Inflammatory Potential of Four Different Phases of Calcium Pyrophosphate Relies on NF-kB Activation and MAPK Pathways

Laure Campillo-Gimenez (1) , Felix Renaudin (1) , Maud Jalabert (1) , Pierre Gras (2) , Marjolaine Gosset (3) , Christian Rey (2) , Stéphanie Sarda (2) , Corinne Collet (1) , Martine Cohen-Solal (1, 4) , Christèle Combes (2) , Frédéric Lioté (1) , Hang-Korng Ea (1, 4)

1 BIOSCAR (UMR_S_1132 / U1132) - Biologie de l'Os et du Cartilage : Régulations et Ciblages Thérapeutiques
2 CIRIMAT - Centre interuniversitaire de recherche et d'ingenierie des matériaux
3 EA 2496 - Pathologies, Imagerie et Biothérapies oro-faciales
4 Hôpital Lariboisière-Fernand-Widal [APHP]

Background: Calcium pyrophosphate (CPP) microcrystal deposition is associated with wide clinical phenotypes, including acute and chronic arthritis, that are interleukin 1b (IL-1b)-driven. Two CPP microcrystals, namely monoclinic and triclinic CPP dihydrates (m- and t-CPPD), have been identified in human tissues in different proportions according to clinical features. m-CPP tetrahydrate beta (m-CPPTb) and amorphous CPP (a-CPP) phases are considered as m- and t-CPPD crystal precursors in vitro. Objectives: We aimed to decipher the inflammatory properties of the three crystalline phases and one amorphous CPP phase and the intracellular pathways involved. Methods: The four synthesized CPP phases and monosodium urate crystals (MSU, as a control) were used in vitro to stimulate the human monocytic leukemia THP-1 cell line or bone marrow-derived macrophages (BMDM) isolated from WT or NLRP3 KO mice. The gene expression of pro- and anti-inflammatory cytokines was evaluated by quantitative PCR; IL-1b, IL-6 and IL-8 production by ELISA; and mitogen-activated protein kinase (MAPK) activation by immunoblot analysis. NF-kB activation was determined in THP-1 cells containing a reporter plasmid. In vivo, the inflammatory potential of CPP phases was assessed with the murine air pouch model via cell analysis and production of IL-1b and CXCL1 in the exudate. The role of NF-kB was determined by a pharmacological approach, both in vivo and in vitro. Results: In vitro, IL-1b production induced by m- and t-CPPD and m-CPPTb crystals was NLRP3 inflammasome dependent. m-CPPD crystals were the most inflammatory by inducing a faster and higher production and gene expression of IL-1b, IL-6, and IL-8 than t-CPPD, m-CPPTb and MSU crystals. The a-CPP phase did not show an inflammatory property. Accordingly, m-CPPD crystals led to stronger activation of NF-kB, p38, extracellular signal-regulated kinase 1/2 (ERK1/2) and c-Jun N-terminal kinase (JNK) MAPKs. Inhibition of NF-kB completely abrogated IL-1b and IL-8 synthesis and secretion induced by all CPP crystals. Also, inhibition of JNK and ERK1/2 MAPKs decreased both IL-1b secretion and NF-kB activation induced by CPP crystals. In vivo, IL-1b and CXCL1 production and neutrophil infiltration induced by m-CPPD crystals were greatly decreased by NF-kB inhibitor treatment. Conclusion: Our results suggest that the inflammatory potential of different CPP crystals relies on their ability to activate the MAPK-dependent NF-kB pathway. Studies are ongoing to investigate the underlying mechanisms.

Mots clés: CPP crystals; microcrystal-induced arthritis; IL-1beta; MAP kinase signaling; NF-kB pathway; macrophages; NLRP3 inflammasome

Domaines: Biologie cellulaire; Matériaux

Laure Campillo-Gimenez, Felix Renaudin, Maud Jalabert, Pierre Gras, Marjolaine Gosset, et al.. Inflammatory Potential of Four Different Phases of Calcium Pyrophosphate Relies on NF-kB Activation and MAPK Pathways. Frontiers in Immunology, 2018, pp.0. ⟨10.3389/fimmu.2018.02248⟩. ⟨hal-02178914⟩