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OP0007 CCR2 inhibition abrogates IL-6–induced activation of matrix metalloproteinases in cartilage
  1. A Latourte1,2,
  2. J Maillet1,2,
  3. J Goossens1,
  4. H-K Ea1,2,
  5. M Cohen-Solal1,2,
  6. E Haÿ1,
  7. P Richette1,2
  1. 1Inserm U1132 BIOSCAR, Hôpital Lariboisière
  2. 2Rheumatology Department, Hôpital Lariboisière, University Paris Diderot, Paris, France

Abstract

Background Interleukin 6 (IL-6) plays a crucial role in many rheumatic diseases, including osteoarthritis (OA) [1]. In cartilage, IL-6 activates chondrocyte catabolism by increasing the production of matrix-degrading enzymes, including matrix metalloproteinase 3 (MMP-3) and MMP-13, but it could have other roles.

Objectives We aimed to identify new biological processes regulated by IL-6 in cartilage.

Methods RNA-seq analysis (Illumina HiSeq platform) was used to determine biological pathways associated with IL-6/IL-6R (100 ng/ml) stimulation in mouse primary articular chondrocytes. Results were further validated by qPCR and western blot analysis. The effect of stimulation with CC chemokine ligand 2 (CCL2; 10 ng/ml), CCL7 and CCL8 (100 ng/ml) was investigated in vitro and ex vivo in mouse femoral head cartilage explants. The impact of targeted inhibition of CCL2 or CCL7 by siRNA or blockade of their common receptor CCR2 by a specific antagonist (RS-504393) was determined in IL-6–treated chondrocytes and/or cartilage explants.

Results Transcriptomic analysis revealed overrepresentation of multiple functional clusters of genes in IL-6–stimulated chondrocytes, with strongly increased expression of signalling molecules and especially cytokines. Two of the 10 top genes upregulated by IL-6 were Ccl7 (log2 fold change [FC] 2.33, adjusted p-value [padj] =3.35x10-62) and Ccl2 (log2 FC 1.85, padj =9.10x10-26), which encode for CCR2 ligands. qPCR and western blot validations confirmed these results and revealed that IL-6 stimulation also increased the mRNA level of Ccl8, another CCR2 ligand not identified by RNA-seq analysis. CCL2 and CCL7 but not CCL8 activated extracellular signal-regulated kinase 1/2 and c-Jun N-terminal kinase signalling and increased MMP-3 and MMP-13 production and activation. CCR2 blockade but not the specific inhibition of CCL2 or CCL7 by siRNA, greatly abrogated the IL-6–induced catabolism in vitro and ex vivo.

Conclusions We identified 2 chemokines, CCL2 and CCL7, as key targets of IL-6 in chondrocytes. Although their main role is to mediate monocyte/macrophage recruitment to the joint, their receptor, CCR2, is also strongly involved in IL-6–induced cartilage catabolism. These results suggest a novel mechanism by which CCL2/CCR2 and CCL7/CCR2 signalling could be involved in rheumatic diseases, especially OA [2].

References

  1. Latourte A, Cherifi C, Maillet J, et al. Systemic inhibition of IL-6/Stat3 signalling protects against experimental osteoarthritis. Ann Rheum Dis Published Online First: 27 Oct 2016. doi:10.1136/annrheumdis-2016–209757.

  2. Raghu H, Lepus CM, Wang Q, et al. CCL2/CCR2, but not CCL5/CCR5, mediates monocyte recruitment, inflammation and cartilage destruction in osteoarthritis. Ann Rheum Dis Published Online First: 13 Dec 2016. doi:10.1136/annrheumdis-2016–210426.

References

Disclosure of Interest None declared

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