Non-Canonical WNT5A Signaling Through RYK Contributes to Aggressive Phenotype of the Rheumatoid Fibroblast-Like Synoviocytes

Angela Rodriguez-Trillo,Fatima Rivas-Tobío,Nerea Mosquera,Antonio Gonzalez,Carmen Pena,Carmen Conde,Antonio Mera-Varela

doi:10.3389/fimmu.2020.555245

Angela Rodriguez-Trillo, Fatima Rivas-Tobío + Show 5 more

Open Access

https://doi.org/10.3389/fimmu.2020.555245

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Abstract

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/Ca2+ 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.

Highlights

Rheumatoid arthritis (RA) is an autoimmune disease characterized by chronic inflammation and progressive destruction of the peripheral joints [1, 2], in which the resident fibroblast-like synoviocytes (FLS) are pivotal
This increase in migration was not observed in the OA FLS, as the healed area with OA FLS was similar with recombinant Wnt5a (rWnt5a) and without treatment (Figure 1C)
Our results indicate that WNT5A contributes to the aggressive phenotype of the FLS in RA patients because it promotes their enhanced migration and invasion, and the expression of inflammatory mediators via the WNT/Ca2+ and RYK/RhoA/ ROCK signaling pathways (Figure 8)

Summary

Introduction

Rheumatoid arthritis (RA) is an autoimmune disease characterized by chronic inflammation and progressive destruction of the peripheral joints [1, 2], in which the resident fibroblast-like synoviocytes (FLS) are pivotal. WNT5A is involved in cytoskeleton remodeling, tissue polarization, cell migration and axon guidance in healthy tissues [11,12,13,14,15,16] and promotes migration and invasion in cancer cells [17,18,19] These actions are mediated through the non-canonical or b-catenin independent WNT pathways, which are the specific pathways transmitting WNT5A signals. Activated Rac and Cdc initiate downstream c-Jun N-terminal kinase (JNK)/activating protein-1 (AP-1) signal transduction, whereas activated RhoA leads to Rho-Kinase (ROCK) activation In this way, the PCP pathway regulates the cytoskeleton remodeling, tissue polarity, coordinated cell migration and axon guidance, and the WNT/ Ca2+ pathway regulates cell migration, fate determination, and axon guidance and cooperates with the PCP pathway in controlling tissue polarity [13, 14, 20]. Dysregulation of the WNT5A signaling has been implicated in the increased migration and invasiveness of cancer cell lines [17,18,19] and the metastatic potential of gastric, breast and nasopharyngeal cancers [19, 25, 26]

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