OP0081 SMOKING CHANGES GENE EXPRESSION AND CITRULLINATION IN JOINTS OF MICE AND MEN

G. Camici ¹F. Brentano ²P. Künzler ²C. Kolling ³R. E. Gay ²B. A. Michel ²S. Gay ²C. Ospelt ^2,*

¹Institute of Physiology, University Zurich, ²Center of Experimental Rheumatology and Center of Integrative Human Physiology (ZIHP), University Hospital Zurich, ³Schulthess Clinic, Zurich, Switzerland

Background: Smoking has been recognized as major risk factor for the development of anti-citrullinated protein antibodies positive rheumatoid arthritis in individuals who carry shared epitope alleles. Presumably smoking alters the immunogenicity of proteins in genetically predisposed individuals or induces the expression of molecules that trigger innate or adaptive immune mechanisms thereby increasing the risk for autoimmunity.

Objectives: To analyze differences in gene expression induced by cigarette smoke in joints of mice, by smoking in human synovial tissues and by cigarette smoke extract (CSE) in rheumatoid arthritis synovial fibroblasts (RASFs).

Methods: Mice were exposed to room air (n=8) or cigarette smoke (n=6) in a whole body exposure chamber for 3 weeks, sacrificed and joints were removed. Synovial tissues were obtained from smoking (n=3) and non smoking (n=5) RA patients undergoing joint replacement surgery. RASFs were incubated with 2% and 5% CSE (n=7). Changes in gene expression were detected using whole genome microarrays and verified with Real-time PCR, ELISA and immunoblotting.

Results: Since it was reported that smoking induces citrullination and peptidyl arginine diminase (PAD) enzymes in the lungs of human smokers, we first measured levels of PAD2 and citrullinated proteins in the joints of smoke-exposed mice. While mRNA levels of PAD2 were not changed by smoke exposure, PAD2 protein levels and the total amount of citrullinated proteins were increased in ankle and elbow joints of mice after smoke exposure. In addition, analysis of changes in gene expression showed a significant induction of vascular endothelial growth factor (VEGF) (2.3 fold, p=0.02) and the heat-shock protein DNAJC6 (2.7 fold, p=0.03) in ankle joints of mice exposed to cigarette smoke. In human synovial tissue samples from RA patients, smokers had 2.3 fold higher mRNA levels of VEGF and 1.8 fold higher mRNA levels of DNAJC6. To test whether synovial fibroblasts might be the source of elevated VEGF and DNAJC6 in joints of smokers, we incubated RASFs with 2% and 5% CSE. Both factors were dose dependently and significantly up regulated by CSE (VEGF: 2% 1.9 fold, 5% 2.7 fold, p<0.0001; DNAJC6: 2% 1.8 fold, 5% 2.3 fold, p=0.02).

Conclusion: In the present study we show that smoking can influence gene expression and protein modifications in the joints of mice and men. Therefore we hypothesize that in addition to systemic changes, smoking might also increase the risk to develop RA by locally changing gene expression and inducing post-translational modifications in the synovium.

Disclosure of Interest: None declared