P75NTR/PRONGF UP REGULATION IN SYNOVIAL TISSUES ACTIVATES INFLAMMATORY RESPONSES IN CHRONIC ARTHRITIS PATIENTS

Introduction: We have recently demonstrated that synovial fluids of JIA patients contain elevated levels of proNGF, the immature form of the Nerve Growth Factor (NGF), and not mature NGF as believed in the past. Moreover, mononuclear cells of JIA patients show a reduced expression of TrkA , the specific receptor of mature NGF, and an enhanced expression of p75NTR, the specific receptor for proNGF. This results in an inverted ratio of TrkA and p75NTR in JIA patients compared to healthy donors. How this altered proNGF/p75NTR axis can influence the inflammatory response is at present unknown. Objectives: In this study, we focused on the involvement of p75NTR and its ligand proNGF in regulating inflammatory responses in the inflamed synovia and in synoviocytes of arthritis patients. Methods: Fibroblast-like synoviocytes (FLS) were obtained by synovial tissue of rheumatoid arthritis patients (RA FLS) after enzymatic digestion. Osteoarthritis fibroblasts (OF) and skin fibroblasts (SF) were used as control. Using Realtime-PCR and western blot, we evaluated TrkA, p75NTR, sortilin, NGF mRNA expression and protein levels. Realtime-PCR and ELISA were used to evaluate cytokine production. Results: Our preliminary data show that, similarly to JIA SFMC, p75NTR is significantly upregulated in FLS of RA patients, while TrkA is much less expressed than p75NTR. Instead, OF and SF showed a higher expression of TrkA than p75NTR. Moreover, FLS express NGF mRNA and release high amounts of proNGF, but not of mature NGF, in the conditioned media. Inflammatory stimuli, such as IL-1b, further upregulate p75NTR expression in FLS and induce the expression of p75NTR in control fibroblasts. In vitro, the addition of proNGF to FLS induces the expression of pro-inflammatory cytokines. This effect is abolished when p75NTR activity was inhibited using LM11A-31, a non-peptide ligand that blocks the binding site of p75NTR for proNGF. Conclusion: These preliminary data suggest that the proNGF found in synovial fluids of chronic arthritis patients is produced and released principally by FLS. The accumulated proNGF binds to its specific receptor p75NTR, which is highly expressed by both FLS and SFMC, inducing pro-inflammatory cytokine expression. Inflammatory stimuli further enhances the expression of p75NTR in FLS, creating a vicious circle that amplify the inflammatory response. The use of p75NTR inhibitors might represent a new therapeutic approach for the treatment of JIA and RA.