【摘要】 目的:建立一种简便易行的小鼠胰腺星状细胞(pancreatic stellate cells,PSCs)分离培养方法,并观察分离培养结果。方法:在原有大鼠胰腺星状细胞分离培养技术基础上,将胰腺组织块碎片进行分离培养并传代,从而得到纯化的星状细胞。采用油红O染色,结合免疫荧光以及蛋白质印迹法鉴定小鼠胰腺星状细胞,通过细胞计数及台盼蓝染色观察细胞生长及活力。结果:组织块法所得小鼠原代胰腺星状细胞油红染色阳性;第3天起细胞快速生长并逐渐活化,传代后的细胞基本活化。活化的PSCs免疫荧光染色显示α-平滑肌肌动蛋白(α-SMA)表达阳性。蛋白质印迹结果表明,活化的PSCs均表达α-SMA、结蛋白,完全活化后结蛋白表达明显增多(P<0.05)。结论:通过组织块法可成功分离出小鼠胰腺星状细胞,其细胞产率、活力和纯度均可满足体外研究需要。更多还原

【Abstract】 Objective:To establish a new method to isolate and culture mouse primary pancreatic stellate cells(PSCs).Methods:Based on techniques of isolating and culturing rat PSCs,we acquired purified PSCs from pancreatic tissue fragments via isolation,culture and passage.Oil red O staining,immunofluorescence as well as western blotting were performed to identify pancreatic stellate cells.Cell growth and vitality were validated by cell counting and trypan blue staining.Results:Primary PSCs were positive for oil red O staining;cells grew rapidly and began being activated from the third day.After passage,most of cells were activated.Immunofluorescence staining showed that α-smooth muscle actin(α-SMA) was positive expressed in activated PSCs.Western blotting showed that α-SMA and desmin were expressed in activated PSCs,and that the expression of desmin in fully activated PSCs was obviously higher(P<0.05).Conclusion:This method can be successfully used to isolate mouse PSCs.The yield,purity and activity of PSCs were higher,and this new method could thus meet the research of pancreatic stellate cells in vitro.更多还原