Tumor necrosis factor alpha, citrullination, and peptidylarginine deiminase 4 in lung and joint inflammation

Abstract

BackgroundThe relationship between lung and joint inflammation in rheumatoid arthritis is poorly understood. Lung inflammation with resultant protein citrullination may trigger anti-citrullinated protein antibodies, inflammation, and arthritis. Alternatively, lung and joint inflammation may be two manifestations of a single underlying pathology. The lung has increased citrullination and TNF-α levels are high in rheumatoid arthritis; however, it is unknown if TNF-α can induce lung protein citrullination. The citrullinating enzyme peptidylarginine deiminase 4 (PAD4) exacerbates TNF-α-induced arthritis, but a role for PAD4 in lung citrullination and TNF-α-induced lung inflammation has not been explored. Our aim was to use TNF-α-overexpressing mice to clarify the intersection of TNF-α, citrullination, PAD4, arthritis, and lung inflammation.MethodsLung protein citrullination in wild-type mice, mice that overexpress TNF-α systemically (TNF+), TNF+PAD4+/+, and TNF+PAD4-/- mice was quantified by both gel electrophoresis using a citrulline probe and western blot. Hematoxylin and eosin (H&E)-stained lung sections from TNF+PAD4+/+ and TNF+PAD4-/- mice were scored for lung inflammation. H&E-stained ankle joint sections from mice that overexpress TNF-α only in the lungs were assessed for arthritis.ResultsTNF+ mice have increased lung protein citrullination. TNF+PAD4-/- mice do not have significantly reduced lung protein citrullination, but do have decreased lung inflammation compared to TNF+PAD4+/+ mice. Mice that overexpress TNF-α only in the lungs do not develop arthritis.ConclusionsPAD4 exacerbates lung inflammation downstream of TNF-α without having a major role in generalized protein citrullination in inflamed lungs. Also, TNF-α-induced lung inflammation is not sufficient to drive murine arthritis.Electronic supplementary materialThe online version of this article (doi:10.1186/s13075-016-1068-0) contains supplementary material, which is available to authorized users.

Peptidylarginine deiminase 2 is required for tumor necrosis factor alpha-induced citrullination and arthritis, but not neutrophil extracellular trap formation

“…This method likely does not detect every citrullinated protein, but use of the citrulline probe rhodamine-phenylglyoxal (Rh-PG) [47] is limited by extensive background and use of the mouse monoclonal anti-peptidylcitrulline antibody F95 [48] is limited by binding of the anti-mouse IgM secondary antibody in murine joint tissue (data not shown). Nonetheless, our finding is consistent with the observation that PAD4 is not required for TNFα-induced lung [31] or serum [25] protein citrullination, as determined by other methods. Thus, we conclude that PAD4 is not required for at least a significant portion of the increased citrullination seen in TNFα-induced arthritis.…”

“…As expected, we found that TNFα overexpression leads to increased citrullination in inflamed joints consistent with the finding that TNFα drives citrullination in the serum [25] and lung [31]. TNFα may also drive citrullination in the human rheumatoid joint providing fodder for continued inflammation particularly in ACPA + individuals.…”

“…Together, these findings suggest that PAD4 may contribute to the increased citrullination seen in arthritic joints. However, no reduction in serum [25] or lung [31] protein citrullination is seen in TNFα-induced arthritis when PAD4 is absent.…”

Mining the Human Tissue Proteome for Protein Citrullination

“…Previous studies have shown that (chronic) inflammation induced by smoking and environmental stimuli such as infection can increase the levels of inflammatory cytokines such as TNF-α in the lung and trigger protein citrullination (44,45). The notion that citrullination is an inflammation-dependent process (46) may be substantiated by our observation that the third highest levels of citrullination was found in placenta, a tissue with strong systemic inflammatory responses during normal pregnancy and acute inflammation during labor (47).…”

Development of Activity-Based Proteomic Probes for Protein Citrullination

“…4a). Using Rh-PG, Bawadekar et al also quantified citrullinated proteins in lysates in a mouse model of TNF-α induced lung inflammation and showed that PADs other than PAD4 are responsible for inflammation-mediated protein citrullination in lungs (Bawadekar et al 2016). Moreover, Carmona-Rivera et al used Rh-PG to show that citrullinated proteins are abundantly generated during NET formation induced by either IgM or rheumatoid factor (Carmona-Rivera et al 2017).…”