Laboratorio de Reumatología Experimental y Observacional, y Servicio de Reumatología, Instituto de Investigación Sanitaria de Santiago (IDIS), Hospital Clinico Universitario de Santiago de Compostela (CHUS), Servizo Galego de Saude (SERGAS), Santiago de Compostela, Spain.
Servicio de Cirugía Ortopédica y Traumatología, Hospital Virxe da Xunqueira, A Coruña, Spain.
Front Immunol. 2020 Oct 15;11:555245. doi: 10.3389/fimmu.2020.555245. eCollection 2020.
We hypothesized that WNT5A could contribute to the enhanced migration and invasiveness of rheumatoid arthritis fibroblast-like synoviocytes (RA FLS), which is one of the incompletely understood aspects of the RA FLS aggressive phenotype. This hypothesis is based on the previous evidence of a WNT5A role in both, RA and cell migration. Migration and invasion of RA FLS were assessed after incubation with recombinant Wnt5a (rWnt5a) or silencing of the endogenous WNT5A expression. The expression of WNT5A, WNT receptors, cytokines, chemokines, and metalloproteinases was quantified with RT-PCR. The WNT pathway was explored with gene silencing, antibody and pharmacological inhibition followed by migration assays and phosphoprotein western blots. Here, we reported that rWnt5a promoted migration and invasion of RA FLS, whereas knockdown of the endogenous WNT5A reduced them. These effects were specific to the RA FLS since they were not observed in FLS from osteoarthritis (OA) patients. Also, rWnt5a induced the expression of IL6, IL8, CCL2, CXCL5, MMP1, MMP3, MMP9, and MMP13 from baseline or potentiating the TNF induction, WNT5A signaling required the RYK receptor and was mediated through the WNT/Ca and the ROCK pathway. These pathways involved the RYK and ROCK dependent activation of the p38, ERK, AKT, and GSK3β kinases, but not the activation of JNK. Together these findings indicate that WNT5A contributes to the enhanced migration and invasiveness of RA FLS through RYK and the specific activation of ROCK and downstream kinases.
我们假设 WNT5A 可以促进类风湿关节炎成纤维样滑膜细胞（RA FLS）的迁移和侵袭，这是 RA FLS 侵袭表型中尚未完全了解的方面之一。这一假设基于 WNT5A 在 RA 和细胞迁移中的作用的先前证据。在用重组 Wnt5a（rWnt5a）孵育或沉默内源性 WNT5A 表达后，评估 RA FLS 的迁移和侵袭。用 RT-PCR 定量检测 WNT5A、WNT 受体、细胞因子、趋化因子和金属蛋白酶的表达。用基因沉默、抗体和药理学抑制来探索 WNT 通路，然后进行迁移测定和磷酸化蛋白 Western 印迹。在这里，我们报告 rWnt5a 促进 RA FLS 的迁移和侵袭，而内源性 WNT5A 的敲低则减少了它们。这些作用是 RA FLS 特有的，因为在骨关节炎（OA）患者的 FLS 中没有观察到这些作用。此外，rWnt5a 诱导基线时或增强 TNF 诱导的 IL6、IL8、CCL2、CXCL5、MMP1、MMP3、MMP9 和 MMP13 的表达，WNT5A 信号需要 RYK 受体，并通过 WNT/Ca 和 ROCK 通路介导。这些途径涉及 RYK 和 ROCK 依赖性激活 p38、ERK、AKT 和 GSK3β 激酶，但不涉及 JNK 的激活。这些发现表明，WNT5A 通过 RYK 并通过 ROCK 和下游激酶的特异性激活，促进 RA FLS 的迁移和侵袭。
