Background/Purpose: The relationship between lung and joint inflammation in rheumatoid arthritis is poorly understood. About 10% of people with rheumatoid arthritis develop interstitial lung disease and people who have early rheumatoid arthritis or anti-citrullinated protein antibodies (ACPAs) without clinical arthritis have increased rates of subclinical lung disease. Also, citrullinated proteins can be found in diseased lungs similar to inflamed joints. Taken together, these observations suggest that one or both of two possible lung/joint relationships may exist. Lung inflammation may be an inciting event in rheumatoid arthritis development possibly by triggering ACPA formation and/or lung inflammation and arthritis may be two manifestations of a similar underlying pathology. However the true mechanism is unknown. Overexpression of TNFα in mice causes both an inflammatory destructive arthritis similar to rheumatoid arthritis as well as severe lung inflammation. We have previously shown that the citrullinating enzyme peptidylarginine deiminase 4 (PAD4) contributes to arthritis in these mice, but its role in lung disease is unknown. Here, we employ mice that overexpress TNFα to address several basic questions about the relationship between TNFα, citrullination, PAD4, arthritis, and lung inflammation.

Methods: Ankle joint and lung tissue from mice that systemically overexpress TNFα (TNF⁺) and wild type littermates as well as TNF⁺PAD4^+/+ and TNF⁺PAD4^-/- littermates were flash frozen and homogenized. Protein lysates were diluted in trichloroacetic acid, incubated with Rh-PG (a citrulline detection probe), quenched, washed, subjected to gel electrophoresis, imaged, stained with Coomassie, and re-imaged to determine the extent of total protein citrullination. Lungs from TNF⁺PAD4^+/+ and TNF⁺PAD4^-/- littermates were fixed, sectioned, stained with hematoxylin and eosin (H&E), and scored in a blinded manner for the severity of several characteristics of lung inflammation. Ankle joints from 22 month old mice that overexpress TNFα only in the lung under the control of the surfactant promoter and littermate controls were fixed, sectioned, stained with H&E, and scored for arthritis severity.

Results: TNF⁺ mice have increased gross protein citrullination in their joints and lungs at 5 months of age, but not 2 or 3.5 months of age compared to wild type. There was no difference between TNF⁺PAD4^+/+ and TNF⁺PAD4^-/- mice in gross protein citrullination in either joints or lungs at 5 months of age, however TNF⁺PAD4^-/- mice had reduced lung disease severity compared to TNF⁺PAD4^+/+ mice. No arthritis was detected in the mice that overexpress TNFα only in the lungs.

Conclusion: PAD4 contributes to both lung and joint inflammation downstream of TNFα, without having a required role in generalized protein citrullination in these tissues. Further, TNFα induced lung inflammation does not drive arthritis in mice.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/tumor-necrosis-factor-alpha-and-peptidylarginine-deiminase-4-in-lung-and-joint-inflammation/