S-腺苷甲硫氨酸抑制miR-34a依赖的肺癌转移的作用

Role of S-adenosylmethionine in suppressing mir-34a dependent lung cancer metastasis

目的探讨S-腺苷甲硫氨酸抑制miRNA-34a依赖的肺癌转移机制。方法选取肺癌细胞A549,分别采用0、50、100、200、500μM的S-腺苷甲硫氨酸(S-adenosylmethionine,SAMe)刺激A549细胞并检测miR-34a以及下游STAT3信号通路的表达情况,同时检测不同刺激条件下A549细胞的侵袭转移情况;实时荧光定量(real-time fluorescence quantification,qPCR)检测miR-34a、E-cadherin、α-catenin、N-cadherin、Vimentin的mRNA表达水平;Western Blot实验检测STAT3、p-STAT3、β-actin的表达水平;Transwell实验检测A549细胞的侵袭能力变化。结果随着SAMe浓度的增加,miR-34a的表达水平呈梯度增加,与对照组相比,经t检验分析,差异有统计学的意义(P<0.05);Western Blot实验显示,与对照组相比,500μM SAMe刺激后,下游的p-STAT3蛋白水平下降,总STAT3和内参蛋白β-actin水平不变;qPCR结果显示,随着药物浓度的增加,表皮蛋白E-cadherin、α-catenin的mRNA水平上升,间质蛋白N-cadherin、Vimentin的mRNA水平下降;Transwell实验显示经50、100、200、500μM的SAMe刺激后,A549细胞的侵袭转移抑制率分别为18.70%、31.24%、47.66%、58.46%,与对照组相比,经t检验分析,差异有统计学的意义(P<0.05)。结论SAMe可通过抑制miR-34a依赖的下游STAT3信号通路抑制肺癌细胞的侵袭转移,提示SAMe在肺癌侵袭转移过程中发挥重要作用。

Objective To investigate the molecular mechanisms of S-adenosine methionine in suppressing lung cancer metastasis with a miRNA-34a-dependent manner.Methods A549 lung cancer cell was selected as the study model of this time research.A549 cells were stimulated with 0,50,100,200,500μM SAMe,respectively and the expression of miR-34a and its related STAT3 signaling pathways were also detected.The invasive capacity of cells was detected under various condition as well;miR-34a,E-cadherin,α-catenin,N-cadherin and Vimentin mRNA expression were detected by qPCR method;Western Blot was used to detect expression of STAT3,p-STAT3 andβ-actin;Invasive capacity of A549 cells was detected by Transwell experiment.Results As the increase of SAMe concentration,miR-34a expression increased gradually,and when it was compared to the control group,the difference was statistically significant through the t-test(P<0.05);Western Blot experiments showed that p-STAT3 protein level of 500μM SAMe stimulation was lower than 0μM treatment,while total STAT3 andβ-actin level had no change;q-PCR results showed that E-cadherin andα-catenin level elevated,but N-cadherin and Vimentin level decreased as the increase of drug concentration;Transwell experiments showed that invasive inhibitory rates of A549 cells in 50,100,200,500μM of SAMe were 18.70%,31.24%,47.66%and 58.46%,respectively.And compared to the control group,the difference had statistical significance by t-tset(P<0.05).Conclusion SAMe could suppress cell metastasis through a miR-34a-dependent manner by inhibiting STAT3 signaling pathway in lung cancer cells,which suggested a potential role of SAMe in metastasis of lung cancer.