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目的模拟体内环境,建立可保持平滑肌细胞与绒毛外细胞滋养层细胞(EVCT)生物学特性的共培养细胞模型,应用于研究平滑肌细胞与滋养细胞理化特性与滋养细胞的侵袭行为。方法利用组织块培养法培养脐动脉平滑肌细胞,组织块培养、胰酶消化和Percoll梯度沉降,收集纯化人早孕绒毛组织的滋养细胞,免疫组化检测细胞的纯度。将滋养细胞与平滑肌细胞分别放入Transwell的上下小室,观察该模型下滋养细胞形态变化、细胞活力、侵袭力改变与分泌功能等特性。结果免疫组化显示EVCT的细胞角蛋白7阳性表达的细胞数占95%以上,SMC a-actin阳性表达的细胞数也超过95%,证实共培养系统中EVCT和SMC纯度均在95%以上,且生物学特性得以维持。上室中的EVCT保持了其侵袭能力,且平滑肌细胞能促进滋养细胞增殖活性、侵蚀能力及MMP2、MMP9的表达。结论成功地建立了平滑肌细胞与滋养细胞原代共培养系统模型,便于研究滋养细胞侵袭和子宫螺旋动脉重铸障碍的分子机制。
Objective To simulate the in vivo environment and establish a co-cultured cell model that can maintain the biological characteristics of smooth muscle cells and villous trophoblast cells (EVCT), and to study the physico-chemical properties of the smooth muscle cells and trophoblast cells and the invasion behavior of trophoblast cells. Methods Umbilical artery smooth muscle cells were cultured with tissue culture method. Tissue culture, trypsinization and Percoll gradient sedimentation were used to collect trophoblast cells. The purity of the cells was detected by immunohistochemistry. The trophoblasts and smooth muscle cells were placed in the upper and lower chambers of Transwell, respectively. The morphological changes of trophoblast cells, changes of cell viability, invasiveness and secretion were observed. Results Immunohistochemistry showed that the number of cytokeratin 7 positive cells in EVCT was more than 95% and the positive cells in SMC a-actin were more than 95%. The purity of EVCT and SMC in the co-culture system was above 95% And biological characteristics are maintained. The upper compartment of EVCT maintained its invasiveness, and smooth muscle cells can promote trophoblast proliferation activity, erosion and MMP2, MMP9 expression. Conclusion The model of primary co-culture system of smooth muscle cells and trophoblast cells has been established successfully and the molecular mechanism of trophoblast invasion and uterine spiral artery recanalization disorder can be easily studied.