RANKL对人子宫内膜癌细胞上皮间质转化的影响

RANKL influenced epithelial-mesenchymal transition of human endometrial cancer cell line

目的:探讨RANKL对人子宫内膜癌细胞转移和上皮间质转化的影响。方法:体外构建针对细胞核因子κB受体活化因子(RANK)的过表达质粒载体(p IRES2-3FLAG-EGFP-RANK),脂质体法转染人子宫内膜癌HEC-1A细胞株,形成HEC-1ARANK细胞(过表达质粒组),并与HEC-1A^Control细胞(空白质粒组)相对照。给予1μg/ml可溶性细胞因子sRANKL处理后,细胞划痕试验检测细胞迁移能力,CCK-8试验检测细胞的增殖能力,实时荧光定量PCR(qRT-PCR)和Western blot技术分别检测EMT相关指标E-cadherin、N-cadherin及Vimentin mRNA及蛋白的表达变化。结果:经1μg/ml sRANKL刺激后,HEC-1ARANK细胞迁移能力增强(P<0.01),增殖效应没有明显改变(P>0.05),上皮性标记物E-cadherin mRNA及蛋白的表达水平下降,间质性标记物N-cadherin、Vimentin mRNA及蛋白的表达水平显著上调(P<0.01)。结论:RANKL可通过诱导HEC-1A细胞发生上皮间质转化促进子宫内膜癌的转移。

Objective： To explore the effect of RANKL on the metastasis and epithelial-mesenchymal transition of human endometrial cancer cell line. Methods：The over-expression plasmid targeting the receptor activator of nuclear factor κB （RANK）,pIRES2-3FLAG-EGFP-RANK,was constructed in vitro,and was transfected into HEC-1A cells with lipofectamine tech-nique. The empty plasmid,pIRES2-3FLAG-EGFP-CON236,was established as blank control group. Both HEC-1AControl cells and HEC-1ARANK cells were treated by sRANKL （1μg / ml）,then the migration ability was detected by wound-healing assay. The proliferative ability was detected by CCK-8 assay. The expressions of EMT related markers such as E-cadherin,N-cadherin and Vimentin were detected by qRT-PCR and Western blot. Results：Wound-healing assay showed that the migration potential of HEC-1ARANK cells was significantly increased（ P 〈0. 01） after sRANKL （1μg / ml） treatment. CCK-8 assay indicated that the value-added effect was not obvi-ous change（P〉0. 05）. After cultured by sRANKL （1μg / ml） in HEC-1ARANK cells,the expres-sion of E-cadherin mRNA and protein was significantly reduced,while the expression of N-cad-herin,Vimentin mRNA and protein got more（P〈0. 01）. Conclusion：RANKL can induce EMT in the HEC-1A cells,promoting them to migrate.