Proceedings of The Physiological Society

Europhysiology 2018 (London, UK) (2018) Proc Physiol Soc 41, PCA341

Poster Communications

Endothelial VEGFR2 KO during CFA induced inflammation is detrimental to murine tibiotalar cartilage integrity

E. D. Vittersø1,2, N. Beazley-Long1,2, L. Donaldson1,2

1. Life Sciences, University of Nottingham, Nottingham, United Kingdom. 2. Arthritis Research UK Pain Centre, Nottingham, United Kingdom.

Introduction The progressive rheumatic disease inflammatory arthritis affects up to 1.1% of the population. The pathophysiology of the disease includes chronic pain, joint inflammation, synovial hyperplasia, degraded bone and cartilage, and dysregulated angiogenesis (Biniecka et al. 2014; Ursum et al. 2013). We hypothesised that endothelial knockout (KO) of vascular endothelial growth factor receptor 2 (VEGFR2) would affect cartilage pathology in Freund's complete adjuvant (CFA) induced inflammation in mice. Methods Tie2CreERT2xVEGFR2fl/flor wildtype mice were dosed with 5mg tamoxifen or vehicle (intra-peritoneal, in sunflower oil, 1mg per day over 5 days) to induce knockout of endothelial VEGFR2. One week later CFA (160mg in 80mL mineral oil) was injected subcutaneously around the ankle joint in KO or control mice under brief isoflurane anaesthesia (2-3% in O2). On day 14 post injection animals were killed, and ankle joints removed and decalcified. Ankles were sectioned, and treated with Toluidine blue, staining nuclei blue and chondrocytes purple. Severity of cartilage loss in the presence of chondrocytes (bleaching), cartilage surface smoothness, and cartilage thickness was graded out of 4 (as described by Pettit et al., 2001), and analysed with one-way ANOVA and Tukey's multiple comparisons test. All animal procedures were performed in accordance with the United Kingdom Animals (Scientific Procedures) Act 1986/Amendment Regulations 2012 and with University of Nottingham Animal Welfare and Ethical Review Group approval. Results An overall significantly exacerbated cartilage pathology was found in CFA injected Cre+Tamoxifen+KO mice (n=5, mean±SEM=2.9±0.4) compared with sham injected KO controls (n=4, 0.6±0.4; p<0.0001). After further investigation this was found to be caused by significantly exacerbated bleaching in the CFA injected mice (3.5±0.4) compared with sham controls (0.4±0.4; p<0.0001), in addition to a reduced cartilage thickness in CFA injected mice (3.0±0.4) compared with sham controls (0.4±0.4; p=0.001). No significant differences were found for surface smoothness. Conclusions Endothelial VEGFR2 KO is detrimental to tibiotalar cartilage integrity during CFA induced inflammation.

Where applicable, experiments conform with Society ethical requirements