能成功感染大鼠脂肪间充质干细胞。PDX．1蛋白定位于细胞核，并可稳定表达，为后续将大鼠脂肪间充质干细胞向胰岛B细胞诱导分化奠定了基础，为糖尿病的干细胞治疗提供依据。

Construction PDX-1 modified adenovirus vector and expression in mesenchymal stem

In this study, recombinanted adenovirus containing Pancreatic duodenal homeobox 1 （Ad-PDX-1） was packaged by adeno- virus expressing system. The expression of PDX-1 was evaluated in mesenchymal stem cells derived from rat adipose tissue （rASCs） through Ad-PDX-1 transduction. It provides an experimental basis for diabetes repair treatment using ASCs-PDX-1 in vivo. Recombi- nant Ad-PDX-1 was successfully packaged through the construction of adenovirus plasmid pAdEasy-CMV-PDX-1 by recombinant adeno- virus expression system PadEasy-1. Ratadipose mensenchymal stem cells （rASCs） were cultured successfully and identified. The ex- pression and location of PDX-1 in rASCs were detected by RT-PCR, immunofluorescence and western blotting. The results showed that recombinant adenovirus with higher titer was successfully packaged, rASCs were infected successfully by Ad-PDX-1. The expression of PDX-l could be detected in nucleus of rASCs steadily.