PM2.5通过活化NF-κB途径诱导支气管上皮细胞中血管内皮生长因子的表达

PM2.5 induces VEGF expression via activation of NF-κB pathway in bronchial epithelial cells

目的探讨细颗粒物(particulate matter 2.5,PM2.5)诱导支气管上皮细胞(bronchial epithelial cell,Beas-2B)表达血管内皮生长因子(vascular endothelial growth factor,VEGF)的分子机制。方法 PM2.5粉末溶解于DMEM培养基中,倍比稀释成不同浓度(0、12.5、25、50、100μg/ml)的溶液,超声30 min后加入血清及双抗(链霉素和青霉素),使终浓度为2%血清的体系,刺激人支气管上皮细胞;双荧光素酶报告基因分析系统检测核因子-κB(nuclear factorkappa B,NF-κB)活化水平和vegf基因启动子的转录活化水平;Western印迹检测NF-κB组成亚基p65磷酸化水平、阻遏蛋白IκBα及VEGF蛋白表达水平。结果 PM2.5呈剂量依赖性诱导Beas-2B细胞VEGF表达;PM2.5诱导Beas-2B细胞中NF-κB活化,在浓度为100μg/ml活化强度最高,阻遏蛋白IκBα降解和p65亚基磷酸化水平升高;抑制NF-κB p65亚基表达后Beas-2B细胞VEGF蛋白表达水平下调,vegf基因启动子的转录活化水平下降。结论 PM2.5通过活化NF-κB途径诱导支气管上皮细胞中VEGF表达。

Objective To investigate the molecular mechanism of vascular endothelial growth factor （ VEGF） expression in bronchial epithelial cells （Beas-2B）induced by particulate matter 2.5（PM2.5）.Methods PM2.5 powder was dis-solved in DMEM medium and diluted into five concentrations , 0,12.5,25,50 and 100μg/ml, respectively.The double an-tibiotics （ streptomycin and penicillin ） and FBS were added into the solution to a 2% final concentration of serum system after being treated by ultrasound for 30 minutes.The cultured Beas-2B cells were then treated with different doses of PM2.5.Subsequently, nuclear factor-kappa B（NF-κB） transactivity and the transcriptional activation of vegf gene promot-er were tested by dual-luciferase reporter gene analysis system while phosphorylation of p 65 , expression levels of IκBαand VEGF were detected by Western blotting .Results PM2.5 induced up-regulation of VEGF expression in Beas-2B cells in a dose-dependent manner , accompanied by NF-κB transactivation at the highest level under 100 μg/ml of PM2.5 treatment. Moreover, PM2.5 induced degradation of the repressor protein IκBαand increase in the phosphorylation level of p 65 sub-unit in Beas-2B cells.Knockdown of NF-κB p65 expression significantly inhibited vegf gene promoter transcriptional activa-tion as well as VEGF protein expression in Beas-2B cells induced by PM2.5.Conclusion PM2.5 induces VEGF expres-sion via activation of NF-κB pathway in bronchial epithelial cells .