转染miR-544a对肺癌细胞侵袭和转移能力的影响

Impact of the transfection of miR-544a on the invasion and metastasis of lung cancer cells

目的探讨miR-544a对肺癌细胞的侵袭和转移能力的影响及其分子机制。方法用miRNA芯片对非小细胞肺癌(NSCLC)细胞系95C和95D进行miRNA谱系分析,并用荧光定量PCR验证其miR-544a表达水平;用Targetscan软件预测miR-544a的靶基因;Transwell实验观察细胞的侵袭转移能力的变化;western blot验证其表达。结果 miRNA芯片及荧光定量PCR结果显示,与95C细胞(1.00±0.00)比较,miR-544a在95D细胞中表达上调(2.95±0.26,t=12.94,P<0.05);Transwell实验结果显示,95C转染miR-544a mimic后增加(32.00±1.00,q=18.67,P<0.01),95D转染miR-544a inhibitor后,侵袭和转移能力降低(11.00±1.00,q=13.67,P<0.01);Targetscan软件预测E-钙粘连素(CDH1)可能为miR-544a的靶基因;western blot显示,95C转染miR-544a mimic后,CDH1表达下调,vimentin表达上调(0.202±0.017,0.574±0.009,q分别为63.86,9.45,P均<0.01);而95D转染miR-544a inhibitor后,CDH1表达上调,而vimentin表达下调(0.769±0.014,0.320±0.020,q分别为18.67,5.99,P均<0.01)。结论 miR-544a可能通过下调CDH1和上调vimentin的表达来促进肺癌细胞的侵袭和转移。

Objective To investigate the impact of miR-544a on the invasion metastasis of lung cancer cells and its molecular mecha- nism. Methods miRNA expressions of non-small cell lung cancer （NSCLC） cell lines, 95C and 95D, were analyzed by miRNA mi- croarray, and the expression level of miR-544a was further determined by real-time quantitative PCR. The target genes of miR-544a were predicted by the Targetscan software. Then, the invasion and metastasis ability of 95C and 95D cells was detected by the Tran- swell test, and the expression of the target genes by western blot. Results The level of miR-544a expression in 95D cells was signifi- cantly higher than that in 95C cells （2.95 ± 0.26 vs 1.00 ± 0.00, t = 12.94, P 〈 0.05）. After 95C ceils were transfected with miR- 544a mimic, the number of cells was increased significantly （32.00± 1.00 vs 13.33 ±0.01, q = 18.67, P 〈0.01 ）, and the expres- sion of E-cadherin （ CDH1 ） down-regnlated （0.202 ± 0.017 vs 0.841± 0. 052, q = 63.86, P 〈 0.01 ） and that of vimentin up-regula- ted （0. 574± 0.009 vs 0.479± 0.011, q = 9.45, P 〈 0.01 ）. After 95 D cells were transfected with miR-544a inhibitor, the number of cells was decreased obviously （ 11.00 ± 1.00 vs 24.67 ± 0.58, q = 13.67, P 〈 0.01 ）, and the expression of CDH1 up-regulated （0.769 ±0.014 vs 0.583 ± 0.010, q = 18. 67, P 〈 0.01 ） and that of vimentin down-regulated （0. 320 ± 0. 020 vs 0.919 ± 0. 019, q = 5.99, P 〈 0.01 ）. Targetscan predicted that CDH1 may be the target gene of miR-544a. Conclusion miR-544a promotes the in- vasion and metastasis of lung cancer cells probably by the down-regulation of CDH1 and the up-regulation of vimentin.