猪小肠上皮细胞分离培养与鉴定  

Isolation Culture and Identification of Porcine Intestinal Epithelial Cells

作　　者：詹康[1] 左晓昕 陈银银[1] 贡笑笑 占今舜[1] 赵国琦[1] 

机构地区：[1]扬州大学动物科学与技术学院,扬州225009

出　　处：《动物营养学报》2015年第5期1477-1484,共8页CHINESE JOURNAL OF ANIMAL NUTRITION

基　　金：江苏省高校优势学科建设工程资助项目;江苏省科技支撑项目(BE2013387)

摘　　要：为建立功能性永生化的猪小肠上皮细胞系,并为猪肠道营养吸收与免疫调控以及仔猪肠道疾病发病机制的研究提供细胞模型,本试验采用组织块培养法来分离纯化猪小肠上皮细胞,并通过细胞角蛋白18、细胞增殖曲线、核型分析来鉴定猪小肠上皮细胞。结果表明:1)采用组织块培养法能够成功培养猪小肠上皮细胞并稳定传11代。2)本试验获得的猪小肠上皮细胞角蛋白18抗原鉴定为阳性,染色体核型为二倍体。3)倒置显微镜下观察培养至第11代的猪小肠上皮细胞仍然保持着上皮细胞的特征,呈现"铺路石"和上皮样形态。培养11代后的猪小肠上皮细胞间隙开始变大,规则不明显,细胞开始凋亡。第15代的猪小肠上皮细胞则出现大量凋亡并从瓶底脱落,仅有很少细胞贴壁生长。综上所述,采用组织块培养法能够获得生物学功能稳定的猪小肠上皮细胞系并能正常传11代,可为细胞的永生化提供试验素材。This experiment was designed to establish a functional immortalized porcine intestinal epithelial cells line and study porcine intestinal nutrient absorption, immune regulation and the pathogenesis of the neonatal porcine intestinal disease, which provided the cell model. Tissue culture method was used in this experiment to separate and purify the porcine epithelial cells. And the methods of cytokeratine 18 immunofluorescence, cell proliferation curve and karyotype analysis were used to identify the porcine intestinal epithelial cells. The results showed that the porcine intestinal epithelial cells could be successfully cultured and passed 11 generations stably by tissue culture method. 2） The porcine intestinal epithelial cells showed positive reaction against cytokeratine 18 antigen, and the karyotype was diploid. 3） The llth generation cells still have characteristics of epithelial cells, showing ＂cobblestone-shape＂ and epithelial-like morphological features. However, the porcine intestinal epithelial ceils were out of shape and went to the procedure of apoptosis after 11 generations. Most of them went to death in lSth generation and seldom were adherent to the dishes anymore. In summary, the experiment succeed to obtain a porcine intestinal epithelial cells line with stable biological functions by tissue culture meth- od, and they pass 11 generations stably, which provided the experimental materials for immortalized cells.

关 键 词：猪小肠上皮细胞 功能性 肠道营养与免疫 永生化 

分 类 号：Q813.1[生物学—生物工程]