ASODNs下调Cx43表达对正常子宫内膜间质细胞生长、黏附及侵袭能力的影响

Biological behavior of normal endometrial stromal cells after downregulating Cx43 with ASODNs

目的通过下调正常子宫内膜间质细胞Cx43蛋白表达，了解内膜间质细胞生物学行为的变化。方法取5例手术患者的正常子宫内膜组织，分离、纯化内膜间质细胞，并分组培养。采用反义寡聚核苷酸（ASODNs）＋Lipo共转染正常内膜间质细胞48h；westernblot法检测间质细胞Cx43蛋白表达；荧光漂白后恢复技术（FRAP）定量分析间质细胞的间隙连接细胞间通讯（GJIC）功能；MTT法检测间质细胞的生长及黏附能力；Transwell小室检测间质细胞的侵袭能力。结果ASODNs＋Lipo处理48h后，间质细胞转染率达65％；ASODNs＋Lipo能有效抑制Cx43蛋白表达，且抑制效果可持续48h以上；ASODNs＋Lipo处理48h后，间质细胞GJIC功能被明显抑制；96～120h，ASODNs＋Lipo组间质细胞的吸光度值高于其余3组；ASODNs＋Lipo处理组间质细胞的黏附及侵袭能力均较其余3组增强。结论ASODNs能有效下调子宫内膜间质细胞Cx43蛋白表达并抑制其GJIC功能，导致细胞增殖速度加快，黏附侵袭能力增强。

Objective To investigate the biological behavior of normal endometrial stromal cells attenuated Cx43 protein by ASODNs. Methods Normal endometrial stromal cells were collected from surgical specimens, then separated, purified and cultured in vitro. The stromal cells were co-transfected with ASODNs＋Lipo for 48h. The expression of Cx43 protein was detected by Western blotting. Fluorescence recovery after photobleaching was applied to determine the function of gap junction intercellu- lar communication （GJIC） in stromal cells after downregulating Cx43.The abilities of growth and adhesion were estimated with MTT, and the ability of invasion was evaluated by transwell migration. Results The ratio of stromal cells transfected with A- SODNs＋Lipo was 65%, and Cx43 protein expression was down-regulated for upon to 48h after ASODNs transfected. The GJIC function was attenuated significantly in the normal endometrial stromal cells after transfection with ASODNs＋Lipo for 48h. The transfection of ASODNs＋Lipo enhanced endometrial stromal cell growth after seeded for 96h. The enhanced adhesion and pro- moted invasion were also observed in stromal cells transfected with ASODNs＋Lipo for 48h. Conclusion ASODNs can down-regulate Cx43 protein expression and functional GJIC in normal stromal cells, resulting in a faster growth, enhanced adhe- sion and promoted invasion of stromal cells.