OP0076 L-ARGININE REPROGRAMS OSTEOCLAST PURINE METABOLISM AMELIORATING BONE LOSS IN RHEUMATOID ARTHRITIS

Abstract

BackgroundBone erosion is a clinical feature of rheumatoid arthritis related to disease severity and poor functional prognosis. Excessive osteoclast differentiation and insufficient osteoblast function are the main reasons for the erosive process in RA. Our previous investigation indicated that L-arginine supplementation not only diminished arthritic inflammation in the serum-induced arthritis (K/BxN) model but also decreased inflammatory joints osteoclast numbers (1).ObjectivesIn the present study, we aim to investigate the metabolic action of L-arginine supplementation in RA, especially on periarticular bone erosion and systemic bone loss. We plan to depict the metabolic features of TNFα induced inflammatory osteoclasts after in vitro L-arginine supplementation.MethodsThree murine arthritis models (serum-induced arthritis (K/BxN) model, collagen-induced arthritis model, and hTNFtg mice model) were analysed in this study. L-arginine was supplemented within the drinking water after the onset of arthritis. Bone parameters for axial skeleton (spine) and peripheral skeleton (tibia) from the respective group were quantified by μCT. HE and TRAP staining were performed to address further the erosion area and osteoclast numbers in periarticular sites. In vitro osteoclast differentiation was conducted with or without L-arginine treatment, in the presence or not of TNFα activation. Seahorse and SCENITH analyses were adopted to delineate the metabolic features. JC-1 staining and transmission electron microscopy (TEM) were used to depict the mitochondria metabolism. RNA-seq and mass spectrometry (MS) were performed to investigate the underlying molecular mechanism.ResultsInflammation was diminished in all three arthritis models after L-arginine supplementation with a significant reduction in arthritic score. Moreover, an amelioration of periarticular bone erosion, systemic bone loss, and decreased osteoclast numbers in periarticular sites were observed in arthritic mice after L-arginine treatment. L-arginine also inhibited osteoclastogenesis in vitro, particularly under TNFα activation. Seahorse and SCENITH analyses indicated TNFα promoted glycolysis while blocking mitochondria-driven oxidative phosphorylations (OXPHOS) in pre-osteoclasts. Meanwhile, JC-1 staining and TEM images also showed that TNFα decreased mitochondria membrane potential and prompted damage of mitochondria. Surprisingly, L-arginine rescued the TNFα inhibition of OXPHOS while promoting ATP production.RNA-seq and MS data confirmed the boost of OXPHOS after L-arginine treatment under TNFα activation. To interfere with OXPHOS, L-arginine inhibited cJun thus altered arginase-1 and arginase-2 expression. Moreover, the increased ATP in L-arginine treated cells facilitated purine metabolism, especially the production of inosine and hypoxanthine, contributing to the inhibition of osteoclastogenesis. Increasing Adenosine deaminase (ADA) is essential for the production of inosine and hypoxanthine due to the decreased inhibitory regulation of the transcription factor c-Jun.ConclusionThese data strongly demonstrated that L-arginine ameliorates bone erosion in RA through metabolic reprogramming and perturbation of purine metabolism in osteoclasts. L-arginine might therefore benefit RA therapy by reducing joint inflammation and also ameliorating bone destruction.