过氧化氢PI3K依赖性促进人支气管上皮细胞血管内皮生长因子表达  

Hydrogen dioxide increases the expression of vascular endothelial growth factor in human bronchiolar epithelium cell via a PI3K-dependent pathway

作　　者：沈湘波[1] 赵海金[1] 蔡绍曦[1] 彭红娟[1] 李文军[2] 佟万成[1] 

机构地区：[1]南方医科大学南方医院呼吸科,广东广州510515 [2]南方医科大学公共卫生与热带医学学院,广东广州510515

出　　处：《南方医科大学学报》2010年第2期228-231,共4页Journal of Southern Medical University

基　　金：国家自然科学基金(30270593);中华医学会慢性呼吸道疾病专项基金(07010130021;08020460124);广东省科技计划基金(2005207007)

摘　　要：目的探讨过氧化氢(H2O2)对人支气管上皮细胞(HBE)VEGF表达的影响及可能的分子调控机制。方法四唑盐(MTT)比色法检测不同H2O2浓度对正常人支气管上皮细胞株HBE135-E6E7活力的影响;实验分4组:正常组,50μmol/L、200μmol/L和600μmol/L,分别处理HBE24h,RT-PCR检测VEGF及β-actin基因表达水平;选择基因表达水平影响较大的50μmol/LH2O2处理细胞24h,双抗体夹心酶联免疫吸附测定(ELISA)检测磷脂酰肌醇3激酶(PI3K)选择性抑制剂Ly294002对H2O2诱导VEGF表达的影响。结果在200μmol/L以下H2O2对细胞活力无显著影响。HBE135-E6E7组成性表达VEGF189和VEGF165。VEGF165 mRNA表达在正常组,50、200和600μmol/L分别为0.379±0.044,0.791±0.042,0.585±0.133,0.720±0.0213VEGF189 mRNA表达在4组中分别为:0.193±0.018,0.270±0.012,0.205±0.074,0.302±0.035。相对于对照组,VEGF165、VEGF189在50μmol/L H2O2处理组即可显著升高VEGF表达(P<0.05),但并无浓度依赖性增加。ELISA结果显示:正常组、50μmol/L H2O2处理组和PI3K预处理上清VEGF浓度分别为:(591.5±9.5)、(768.9±21.3)、(489.3±10.9)pg/ml。H2O2处理组显著升高VEGF表达,而Ly294002可显著降低H2O2诱导的VEGF表达(P<0.05),甚至比正常水平更低(P<0.05)。结论氧化应激可以PI3K通路依赖性地增加HBEVEGF表达,这可能是哮喘慢性气道炎症产生和维持的分子机制之一。Objective To investigate the effect of hydrogen dioxide（H2O2） on the expression of vascular endothelial growth factor（VEGF） in human bronchiolar epithelial（HBE） cells.Methods MTT assay was used to assess HBE cell viability after exposure to different concentrations of H2O2.VEGF/β-actin gene fragments were amplified simultaneously by RT-PCR from the total HBE cell RNA,and VEGF protein expression in the cells was detected using ELISA.Results The exposure to 200 μmol/L H2O2 did not obviously affected the cell viability.Compared with those in the control cell,VEGF165/β-actin and VEGF189/β-actin ratios were significantly increased in the cells after treatment with 50,200,and 600 μmol/L H2O2（P〈0.05）.The protein expression of VEGF significantly increased after 50 μmol/L H2O2 treatment（P〈0.05）,but significantly decreased with pretreatment with the PI3K inhibitor Ly294002（P〉0.05）.Conclusion Oxidative stress increases the expression of VEGF via a PI3K-dependent pathway in human bronchiolar epithelial cells,which may play an important role in the onset and maintenance of chronic inflammation in asthma.

关 键 词：哮喘 氧化应激 血管内皮生长因子 人支气管上皮细胞 PI3K 

分 类 号：R562.2[医药卫生—呼吸系统]