两类金属蛋白酶在炎性细胞模型中的变化及中药RDQ的天然抑制效应(英文)  

Change in the Activity of MMPs and ADAMs (TACE) in Inflammation Cell Model and Natural Suppressive Effect of Chinese Herb RDQ on These Two Subclasses of Metalloproteinase Family

作　　者：李小鸥[1] 闫媛[2] 王震[2] 王开富[2] 杨渝珍[2] 王宏伟[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院儿科学系,武汉市430030 [2]华中科技大学同济医学院基础医学院生物化学与分子生物学系,武汉市430030

出　　处：《医学分子生物学杂志》2007年第3期189-194,共6页Journal of Medical Molecular Biology

基　　金：国家自然科学基金(No.30371309)~~

摘　　要：目的探讨金属蛋白酶(MPs)家族的MMPs和ADAMs两亚类成分在前体TNF-α(mTNF-α)向成熟型TNF-α(sTNF-α)转化过程所起的作用,MMPs在炎症过程中活性的变化和中药RDQ抗炎保护功效的分子机制,为人工干预炎症过程提供依据和手段。方法以LPS刺激HL-60细胞为天然模型,用细胞学和分子生物学技术在蛋白质和mRNA水平上研究MMPs、ADAM17、TNF-α在炎症过程中的表达和活性变化。结果①LPS刺激后,HL60细胞上清中MMPs的活性随刺激时间延长逐渐减弱,胞浆裂解物中的MMPs则随刺激时间延长活性增加但LPS刺激对MMPs mRNA的表达影响不大;②LPS刺激能使HL-60细胞中ADAM17mRNA和TNF-αmRNA表达在6h同时达高峰;③RDQ能明显降低LPS诱导的ADAM17 mR- NA的增加,并能抑制sTNF-α的分泌。结论①ADAM17对sTNF-α的产生起主导作用;②纯中药注射液热毒清(RDQ)能调节TACE的表达,最终抑制TNF-α的分泌。Objective This study was aimed to investigate the altered activity of two subclasses of metalloproteinases （MPs） family, matrix metalloproteinases （MMPs） and a disintegrin and metalloproteinases （ADAMs）, during the transformation of precursor TNF-α （mTNF-α） to mature TNF- α （sTNF-α） , and to explore the innate inhibitory effect of the Chinese herbal Re Du Qing （RDQ） on inflammation. Methods HL-60 cells were treated in vitro by lipopolysaccharide （LPS） and RDQ. MPs activity was measured on SDS-PAGE substrate gel embedded with gelatin （zymography, Gelatin-PAGE）. The mRNA expression of MMPs （MMlY2, MMP7 and MMP9）, ADAM17 （also called TNF-α converting enzyme, TACE）, and sTNF-α, at various time points after LPS stimulation, was examined by Dot blot hybridization and RT-PCR, respectively. The amount of released sTNF-α was detected by MTT colorimetric assay. RT-PCR was used to assess the inhibitory effect of RDQ on TACE at mRNA level. Result Zymographic analysis revealed that the activity of MMPs in HL-60 cells gradually decreased with the time of LPS stimulation, whereas the alteration of MMPs at mRNA expression level was not significant, as detected by RT-PCR. Dot-blot results indicated that the expression of TACE and TNF-α at mRNA level increased simultaneously. The Chinese herb RDQ displayed obvious inhibitory effect on the cytotoxicity of sTNF-α induced by LPS stimulation at the transcriptional level of TACE gene. Conclusion TACE subclass of MPs family plays a crucial role in controlling the secretion of TNF-α induced by LPS. The Chinese herb RDQ can regulate TACE mRNA expression and inhibit sTNF-α secretion. TACE may serve as a novel target for inflammatory therapy.

关 键 词：肿瘤坏死因子 脂多糖 肿瘤坏死因子转换酶 金属蛋白酶 热毒清 

分 类 号：Q55[生物学—生物化学]