A549细胞中NOX4介导IL-6自分泌及细胞侵袭的研究

Effect of NOX4 on IL-6 autocrine and cell invasion in A549 cells

目的观察在非小细胞肺癌(NSCLC)细胞A549中NADPH氧化酶4(NOX4)对炎症因子IL-6表达的影响,并研究NOX4、IL-6对A549细胞侵袭过程的作用。方法通过质粒p CMV-NOX4转染细胞,制备NOX4稳定过表达的A549细胞株,采用RT-PCR及Western blotting法对NOX4的表达进行检测;采用ELISA法检测IL-6的分泌水平;采用侵袭小室法检测A549细胞的侵袭能力;采用Western blotting法检测磷酸化Akt表达水平。结果与对照组相比,经质粒p CMV-NOX4转染后,A549细胞中NOX4的mRNA表达和蛋白表达水平均显著升高;NOX4过表达的A549细胞中IL-6表达量增高;NOX4过表达可促进A549细胞的侵袭能力及上调细胞中Akt活性,当采用P6或Siltuximab阻断IL-6/JAK2/STAT3信号时,上述效应被部分逆转。结论 A549细胞中,NOX4过表达能提高IL-6的自分泌水平,IL-6/JAK/STAT3信号参与介导NOX4促进A549细胞侵袭的效应。阻断这两个信号通路可能是针对NSCLC侵袭转移的一个潜在治疗手段。

Objective To study the effect of NOX4 on IL-6 expression in non-small cell lung cancer（ NSCLC）A549 cells and identify the roles of NOX4 and IL-6 on A549 cell invasion. Methods The stable cell line expressing NOX4 was generated by transfection of p CMV-NOX4 into A549 cells,and NOX4 expression was determined by RT-PCR and western blotting. The expression of IL-6 was evaluated by ELISA. The cell invasive ability was observed by the transwell test. The level of phosphorylated Akt was identified by western blotting.Results Compared with the control,the mRNA and protein levels of NOX4 were increased in transfected A549 cells. NOX4 overexpression enhanced IL-6 production in A549 cells,promoted the A549 cell invasion and enhanced Akt phosphorylation. The IL-6 / JAK2 / STAT3 signaling was blocked when treated with P6 or Siltuximab. The NOX4-mediated A549 cell invasion was partially reversed by P6 or Siltuximab treatment.Conclusion In A549 cells,NOX4 overexpression enhances the production of IL-6 and the IL-6 / JAK / STAT3 pathway is involved in NOX4 promoting invasion and metastasis of A549 cells. This study suggests a rationale for a combination of anti-IL-6 / JAK / STAT3 and anti-NOX4 / Akt therapy to suppress the invasion and metastasis of NSCLC.