Transient Receptor Potential Canonical 5 (TRPC5) is protective in chronic arthritis induced by Complete Freund’s Adjuvant (CFA)

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterised by inflammation of diarthrodial joints. Sensory neurons expressing transient receptor potential (TRP) channels, a wide family of non-selective cation channels, have been shown to play a role in the pathogenesis of RA¹. We investigated the role of transient receptor potential canonical 5 (TRPC5) in a model of unilateral arthritis.

All experiments were conducted under the guidelines of the United Kingdom Home Office Animals (Scientific Procedures) Act 1986. Male 129S1/SvImJ wildtype (WT) and TRPC5 knockout (KO) mice littermates (n=8-9, 20-25 g) were used >8 weeks of age. TRPC5 KO mice were bred from heterozygotic mice provided by Dr. D.E. Clapham (Howard Hughes Medical Institute, Boston, U.S.A)². Arthritis was induced by Complete Freund’s Adjuvant(CFA [10 μg, i.art.]), the contralateral joint received saline (10 µl); inflammation was allowed to progress to 14 days. Nociceptive behavioural tests were carried out weekly to examine primary and secondary hyperalgesia. Ex vivo analysis, including qPCR of the synovial membrane and analysis of pro-inflammatory cytokines were carried out as previously described^1,3. Separate studies involved chronic pharmacological antagonist of TRPC5 1h prior to induction of arthritis and daily thereafter. TRPC5 WT and KO mice received ML204⁴ (2mg/kg, i.p., n=6) or vehicle (2% DMSO in saline, n=4). Results are presented as mean ± s.e.m. and analysed by two-way ANOVA and Bonferronni post hoc test.

WTmice developed primary hyperalgesia with a significant reduction in withdrawal threshold compared to baseline responses (334.7±25.7 gf vs 452.2±8.2gf; p<0.01]). Primary hyperalgesia was exacerbated in TRPC5 KO mice compared to WTmice on day 14 (267.4±34.2 vs 334.7±25.47 gf; p<0.05]). This was associated with similar deficits in weight-bearing on the ipsilateral hind-limb which was significantly reduced compared to WT mice on day 14 (37.1±0.9 % vs 44.6±1.6 %; p<0.01]). Pharmacological antagonism of TRPC5 (ML204, 2mg/kg)in WT mice produced similar results observed in TRPC5 KO mice where nociceptive responses were significantly exacerbated compared to vehicle treated WT mice. Weight-bearing asymmetry was augmented in antagonist-treated WT mice compared to vehicle treated WT mice (p<0.05). Cytokine concentrations were increased in the ipsilateral synovial lavage fluid of TRPC5 KO mice in comparison to WT mice; this was observed in chronic administration of ML204 in WT mice. Collectively, our results highlight a protective role for TRPC5 in inflammatory joint conditions. We present in vivo evidence for a role of TRPC5 in regulating joint inflammation. Genetic deletion and pharmacological antagonism of TRPC5 resulted in a significantly exacerbated inflammatory response highlighted by heightened nociceptive responses, and increased secretion of pro-inflammatory cytokines.

References:

1. Fernandes, E.S., et al. Arthritis and rheumatism 63, 819-829 (2011).

2. Riccio, A., et al. Cell 137, 761-772 (2009).

3. Keeble, J., et al. Arthritis and rheumatism 52, 3248-3256 (2005).

4. Miller, M., et al. The Journal of biological chemistry 286, 33436-33446 (2011).

Supported by Arthritis Research UK.