胆囊Cajal间质细胞的体外分离、培养与鉴定-东南大学学报医学版

胆囊Cajal间质细胞的体外分离、培养与鉴定

单位：东南大学附属中大医院普外科, 江苏南京 210000

分类号：R657.42

出版年·卷·期（页码）：2015·34·第三期（390-393）

摘要：

目的:探讨兔胆囊Cajal间质细胞(ICC)的原代分离、培养及鉴定的方法.方法:选择出生后4~5周龄的新西兰大白兔,经耳缘静脉注入空气处死,在无菌条件下取出胆囊,在解剖显微镜下剥去胆囊黏膜和黏膜下层,利用Ⅱ型胶原酶消化法分离胆囊原代ICC,接种于含有干细胞因子的M199培养基中进行培养.倒置显微镜下连续观察细胞的形态和生长情况,用酪氨酸蛋白激酶受体c-kit特异性抗体免疫荧光染色进行鉴定.结果:细胞稳定贴壁后,在倒置显微镜下观察可见ICC呈梭形、三角形,有突起;随着培养时间的延长,突起细长化并彼此相互连接形成网络.该类细胞c-kit抗体免疫荧光染色呈阳性.结论:成功建立了一套胆囊原代ICC培养的方法,为研究ICC的生物学特性及其与胆囊动力障碍性疾病的关系奠定了基础.

Objective: To investigate the method of isolation, culture and identification of interstitial cells of Cajal (ICC) from the rabbit gallbladder in vitro. Methods: ICC were derived from gallbladder tissue of New Zealand white rabbit by enzymatic dissociation of collagenase type Ⅱ and cultured in M199 medium. The morphological characteristics of ICC was observed with an inverted microscope, and the cell phenotype was identified by immunofluorescence staining using specific antibody against receptor tyrosine kinase c-kit with the confocal microscopy. Results: ICC exhibited a few triangle or spindle shaped axis-cylinders after cells attachment, and the long processes were observed to build synapse and form the net-work structure of cells after 3 days. Cultured ICC were positive for immunofluorescence staining of c-kit antibody. Conclusion: The enzymolysis method for primary culture of ICC from rabbit gallbladder is feasible, which may provide a basis for research into the biological function of ICC and the relationship between the ICC and gallbladder motility disorders.

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