NF-κB在马钱子碱抑制非小细胞肺癌细胞环氧化酶2生成中的作用研究

Effect of NF-κB on inhibition of non-small cell lung cancer cell cyclooxygenase-2 by brucine

目的:探讨马钱子碱抑制环氧化酶2(COX-2),从而诱导非小细胞肺癌细胞凋亡的分子机制。方法:构建COX-2启动子若干转录因子缺失突变体与含COX-2 mRNA 3'-UTR的报告质粒,与Renillia共转染至非小细胞肺癌A549细胞,测报告基因luciferase活性研究COX-2启动子受马钱子碱抑制的最小顺式作用元件;采用蛋白质免疫印迹法研究马钱子碱对IκBα磷酸化与p65进核的影响。结果:马钱子碱显著性抑制LPS诱导的COX-2启动子的激活,而对COX-2 mRNA转录后调控影响不大,COX-2启动子-262位附近NF-κB是马钱子碱抑制COX-2启动子活性的重要顺式作用元件。马钱子碱能抑制IκBα的磷酸化,并能抑制p65的进核。结论:马钱子碱抑制NF-κB的激活,进而从转录水平COX-2的基因表达,促进A549细胞凋亡。

Objective： To study the molecular mechanism of cyclooxygenase-2（COX-2）,one of effective ingredient of brucine,in inducing non-small cell lung cancer cell apoptosis.Method： COX-2 promoter,transcription factor deletion mutants and COX-2 mRNA 3′-UTR-containing report plasmids were transfected with Renillia to non-small cell lung cancer A549 cell,in order to detect the activity of report gene luciferase and minimum cis-acting element of COX-2 promoter inhibited by brucine.The influence of brucine on IκB phosphorylation and the nuclear translocation of p65 were detected by immunoblotting assay.Result： Brucine significantly suppressed LPS-induced COX-2 promoter activation,but revealed minor impact on COX-2 mRNA stability.NF-κB in the vicinity of COX-2 promoter-262 was an important cis-acting element of brucine for inhibiting the activity of COX-2 promoter.Brucine was found to inhibit the phosphorylation of IκBα as well as the nuclear translocation of p65.Conclusion： Brucine can improve A549 cells apoptosis by inhibiting the activity of NF-κB and the subsequent COX-2 gene expression.