表皮生长因子受体-细胞外信号调节激酶信号通路下调肺癌细胞中微小RNA145的表达

EGFR-ERK signaling pathway down-regulates miRNA-145 in lung cancer cells

目的探讨肺癌细胞中表皮生长因子受体（EGFR）的活化与微小RNAl45（miR-145）下调之间的关系及其分子机制。方法选择正常肺上皮细胞BEAS-2B、EGFR野生型和突变型肺癌细胞A549、H292、H1650和H1975，采用实时荧光定量PCR和Western blot法检测各细胞中miR-145的表达水平和EGFR的活化水平，并分析二者的相关性。分别以EGF、特异性EGFR酪氨酸激酶抑制剂AG1478或ERK1／2抑制剂U0126处理H1975、A549和BEAS^B细胞，观察各细胞中miR-145的表达。结果肺癌细胞中EGFR的活化与miR-145的表达呈显著负相关（r=-0．926，P=0．024）；EGF可下调miR-145的表达，以BEAS-2B和A549细胞内miR-145的表达下调最为明显，分别下调53．0％（t=30．993，P=0．001）和42．6％（t=14．326，P=0．005）；AG1478可恢复EGFR活化诱导miR．145的下调，经AG1478处理后，H1975细胞内miR-145的表达上调67．5％（t=8．269，P=0．014）。EGFR活化后可激活下游信号蛋白分子ERK1／2，U0126可逆转EGFR活化所诱导的miR-145的下调。结论在肺癌细胞中，EGFR通过ERK1／2信号分子下调miR-145的表达。

Objective To investigate the relationship between EGFR activation and down-regulation of miRNA-145 in lung cancer. Methods Normal human lung epithelia cell line （BEAS-2B） , human lung adenocarcinoma cell lines with wild-type EGFR （A549 and H292 ） and human lung adenocarcinoma cell lines with EGFR mutation （H1975 and H1650） were chosen in this study. The levels of miRNA-145 and p- EGFR were determined by quantitative real-time PCR （qRT-PCR） and Western blotting, respectively, and the relationship between p-EGFR and miRNA-145 levels was analyzed. The miRNA-145 levels were determined by qRT-PCR after activating EGFR with EGF or blocking EGFR signal pathway with AG1478. In addition, ERK1/2 inhibitor U0126 was used to inhibit ERKI/2 activation and then the expression of miRNA-145 was detected. Results The miRNA-145 levels were closely negatively related with p-EGFR in lung cancer cells （ r = - 0. 926, P = 0. 024）. EGF down-regulated miRNA-145 expression, particularly in BEAS-2B cells （53.0% ; t = 30.993, P = 0. 001 ） and A549 cells （42.6% ; t = 14. 326, P = 0. 005 ）. The miRNA-145 was up-regulated after inhibiting p-EGFR with AG1478, and significantly enhanced by 67.5% in H1975 cells when treated with AG1478 （ t = 8. 269, P = 0. 014 ）. The ERK1/2 signal pathway was activated by p-EGFR. U0126 restored the miRNA-145 down-regulation induced by EGFR-aetivation in lung cancer cells. Conclusion The activation of EGFR down-regulates miRNA-145 expression through ERK1/2 in lung cancer cells.