摘要
目的:探讨桔皮素(TGN)对非小细胞肺癌(NSCLC)细胞生长和侵袭的影响及其分子机制。方法:体外培养非小细胞肺癌A549细胞,分别用不同浓度的TGN处理,MTT比色法检测细胞活性,Annexin V-FITC/PI染色及流式细胞术检测细胞凋亡率,Transwell检测细胞侵袭,RT-PCR分析MMP-2和MMP-9的mRNA表达水平,Western blotting检测Ki67、Cyt C、caspase-3、cleaved caspase-3、MMP-2、MMP-9、Akt、p-Akt以及p-PI3K表达水平。结果:桔皮素剂量依赖性地抑制A549细胞增殖(P<0.05),同时伴随有增殖标记分子Ki67表达水平的下调。分析发现,桔皮素诱导细胞中Cyt C、caspase-3和cleaved caspase-3的表达上调(P<0.01),加速A549细胞的凋亡。此外,桔皮素作用后,A549细胞中侵袭相关蛋白MMP-2和MMP-9的表达量下降,且侵袭数目随桔皮素浓度增加而减少。进一步研究表明,桔皮素作用后A549细胞中p-Akt和p-PI3K表达水平降低(P<0.05),阻断PI3K/Akt信号通路后,不同浓度TGN对细胞活性影响没有变化。结论:桔皮素能抑制A549细胞生长及侵袭,促进细胞凋亡,可能通过抑制PI3K/Akt信号通路的激活起作用。因此,本研究将为非小细胞肺癌的防治提供新的研究方向。
AIM： To study the influences of tangeretin （ TGN） on the growth and invasion of non-small-cell lung cancer （ NSCLC） cells, and to explore the molecular mechanisms.METHODS： The A549 cells were treated with different concentrations of TGN in vitro.The relative cell activity was determined by MTT assay.The apoptotic rate was an-alyzed by flow cytometry with Annexin V-FITC/PI staining.The number of the invasive cells was measured by Transwell as-say.The mRNA expression of MMP-2 and MMP-9 was detected by RT-PCR, and the protein levels of Ki67, Cyt C, caspase-3, cleaved caspase-3, MMP-2, MMP-9, Akt, p-Akt and p-PI3K were determined by Western blotting analysis. RESULTS：TGN inhibited the proliferation of A549 cells in a dose-dependent manner （ P〈0.05） along with the low ex-pression level of proliferation biomarker Ki67.TGN up-regulated the protein levels of Cyt C, caspase-3 and cleaved caspase-3 （P〈0.01） and promoted the apoptosis of A549 cells in a dose-dependent manner.Moreover, TGN down-regu-lated the invasion-related molecules MMP-2 and MMP-9 at the mRNA and protein levels, and the number of invasive cells reduced with the increase in the concentration of TGN.The protein levels of p-Akt and p-PI3K in the A549 cells was re-duced （P〈0.05）, and no difference of the cell viability in the cells treated with different concentrations of TGN was ob-served after blocking PI3K/Akt signaling pathway using LY294002.CONCLUSION： TGN inhibits the growth and inva-sion of A549 cells and promotes the cell apoptosis by potentially inhibiting PI3K/Akt signaling pathway activation.There-fore, this study will provide a new target for the prevention and control of NSCLC.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2015年第3期452-456,共5页
Chinese Journal of Pathophysiology
