Implication of prostaglandin E2 in TNF-α-induced release of m-calpain from HCS-2/8 chondrocytes.: Inhibition of m-calpain release by NSAIDs
Implication of prostaglandin E2 in TNF-α-induced release of m-calpain from HCS-2/8 chondrocytes.: Inhibition of m-calpain release by NSAIDs
复制标题
DOI：
10.1016/j.joca.2004.08.001
复制
发表时间：
2004-11-01
影响因子：
7
通讯作者：
Shimizu, K
中科院分区：
文献类型：
--
作者：
Fushimi, K;Nakashima, S;Shimizu, K
目的：钙蛋白酶是一类钙离子依赖的细胞内中性半胱氨酸蛋白酶。然而，在关节炎关节的滑液中检测到m-钙蛋白酶，并显示具有体外蛋白聚糖酶活性。在关节炎期间，m-钙蛋白酶释放到细胞外间隙的机制尚未得到很好的表征。在本研究中，我们分析了m-钙蛋白酶释放培养的软骨细胞刺激的促炎细胞因子，肿瘤坏死因子-α（TNF-α）。非甾体抗炎药（NSAID）对m-钙蛋白酶释放的影响也examined.Methods：人软骨细胞HCS-2/8细胞刺激TNF-α的存在或不存在的NSAID。使用抗-m-钙蛋白酶抗体通过Western印迹分析定量细胞和培养基中的m-钙蛋白酶。Western印迹进行光密度分析和带强度determined.Results：TNF-α（10 ng/ml）刺激的m-钙蛋白酶释放与短暂增加细胞m-钙蛋白酶在HCS-2/8细胞。检查的NSAID（阿司匹林、洛索洛芬-SRS、双氯芬酸钠、吲哚美辛和NS 398）抑制10 ng/ml TNF-α诱导的m-钙蛋白酶释放和前列腺素E-2（PGE（2））产生。外源性加入PGE（2）可加速m-钙蛋白酶的释放，以响应较低浓度的TNF-α（1 ng/ml）。AH 6809（一种EP 1/2拮抗剂）有效抑制TNF-α诱导的m-钙蛋白酶释放，而SC 19220（一种EP 1拮抗剂）无效。结论：这些结果表明TNF-α刺激软骨细胞HCS-2/8细胞中m-钙蛋白酶的上调和释放，并且产生的PGE（2）对EP 2-PGE（2）受体的刺激在这个过程中有很深的参与。(C)2004年国际骨关节炎研究学会。由爱思唯尔有限公司出版。保留所有权利。
Objective: Calpains are known as Ca2+-dependent intracellular neutral cysteine proteases. However, m-calpain is detected in synovial fluid of arthritic joints and is shown to possess the proteoglycanase activity in vitro. The mechanism of m-calpain release into the extracellular spaces during arthritis has not yet been well characterized. In the present study, we have analyzed m-calpain release from cultured chondrocytes stimulated by a proinflammatory cytokine, tumor necrosis factor-alpha (TNF-alpha). The effects of non-steroidal anti-inflammatory drugs (NSAIDs) on m-calpain release were also examined.Methods: Human chondrocytic HCS-2/8 cells were stimulated by TNF-alpha in the presence or absence of an NSAID. m-Calpain in the cells and culture medium was quantified by Western blot analysis using an anti-m-calpain antibody. Western blots were subjected to densitometric analysis and band intensities were determined.Results: TNF-alpha (10 ng/ml) stimulated m-calpain release with transient increase in cellular m-calpain in HCS-2/8 cells. NSAIDs examined (aspirin, loxoprofen-SRS, diclofenac sodium, indomethacin and NS398) inhibited m-calpain release and production of prostaglandin E-2 (PGE(2)) induced by 10 ng/ml TNF-alpha. Exogenously added PGE(2) accelerated the release of m-calpain in response to a lower concentration of TNF-alpha (1 ng/ml). AH6809, an EP1/2 antagonist, but not SC19220 (an EP1 antagonist), effectively inhibited TNF-alpha-induced m-calpain release. In contrast, butaprost, an EP2 agonist, accelerated release of m-calpain by 1 ng/ml TNF-alpha.Conclusions: These results suggest that TNF-alpha stimulates upregulation and release of m-calpain in chondrocytic HCS-2/8 cells, and that stimulation of EP2-PGE(2) receptor by produced PGE(2) is deeply involved in this process. (C) 2004 OsteoArthritis Research Society International. Published by Elsevier Ltd. All rights reserved.