猪胎盘绒毛滋养层细胞体外原代培养

doi:10.11843/j.issn.0366-6964.2017.01.009

摘要/Abstract

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48 h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:

This research was conducted to establish a simple and effective method for isolation and purification of pig chorionic trophoblast cells. Single cell suspension was prepared from pig fullterm placentas by the method of trypsin-DNaseⅠdigestion and two density (35% and 45%, v/v) Percoll gradient centrifugation.Morphology observation,growth curve measurement, immunofluorescence and transmission electron microscopy (TEM) were applied to cells identification. The results showed that the cells were polygon or quasi-circular in shape, which exhibited epithelioid and paving-like spreading growth. After 48 hours culture, some cells started developing into syncytiotrophoblasts. Immunofluorescence assay showed that the proportion of Cytokeratin 7 positive cells was more than 91%, indicated the high purity of pig chorionic trophoblast cells. The results of TEM revealed that the cells had typical structural features of trophoblasts. These results indicated that our research could be used to obtain high-purity pig chorionic trophoblast cells simply and effectively by the method of trypsin-DNaseⅠdigestion and 2 density (35% and 45%, v/v) Percoll gradient centrifugation.

中图分类号:

S826.2

董书圣，田亮，颜培实. 猪胎盘绒毛滋养层细胞体外原代培养[J]. 畜牧兽医学报, 2017, 48(1): 75-82.

DONG Shu-sheng, TIAN Liang, YAN Pei-shi. Primary Culture of Villous Cytotrophoblasts Isolated from Pig Fullterm Placenta[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2017, 48(1): 75-82.

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https://www.xmsyxb.com/CN/Y2017/V48/I1/75

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