Monocytes are important players in immunity and their heterogeneity reveals multiple functions. Recent publications suggest that the mouse Ly6Chigh monocyte subset and its human counterpart CD14+/CD16low may represent a valuable target for innovative immunotherapeutic strategies against immune-mediated inflammatory disorders. The present work aims at evaluating such hypothesis in vivo in a preclinical model of rheumatoid arthritis (RA) and in vitro in human CD14+/CD16low primary monocytes. PBEF (Pre-B cell colony-enhancing factor/Visfatin), a pro-inflammatory adipokine described as a new marker of inflammation in RA, is expressed by monocytes and induces IL-6 production. Using RNAi-mediated gene silencing of PBEF, we investigated the therapeutic potential of the targeting of the mouse Ly6Chigh and the human CD14+/CD16low monocyte subsets for immuno-intervention in RA.
Method: Collagen-induced arthritis (CIA) was induced in male DBA/1 mice. We first provided an in-depth characterization of the monocyte subsets in CIA mice using multiparametric analysis by flow cytometry. Small interfering (si)RNAs against mouse PBEF (siPBEF) or non targeting siRNAs (siCT) sequences were formulated with the cationic liposome RPR209120/DOPE and injected intravenously from arthritis onset (0,5 mg/kg). Clinical and biological features of the disease were investigated. In vitro, human CD14+/CD16low monocytes were transfected with human siPBEF or siCT formulated with the cationic liposome. Inhibition of PBEF expression and impact on pro-inflammatory cytokines was quantified.
Result: CIA mice present Ly-6Chigh monocytosis that infiltrate into the arthritic joints and highly express TNFα. The lipoplex formulation used is preferentially uptaken by the Ly-6Chigh monocytes from the blood, spleen and joints of CIA mice and by human CD14+/CD16low monocytes from PBMCs in vitro. Weekly systemic injection of siPBEF lipoplexes significantly decreased clinical disease activity scores compared with siCT treated animals as evidenced by paw swelling measures. The anti-PBEF siRNA lipoplexes efficiently triggered in vivo the inhibition of PBEF protein expression within Ly-6Chigh monocytes/macrophages from blood and inflamed joints. Importantly, PBEF silencing significantly decreased several pro-inflammatory mediators including TNF-α, IL-6, IFN-γ and IL-17A, while increasing anti-inflammatory IL-10 cytokine production in the spleen. Finally, inhibition of PBEF expression in human primary CD14+/CD16low monocytes leads to a decreased IL-6 production.
Conclusion: These results provide novel evidence that silencing of PBEF within Ly-6Chigh monocytes efficiently reduces experimental arthritis, and within CD14+/CD16low monocytes decreases their pro-inflammatory profile, emphasizing the therapeutic potential of Ly-6Chigh / CD14+/CD16low monocytes targeting for anti-inflammatory intervention in RA.
Disclosure: J. Presumey, None; G. Courties, None; V. Escriou, None; D. Scherman, None; D. Kyburz, None; S. Gay, None; Y. M. Pers, None; C. Jorgensen, None; F. Apparailly, None.