Agere Solomon A, Akhtar Nahid, Watson Jeffery M, Ahmed Salahuddin
Department of Pharmaceutical Sciences, College of Pharmacy, Washington State University, Spokane, WA, United States.
Department of Chemistry and Biochemistry, Gonzaga University, Spokane, WA, United States.
Front Immunol. 2017 Oct 18;8:1341. doi: 10.3389/fimmu.2017.01341. eCollection 2017.
Regulated on activation, normal T expressed, and secreted (RANTES)/CC ligand 5 (CCL5) participates in rheumatoid arthritis (RA) pathogenesis by facilitating leukocyte infiltration, however, its other pathological functions are not fully defined in RA. In the present study, we evaluated the effect of RANTES/CCL5 on tissue degrading enzymes matrix metalloproteinase-1 (MMP-1) and MMP-13 expression and its contribution to the progressive joint damage by RA synovial fibroblasts (RASFs). Our results showed that RANTES/CCL5 dose dependently induced MMP-1 and MMP-13 expression in monolayers and three-dimensional (3D) micromass of human RASFs, which correlated with an increase in collagenase activity. This activation by RANTES/CCL5 was observed in RASF, but not in osteoarthritis SFs (OASFs). Evaluation of the signaling events showed that RANTES/CCL5 selectively activated PKCδ, JNK, and ERK proteins to induce MMP expression in human RASFs. Pretreatment with a functional antagonist (Met-RANTES) or heparinase III [an enzyme that selectively digests heparan sulfate proteoglycans (HSPGs)] completely abrogated RANTES/CCL5-induced MMP-1 and MMP-13 expression. Interestingly, the inhibition of RANTES/CCL5 using small-interfering RNA approach reduced the ability of interleukin-1β (IL-1β) to induce MMP-1 and MMP-13 expression, asserting its mediatory role in tissue remodeling. In the inhibitor study, only the selective inhibition of HSPGs or PKCδ, ERK, and JNK markedly inhibited RANTES/CCL5-induced MMP-1 and MMP-13 production. Circular dichroism spectroscopy results demonstrated the degradation of collagen triple-helical structure upon exposure to the conditioned media from RANTES/CCL5 stimulated RASFs, which was reverted by a broad-spectrum MMP inhibitor (GM6001). These findings suggest that RANTES/CCL5 not only upregulates MMP-1 and MMP-13 expression by partly utilizing HSPGs and/or PKCδ-JNK/ERK pathways but also mediates IL-1β-induced MMP-1 and MMP-13 expression.
调节激活正常T细胞表达和分泌因子（RANTES）/CC趋化因子配体5（CCL5）通过促进白细胞浸润参与类风湿关节炎（RA）的发病机制，然而，其在RA中的其他病理功能尚未完全明确。在本研究中，我们评估了RANTES/CCL5对组织降解酶基质金属蛋白酶-1（MMP-1）和MMP-13表达的影响及其对RA滑膜成纤维细胞（RASFs）所致进行性关节损伤的作用。我们的结果表明，RANTES/CCL5在人RASFs的单层和三维（3D）微团中剂量依赖性地诱导MMP-1和MMP-13表达，这与胶原酶活性增加相关。RANTES/CCL5的这种激活作用在RASF中观察到，但在骨关节炎滑膜成纤维细胞（OASFs）中未观察到。对信号转导事件的评估表明，RANTES/CCL5选择性激活PKCδ、JNK和ERK蛋白以诱导人RASFs中MMP的表达。用功能性拮抗剂（Met-RANTES）或肝素酶III（一种选择性消化硫酸乙酰肝素蛋白聚糖（HSPGs）的酶）预处理可完全消除RANTES/CCL5诱导的MMP-1和MMP-13表达。有趣的是，使用小干扰RNA方法抑制RANTES/CCL5降低了白细胞介素-1β（IL-1β）诱导MMP-1和MMP-13表达的能力，证实了其在组织重塑中的介导作用。在抑制剂研究中，仅选择性抑制HSPGs或PKCδ、ERK和JNK可显著抑制RANTES/CCL5诱导的MMP-1和MMP-13产生。圆二色光谱结果表明，暴露于RANTES/CCL5刺激的RASFs的条件培养基后，胶原三螺旋结构发生降解，而广谱MMP抑制剂（GM6001）可使其恢复。这些发现表明，RANTES/CCL5不仅通过部分利用HSPGs和/或PKCδ-JNK/ERK途径上调MMP-1和MMP-13表达，还介导IL-1β诱导的MMP-1和MMP-13表达。
