低氧诱导大鼠外周血单核细胞表达TNF-α与PKC膜易位及活性变化关系的研究  

A study of relations among PKC membrane translocation,activity changes and expression of TNF-alpha in rat monocytes exposed to hypoxia

作　　者：徐智[1] 吴国明[1] 钱桂生[1] 王兴胜[1] 陈维中[1] 薛桥[2] 王士雯[2] 

机构地区：[1]第三军医大学新桥医院全军呼吸内科研究所,重庆400037 [2]中国人民解放军301医院老年心血管病研究所,北京100853

出　　处：《重庆医学》2008年第23期2716-2719,共4页Chongqing medicine

基　　金：国家自然科学基金资助项目(30500230)

摘　　要：目的研究低氧诱导大鼠外周血单核细胞（peripheral blood monouclear cells，PBMCs）膜内PKC膜易位及活性变化对肿瘤坏死因子α（TNF-α）表达的作用，探讨缺氧与全身炎症反应综合征（system inflammation response syndrome，SIRS）的关系。方法采用明胶法分离大鼠外周血单核细胞，于低氧条件下（3％O2，5％CO2，92％N2）培养0、1、3、6、9、12、24h后，收集细胞，分别采用免疫沉淀法、SDS-PAGE、PKC活性检测试剂盒及逆转录PCR（RTPCR）检测PKC膜易位、PKC活性及TNF-α表达量，采用SPSS10．0分析以上指标变化的相关性。结果低氧1～9h，大鼠外周血单核细胞膜PKCα含量及活性明显增高（P〈0．01）。低氧1～24h，大鼠外周血单核细胞膜PKCε含量及活性无明显变化。缺氧1～9h，大鼠外周血单核细胞TNF-α mRNA表达明显增高（P〈0．01）。低氧1～24h，PKCα膜蛋白含量及活性变化与TNF-α mRNA表达水平间呈显著正相关（P〈0．01～0．05）。结论低氧可诱导PKCα向膜易位并被激活，活化的PKCα可增强促炎症因子TNF-α的表达。这些变化可能与急性呼吸窘迫综合征（acute respiratory dysfunction，ARDS）时血浆中大量促炎症因子持续存在密切相关。Objective To investigate the role of PKC membrane translocation,activity changes in the expression of TNF-α in rat peripheral blood monouclear cells （PBMCs） exposed to hypoxia,and to explore the relationship between hypoxia and system inflammation response syndrome （SIRS）. Methods Purified rat PBMCs were exposed to hypoxia （3%O2,5% CO2, 92% N2） for 0,1,3, 6,9,12,24h. The PKC membrane translocation was assayed by immuno-precipitation and SDS-PAGE. The PKC activity was assayed by PKC kit. The expression of TNF-α was detected by reverse transcriptase PCR （RT-PCR）. The dependability among above indexes was analysis by SPSS10.0. Results The PKCα membrane translocation and activity increased significantly during 1-9h of hypoxia （P〈0.01）. There were no significant changes of PKCε membrane translocation and activity during 1- 24h of hypoxia. There were significant positive correlations among the PKCα membrane translocation, activities of PKCa and the expression of TNF-α （P〈0. 01-0. 05）. Conclusion Hypoxia enhances the PKCα membrane translocation, the activities of PKCα and up-regulates the expression of TNF-α in mRNA subsequently. These changes might be one of the mechanisms of a mass of pro-inflammatory factors existing persistently in blood plasma in acute respiratory distress syndrome （ARDS） patients.

关 键 词：低氧 PKC 肿瘤坏死因子Α 

分 类 号：R365.73[医药卫生—病理学]