Abstract
Peptidylarginine deiminase 4 (PAD4) contributes to the production of citrullinated proteins as autoantigens for anti-citrullinated protein antibodies (ACPAs) in rheumatoid arthritis (RA). PAD4 can also self-deiminate via autocitrullination. However, the role of this process in RA pathogenesis has not been elucidated. This study aimed to clarify PAD4 function before and after autocitrullination and identify citrullinated PAD4 in the synovial fluid of patients with RA. The autocitrullination of recombinant human PAD4 (rhPAD4) was catalyzed in vitro and determined using anti-modified citrulline immunoblotting. Monocyte chemotaxis was evaluated using Boyden chambers, and citrullinated rhPAD4's ability to induce arthritis was assessed in a C57BL/6J mouse model. Citrullinated PAD4 levels were measured in the synovial fluid of patients with RA and osteoarthritis using a novel enzyme-linked immunosorbent assay. Chemotactic findings showed that citrullinated rhPAD4 recruited monocytes in vitro, whereas unmodified rhPAD4 did not. Compared to unmodified rhPAD4, citrullinated rhPAD4 induced greater inflammation in mouse joints through monocyte migration. More citrullinated PAD4 was found in the synovial fluid of patients with RA than in those with osteoarthritis. Citrullinated PAD4 was even detected in ACPA-negative patients with RA. The autocitrullination of PAD4 amplified inflammatory arthritis through monocyte recruitment, suggesting an ACPA-independent role of PAD4 in RA pathogenesis.