猪胎盘绒毛滋养层细胞体外原代培养

发布时间：2018-08-11 11:50

【摘要】：本研究旨在建立一种简便有效的猪胎盘绒毛滋养层细胞体外分离纯化方法。采用胰酶DNase I酶联合消化法消化母猪自然分娩后胎盘绒毛组织,以35%、45%2个Percoll密度梯度法进行猪胎盘绒毛滋养层细胞的分离纯化,并通过观察细胞形态、测定生长曲线、免疫荧光染色、透射电镜等鉴定猪胎盘绒毛滋养层细胞。结果表明,分离培养的猪胎盘绒毛滋养层细胞呈上皮样平铺成片生长,形态为多角形或类圆形,培养48h后部分细胞开始融合成多核合胞体滋养层细胞。免疫荧光染色结果显示,细胞角蛋白7染色阳性占91%以上,表明分离获得的猪胎盘绒毛滋养层细胞纯度较高。透射电镜观察显示,所获细胞具有典型滋养层细胞结构特征。综上表明,采用胰酶DNase I酶联合消化法和35%、45%2个Percoll密度梯度法进行分离纯化,可简便有效地获得较高纯度的猪胎盘绒毛滋养层细胞。
[Abstract]:The aim of this study was to establish a simple and effective method for isolation and purification of porcine placental villus trophoblastic cells in vitro. Placental villi of sows were digested with trypsin DNase I enzyme and digested. Two Percoll density gradient methods were used to isolate and purify placental villus trophoblastic cells from porcine placenta. Cell morphology was observed and growth curve was measured. Porcine placental villus trophoblastic cells were identified by immunofluorescence staining and transmission electron microscopy. The results showed that the cultured porcine placental villus trophoblastic cells grew as epithelioid lamina and were polygonal or round in shape. After 48 hours of culture, some of the cells began to fuse into polynuclear syncytiotrophoblastic cells. The results of immunofluorescence staining showed that cytokeratin 7 was positive in more than 91%, indicating that the purity of the isolated porcine placental villus trophoblastic cells was high. Transmission electron microscopy (TEM) showed that the cells were characterized by typical trophoblast cells. The results showed that high purity of porcine placental villus trophoblastic cells could be obtained by using trypsin DNase I enzyme digestion method and Percoll density gradient method.
【作者单位】：南京农业大学动物科技学院;
【基金】：江苏省自然科学基金-青年基金项目(BK20150672)
【分类号】：S828

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