摘要
目的:利用干细胞和非干细胞生长对血清的不同要求,获取大鼠胰岛中所含的干细胞并探讨转分化前后PDX-1的表达差异。方法:将雄性SD大鼠采用胶原酶V型通过胰管对胰腺进行灌注,切取胰腺,消化、离心获得胰岛细胞沉淀物。将细胞悬液首先接种于含10%胎牛血清Ham’sF-10培养液中连续培养36h;然后改用无血清Ham’sF-10培养液培养14d。最后以含KGF、BSA及ITS的无血清DMEM/F12(8mmol/l葡萄糖)培养液连续培养28d。在无血清Ham’sF-10培养液培养第14d以及含KGF、BSA及ITS的无血清DMEM/F12(8mmol/l葡萄糖)培养液培养第28d分别加入双硫腙染色观察三维细胞团,同时取部分贴壁细胞应用免疫细胞化学分析所获得干细胞的胰十二指肠同源框(PDX-1)的表达。结果:经双硫腙染色,转分化培养后的部分细胞呈红色;免疫组化结果显示经转分化培养后,PDX-1在胞核表达更加明显。结论:胰腺干细胞经转分化培养后部分细胞可表现出胰岛细胞的性质;不同纯度胰岛所携带的干细胞经转分化培养后的子代细胞仍然保持干细胞的特征;KGF可能具有促进PDX-1表达的作用。
Objective： To obtain the stem cells in different pancreatic islets purity utilizing free-serum culture medium and investigate the PDX-1 mRNA expression distinction before and after transdifferentiation culture. Methods： Pancreatic islets of male SD rats （islet donors ） were isolated by injection of collagenase into pancreatic ducts, and maintained in serum Ham＇s F-10 for 36h. Then the cells were kept in free-serum Ham＇s F-10 for 14 days.The last stage was to shift culture media from free-serum Ham＇s F-10 to free-serum DMEM/F12 that contained KGF, BSA , glucose and ITS. On the 28d, Dithizon was added into the culture flask for observation of 3D cell. The PDX-1 positive cells was investigated by immunocytochemistry. Results： After transdifferentiation culture, some 3D cells were red. The pdx-1 was remarkable in cell nucleus. Conclusion： After transdifferentiation culture, some pancreatic stem cell display the character of islet cell. The daughter cells of pancreatic stem cells have character of stem cells after transdifferentiation. KGF may have effect on promoting the expression of PDX-1.
出处
《重庆医科大学学报》
CAS
CSCD
北大核心
2009年第1期33-36,共4页
Journal of Chongqing Medical University
基金
重庆市卫生局科研基金资助(项目号:04-2-121)
