人胰腺干细胞的体外分离培养、鉴定和分化特性

Isolation, culture and identification of human pancreatic stem cells and their differentiation characteristics in vitro

目的 体外分离培养人胰腺干细胞 ,进行鉴定并初步观察其分化为胰岛内分泌细胞的能力。方法 用酶消化法从人胎儿胰腺分离胰腺干细胞 ,培养并传代 ,应用胰腺干细胞特异性标志物CK 19、胰岛 β细胞特异性成分胰岛素和α细胞特异性成分胰高血糖素的抗体进行免疫细胞化学染色鉴定细胞种类和分化特性。结果 分离培养的人胎儿胰腺干细胞呈梭形 ,最初 1周生长较慢 ,以后增殖较快 ,约培养 3周后即可传代培养。传代细胞约生长 2周即可再次传代 ,免疫细胞化学染色显示细胞呈CK 19免疫阳性反应。经诱导分化后 ,细胞形成类胰岛样组织 ,此时免疫细胞化学染色显示部分细胞呈胰岛素免疫阳性反应 ,少数细胞呈胰高糖素免疫阳性反应。结论 人胰腺干细胞能在体外培养条件下快速增殖 ,并具有多向分化潜能 ,能分化形成胰岛

Objective To isolate, culture and identify the human pancreatic stem cells in vitro and to observe the potency of the cells to differentiate into endocrine cells of pancreatic islet. Methods Stem cells were isolated from the human fetal pancreas by enzyme digestion for transfer of culture in vitro . The potency and types of the cell differentiation were then identified by using the pancreatic stem cell marker cytokeratin 19 (CK 19), the pancreatic β cell marker insulin and the pancreatic α cell marker glucagons with immunocytochemical staining. Results The shape of human pancreatic stem cells cultured in vitro were fusiform. The cells grew slowly in the first week but then proliferated quickly. The cells could be used for subculturing in vitro after 3 weeks. The subcultured cells could be used for subculturing again after another 2 weeks and the cells were found to be of CK 19 immunoreactive positive. After induced differentiation, the cells formed the pancreatic islet like structures. Part of the cells were of insulin immunoreactive positive but few of them were of glucagon immunoreactive positive. Conclusion Human pancreatic stem cells can proliferate quickly, may be of multipotency and can be differentiated into β and α cells of pancreatic islet in vitro.