Session Title: Rheumamtoid Arthritis - Human Etiology and Pathogenesis
Session Type: Abstract Submissions (ACR)
Background/Purpose: Rheumatoid arthritis (RA) is an autoimmune disease characterised by inflammation followed by tissue rebuilding or fibrosis. Failure by the body to effectively regulate inflammation is a hallmark of RA. It has been suggested that periodontal disease is one mechanism whereby tissue inflammation is triggered in RA. One of the organisms implicated in periodontal disease: Porphorymonas gingivalis, is an anaerobic pathogen that is known to produce peptidyl arginine demiminase (PAD), the only known bacterial PAD which causes citrullination. Cleavage of extracellular matrix (ECM) substrates in RA is known to lead to the production of ECM damage-associated molecular patterns, or DAMPs that can then be available for citrullination, thereby mediating chronic inflammation in RA.
Methods: We investigated the ability of proteases produced by Porphorymonas gingivalis to cleave extracellular matrix substrates which are found in the joint and also the human oral mucosa: fibrinogen, fibronectin and type I collagen. Culture supernatants of the anaerobe Porphorymonas gingivalis (strain W83 from ATCC) were produced from 24 hour cultures using full anaerobic conditions (3M Concept Plus anaerobic incubator). After culture, bacterial supernatants were extracted and incubated with the substrates at 0.5 mg/ml at 37o C, with collection of digestion products from 0-320 mins. The cleavage patterns of the substrates described was evaluated by SDS-PAGE and Western blotting in the presence of selective protease inhibitors.
Results: We found that culture supernatants from Porphorymonas gingivalis are effective at cleaving all the substrates tested. The substrate demonstrating the most rapid digestion profile in conditions of 37o C were in order of rapidity of cleavage: fibrinogen (30 mins for complete cleavage), fibronectin (180 mins for complete cleavage) and type I collagen (320 mins). In the presence of arginine inhibitor (100 micromolar NMLA, NG-methyl-L-arginine ) and 50 nM 1400 W (a potent selective inducible NOS inhibitor), digestion of all three substrates was strongly inhibited. In the presence of gingipain inihibitors KYT-1 and KYT-36, greater inhibition of cleavage was demonstrated for KYT-36 than for KYT-31.
Conclusion: Our data demonstrate that selective proteases can cleave extracellular matrix protein substrates shared in the oral mucosa and the arthritic joint. Inhibition of cleavage of such substrates may delay the production of ECM DAMPs that can then be available for citrullination in RA. Therapeutic strategies aimed at inhibiting such cleavage of ECM substrates may be a novel therapeutic target in RA.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/proteases-produced-by-porphyromonas-gingivalis-can-cleave-and-citrullinate-substrates-found-in-the-joint-and-oral-mucosa-implications-for-autoimmunity-in-rheumatoid-arthritis/