小鼠胰腺干细胞的分离、培养与鉴定

Isolation,culture and identification of pancreatic stem cells from newborn mouse

目的从新生小鼠中分离培养并鉴定胰腺干细胞。方法利用组织块法分离新生小鼠胰腺源性干细胞,经RPMI1640+5%胎牛血清+双抗+1 mmol/L丙酮酸钠+1 mmol/L非必需氨基酸+10 ng/mL表皮生长因子+10 ng/mL成纤维生长因子+2 mmol/L谷氨酰胺的溶液培养,利用免疫组化分析胰腺源性干细胞的兔抗巢蛋白抗体(Nestin)、胰肠同源域因子(PDX-1)和葡葡糖转运子(GLUT-2)抗原表达;经10 mmol/L尼克酰胺诱导产生胰岛样细胞团并利用双硫腙(DTZ)染色分析其胰岛素分泌情况。结果分离培养的胰腺源性干细胞呈Nestin、PDX-1和GLUT-2阳性;诱导的胰岛样细胞团呈DTZ染色阳性。结论分离获得的新生小鼠胰腺源性干细胞在体外可分化为类胰岛,并具有胰岛素分泌功能。

Objective To separate, culture and identify the pancreatic stem cells （PSCs） from newborn mouse. Methods The pancreas-derived stem cells were separated from newborn mouse by tissue block method and euhured in the liquid medium （RPMI1640 ＋ 5% FBS ＋ double-antibiotics ＋ 1 mmol/L sodium pyrnvate ＋ 1 mmol/L Non-essential amino acid ＋ 10 ng/mL EGF ＋ 10 ng/mL bFGF ＋ 2 mmol/L Glutamine）. The expressions of Nestin, PDX-1 and GLUT-2 anti- gens were identified and analyzed by immunohistoehemistry. The pancreas-derived stem cells were induced by 10 mmol/L NIC and differentiated into panereatie islet-like eell clusters, which was stained with DTZ for analyzing the seeretion of in- sulin. Results In the isolated and cultured pancreas-derived stem cells, Nestin, PDX-1 and GLUT-2 expressed positive ly, in the induced panereatie islet-like cell elusters, DTZ expressed positively. Conclusions The pancreas-derived stem cells isolated from newborn mouse are PSCs, which can differentiate into similar islets of pancreas, and have islets of pan- ereas function.