THU0515 C5orf30 A Novel Regulator of Tissue Damage in Rheumatoid Arthritis
Authors: M. Muthana, H. Davies, S. Khetan, F. Wright, S. Hawtree, B. Ciani, A. Wilson
Journal: Annals of the Rheumatic Diseases (2014)

<h3>Background</h3> A recent genome wide association study identified the variant rs26232 in the first intron of the uncharacterized gene, <i>C5orf30</i>, as a rheumatoid arthritis (RA) susceptibility variant<sup>1</sup>. In addition, it has been associated with severity of radiological joint damage suggesting a role in tissue breakdown<sup>2</sup>. To date there is no function assigned for <i>C5orf30</i> and neither the gene or protein show homology to any known functional sequences. However, C5orf30 <i>i</i>s highly conserved in chimpanzee, dog, cow, mouse, chicken, and zebrafish (orthologs). <h3>Objectives</h3> The aim of this study is to determine the biological roles of C5orf30 in rheumatoid arthritis. <h3>Methods</h3> Immunohistochemistry on synovial samples was used to determine expression of C5orf30 including co-localisation using antibodies to macrophages (CD68), fibroblasts (5B5), T (CD3) &amp; B (CD19) cells. Real time PCR and western blotting were used to examine C5orf30 transcript and protein levels in fibroblast-like synovial cells (FLS treated with TNF &amp; hypoxia). To investigate gene function siRNA was used to knockdown (KD) either <i>C5orf30</i> or a non-targeting control (NTC) in synovial FLS <i>in vitro</i>. After knockdown cell viability, proliferation, invasion and migration were all assessed and an Illumina BeadChip was used to analyse effects of siRNA-mediated <i>C5orf30</i> repression on global gene expression. <h3>Results</h3> Confocal microscopy revealed C5orf30 to be strongly expressed in both the cytoplasmic compartment of RA synovial lining cells including macrophages and fibroblasts, but not T &amp; B cells. C5orf30 was undetectable in arthroscopy sections obtained from osteoarthritis or control synovium. C5orf30 was expressed in FLS and was found to be up-regulated by hypoxia (8-fold) and down-regulated by TNF treatment (0.5-fold). We found that <i>C5orf30</i>KD did not affect cell viability and proliferation but increased the invasiveness of FLS assayed using Matrigel (<i>p</i>=0.01) and increased FLS migration in a scratch wound assay (<i>p</i>=0.02) (n=6). In support of this gene profiling studies revealed upregulation of cell migration, adhesion, angiogenesis, and immune and inflammatory pathways as a result of <i>C5orf30KD</i>. <h3>Conclusions</h3> C5orf30 is expressed in synovial cells and to a much lesser extent in circulating peripheral blood leukocytes obtained from RA patients. <i>C5orf30</i> knockdown increased FLS migration and invasion into matrigel confirming that C5orf30 is a negative regulator of tissue breakdown. <h3>References</h3> Stahl EA, Raychaudhuri S, Remmers EF, et al. Genome-wide association study meta-analysis identifies seven new rheumatoid arthritis risk loci. Nat Genet 2010;42:508-14. Teare MD, Knevel R, Morgan MD, et al. Allele-Dose Association of the C5orf30 rs26232 Variant With Joint Damage in Rheumatoid Arthritis. Arthritis Rheum. 2013;65:2555-61. <h3>Disclosure of Interest</h3> : None declared <h3>DOI</h3> 10.1136/annrheumdis-2014-eular.4284