山萘酚通过下调ERRα抑制非小细胞肺癌A549细胞的侵袭和迁移

Kaempferol inhibits invasion and migration of non-small cell lung cancer A549 cells by down-regulating ERRα expression

目的:探讨山奈酚(kaempferol)对非小细胞肺癌(non-small cell lung cancer, NSCLC)A549细胞侵袭和迁移的影响及其作用机制。方法:A549细胞培养完成后,CCK-8法检测不同浓度山奈酚对A549细胞增殖的影响,Transwell和划痕实验检测A549细胞侵袭和迁移能力,Western blotting检测EMT相关蛋白的表达,qRT-PCR和Western blotting检测山奈酚对雌激素相关受体α(estrogen related receptor alpha, ERRα)mRNA和蛋白表达的影响。构建ERRα过表达载体pLV-ERRα转染A549细胞后,Transwell实验、划痕愈合实验和Western blotting检测A549细胞侵袭、迁移和EMT相关蛋白的表达情况。结果:山奈酚浓度依赖性抑制A549细胞增殖,选择5、10和20μmol/L山奈酚进行后续实验。经山奈酚处理后,A549细胞侵袭细胞数目显著减少、划痕愈合率显著降低、N-cadherin和Snail-2的表达显著下调,E-cadherin的表达显著上调,ERRa mRNA和蛋白水平显著降低(均P<0.01)。转染pLV-ERRα后,过表达ERRα的A549细胞的侵袭细胞数、划痕愈合率均显著增加,细胞中N-cadherin、Snail-2表达上调,而E-cadherin的表达下调(均P<0.05或P<0.01);该细胞再经山奈酚处理后,上述各指标均发生逆转变化(均P<0.05或P<0.01)。结论:山奈酚通过抑制ERRα降低NSCLC A549细胞侵袭和迁移能力,并抑制其EMT,为肺癌的临床治疗提供了实验依据。

Objective: To study the effects of kaempferol on invasion and migration of non-small cell lung cancer(NSCLC) A549 cells and the related mechanisms. Methods: CCK-8 was used to detect the effect of different concentrations of kaempferol on the proliferation of A549 cells. Transwell assay and wound healing assay were used to detect the ability of cell invasion and migration. The expressions of EMT-related proteins(N-cadherin, Snail-2 and E-cadherin) were detected by Western blotting. The effect of kaempferol on the mRNA and protein expressions of estrogen related receptor alpha(ERRα) was determined by qRT-PCR. ERRα over-expression vector(pLV-ERRα) was transfected into A549 cells, and the cell invasion, migration and the expression of EMT related proteins were detected by Transwell assay, wound healing assay and Western blotting, respectively. Results: Kaempferol dose-dependently inhibited the proliferation of A549 cells, and kaempferol at concentrations of 5, 10 and 20 μmol/L was used in the following experiments. After the treatment with different concentrations of kaempferol, the number of invasive cells and wound closure rate, the expression of N-cadherin and Snail-2, and the mRNA and protein levels of ERRα decreased significantly, while the expression of E-cadherin increased significantly,(all P<0.01). After transfection with pLV-ERRα, the number of invasive cells, scratch closure rate and the expressions of N-cadherin and Snail-2 of A549 cells over-expressing ERRα significantly increased, while the expression of E-cadherin decreased significantly(all P<0.05 or P<0.01); and all these indices were attenuated after the treatment with kaempferol(all P<0.05 or P<0.01). Conclusion: Kaempferol inhibits the invasion and migration as well as EMT of non-small cell lung cancer A549 cells by down-regulating ERRα, which may provide experimental basis for the clinical treatment of lung cancer.