Interleukin-1 Acts via the JNK-2 Signaling Pathway to Induce Aggrecan Degradation by Human Chondrocytes
Interleukin-1 Acts via the JNK-2 Signaling Pathway to Induce Aggrecan Degradation by Human Chondrocytes
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DOI：
10.1002/art.39099
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发表时间：
2015-07-01
影响因子：
13.3
通讯作者：
Saklatvala, Jeremy
中科院分区：
文献类型：
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作者：
Ismail, Heba M.;Yamamoto, Kazuhiro;Saklatvala, Jeremy
Aggrecan使关节软骨能够承受负荷并抵抗压缩。聚集蛋白聚糖损失发生在骨关节炎和类风湿性关节炎的早期，并且可以由炎性细胞因子如白细胞介素-1（IL-1）诱导。IL-1诱导ADAMTS蛋白酶特有的特异性聚集蛋白聚糖的切割。本研究的目的是确定的细胞内信号转导途径，IL-1导致聚集蛋白聚糖降解的人软骨细胞，并探讨如何聚集蛋白聚糖酶的活性是由软骨cytos.MethodsWe开发了一种基于细胞的测定结合小干扰RNA（siRNA）诱导敲低与聚集蛋白聚糖降解测定。在用针对IL-1信号传导途径分子的siRNA转染后，用牛聚集蛋白聚糖覆盖人关节软骨细胞。IL-1刺激后，用AGEG和ARGS新表位抗体检测释放的聚集蛋白聚糖片段。用酶联免疫吸附法测定血清中的蛋白聚糖酶活性和金属蛋白酶组织抑制剂3水平。低密度脂蛋白受体相关蛋白1（LRP-1）脱落进行了分析，通过Western blotting.ResultsADAMTS-5是一个主要的聚集蛋白聚糖酶在人软骨细胞，调节聚集蛋白聚糖降解响应IL-1。肿瘤坏死因子受体相关6（TRAF-6）/转化生长因子活化激酶1（TAK-1）/MKK-4信号轴对于IL-1诱导的聚集蛋白聚糖降解是必不可少的，而NF-B不是。在3种MAPK（ERK、p38和JNK）中，只有JNK-2在聚集蛋白聚糖降解中显示出显著作用。软骨细胞组成性地分泌聚集蛋白聚糖酶，该聚集蛋白聚糖酶被LRP-1持续地内吞，保持聚集蛋白聚糖酶的细胞外水平低。IL-1以JNK-2依赖的方式诱导细胞聚集蛋白聚糖酶的活性，这可能是由于IL-1通过JNK-2依赖的方式诱导LRP-1的脱落，从而减少聚集蛋白聚糖酶的内吞作用。结论TRAF-6/TAK-1/MKK-4/JNK-2信号轴介导IL-1诱导的聚集蛋白聚糖酶分解。聚集蛋白聚糖酶的水平受其内吞作用控制，其可在IL-1刺激后由于LRP-1脱落而降低。
ObjectiveAggrecan enables articular cartilage to bear load and resist compression. Aggrecan loss occurs early in osteoarthritis and rheumatoid arthritis and can be induced by inflammatory cytokines such as interleukin-1 (IL-1). IL-1 induces cleavage of specific aggrecans characteristic of the ADAMTS proteinases. The aim of this study was to identify the intracellular signaling pathways by which IL-1 causes aggrecan degradation by human chondrocytes and to investigate how aggrecanase activity is controlled by chondrocytes.MethodsWe developed a cell-based assay combining small interfering RNA (siRNA)-induced knockdown with aggrecan degradation assays. Human articular chondrocytes were overlaid with bovine aggrecan after transfection with siRNAs against molecules of the IL-1 signaling pathway. After IL-1 stimulation, released aggrecan fragments were detected with AGEG and ARGS neoepitope antibodies. Aggrecanase activity and tissue inhibitor of metalloproteinases 3 levels were measured by enzyme-linked immunosorbent assay. Low-density lipoprotein receptor-related protein 1 (LRP-1) shedding was analyzed by Western blotting.ResultsADAMTS-5 is a major aggrecanase in human chondrocytes, regulating aggrecan degradation in response to IL-1. The tumor necrosis factor receptor-associated 6 (TRAF-6)/transforming growth factor -activated kinase 1 (TAK-1)/MKK-4 signaling axis is essential for IL-1-induced aggrecan degradation, while NF-B is not. Of the 3 MAPKs (ERK, p38, and JNK), only JNK-2 showed a significant role in aggrecan degradation. Chondrocytes constitutively secreted aggrecanase, which was continuously endocytosed by LRP-1, keeping the extracellular level of aggrecanase low. IL-1 induced aggrecanase activity in the medium in a JNK-2-dependent manner, possibly by reducing aggrecanase endocytosis, because IL-1 caused JNK-2-dependent shedding of LRP-1.ConclusionThe signaling axis TRAF-6/TAK-1/MKK-4/JNK-2 mediates IL-1-induced aggrecanolysis. The level of aggrecanase is controlled by its endocytosis, which may be reduced upon IL-1 stimulation because of LRP-1 shedding.