雷公藤甲素对TNF-α诱导的肾系膜细胞MCP-1和ICAM-1表达干预及其机制的研究  

Effects of Triptolide on TNF-α Induced Mesangial Cell MCP-1,ICAM-1 Expression and its Mechanism

作　　者：朱彩凤[1] 朱斌[1] 魏升[1] 陈洪宇[1] 

机构地区：[1]浙江中医药大学附属广兴医院(杭州市中医院)肾内科,杭州310007

出　　处：《中国中西医结合肾病杂志》2011年第6期488-492,共5页Chinese Journal of Integrated Traditional and Western Nephrology

基　　金：浙江省自然基金资助项目(No.Y207829);浙江省中医药科技计划项目(No.2005Y015);国家"十一五"支撑计划项目(No.2006BAI04A07)

摘　　要：目的:观察雷公藤甲素对TNF-α诱导的肾系膜细胞(GMC)MCP-1,ICAM-1表达的干预并探讨其作用机制。方法:把培养的GMC按刺激及干预情况分为正常对照组、TNF-α刺激组、雷公藤甲素低(5ng/ml),中(10ng/ml),高浓度(15ng/ml)干预组,以及吡咯烷二硫代氨基甲酸盐(PDTC)干预组(阳性对照组)、PDTC与雷公藤甲素(10ng/ml)联合干预组。TNF-α刺激干预诱导24h后,分别提取细胞上清液、细胞质、细胞核等成分,采用ELISA法检测MCP-1,ICAM-1,IκB,IκBα,ELISA结合EMSA方法检测各组NF-κB p65,以RT-realtime PCR方法检测MCP-1,ICAM-1的mRNA。结果:TNF-α刺激组MCP-1、ICAM-1的mRNA及蛋白表达、NF-κB p65分子水平较对照组显著增高(P<0.01);各浓度雷公藤甲素或PDTC干预后上述指标显著下降(P<0.01),其中PDTC与雷公藤甲素联合干预组较单用PDTC干预组MCP-1、ICAM-1更低。相关分析表明NF-κB p65与MCP-1及ICAM-1的蛋白和mRNA表达水平呈正相关。GMC经TNF-α刺激后IκB有所下降,雷公藤甲素呈剂量依赖性上调κB,TNF-α刺激后磷酸化的IκBα较正常对照组显著升高(P<0.05),低浓度雷公藤甲素可显著下调其水平(P<0.05)。结论:雷公藤甲素能显著抑制GMC分泌MCP-1、ICAM-1等促炎因子的mRNA及蛋白表达,其机制可能是促进IκB表达上调,并抑制IκBα磷酸化,从而阻断细胞核NF-κB p65活化所致。Objective：To observe the effects of triptolide on TNF-α induced secretion of MCP-1,ICAM-1 and its mechanism in glomerular mesangial cells.Methods：Cells were divided into normal control cells,TNF-α stimulated cells,triptolide （low（5 ng/ml）,median（10 ng/ml） and high（15 ng/ml） concentration） incubated cells stimulated with TNF-α,PDTC intervention cells,cells incubated with both PDTC and triptolide.Supernatant MCP-1,ICAM-1,IκB,IκB α were detected using ELISA method.Cell nucleus NF-κB p65 was detected by ELISA combined with EMSA method.MCP-1 and ICAM-1 and mRNA were measured using RT-realtime-PCR.Results：In the TNF-α stimulated cells,of MCP-1,ICAM-1 Protein and mRNA,NF-κB p65 were significantly increased as compared with the control group （P0.01）;In the triptolide treated cells and PDTC treated cells,MCP-1,ICAM-1 protein and mRNA,NF-κB p65 were decreased significantly（P0.01）.MCP-1,ICAM-1 were decreased more significantly in the cells treated with both triptolide and PDTC as compared with the cells treated with PDTC or triptolide.NF-κB p65 positively correlated to MCP-1 and ICAM-1.IκB was decreased in the TNF-α stimulated cells.Triptolide increased IκB in a dose dependent manner.TNF-α stimulation also significantly increased IκBα phosphorylation （P0.05）.But low concentration of triptolide significantly inhibited the IκBα phosphorylation （P0.05）.Conclusion：Triptolide can upregulate IκB and inhibit IκBα phosphorylation to block NF-κB p65 acitivation,which then repress the secretion of MCP-1,ICAM-1 in TNF-α stimulated GMCs.

关 键 词：雷公藤甲素 MCP-1 ICAM-1 NF-κBp IΚB 

分 类 号：R285.5[医药卫生—中药学]