330. Characterizing a Population with Serological Features of Anca-Associated Vasculitis and Systemic Lupus Erythematosus

Abstract

serum of patients with SLE (n1⁄423) and healthy controls (n1⁄4 11) which were age and gender matched. Endotoxin was removed using DetoxiGel columns to a level below 0.225 endotoxin U/ml. We utilized an established in vitro model of anoxia/reoxygenation to I/R injury. Cardiomyocytes were isolated from 1–2 day old rat pups and when beating synchronously were treated with 500mg/ml polyclonal IgG from each group and the following day exposed to simulated I/R injury using a hypoxic chamber (argon, 5% CO2) followed by reoxygenation. Apoptosis was measured by assessment of caspase-3 cleavage using immunoblot and TUNEL. Results: In cells exposed to simulated I/R injury caspase-3 cleavage was not significantly increased in the presence of IgG from healthy volunteers (mean increase in caspase-3 cleavage of cells treated with healthy control IgG above untreated cells exposed to simulated I/R injury is 12.28% S.D. 26.01, n1⁄410). However, in the presence of IgG from patients with SLE/aPL þve and SLE/aPL –ve, caspase-3 cleavage was increased above untreated cells exposed to simulated I/R injury by 58.74% ( S.D. 17.8, n1⁄4 6) and 63.98 ( S.D. 17.9, n1⁄48) and therefore significantly increased in comparison with healthy controls (P1⁄4 0.0001). The effect observed with IgG from SLE patients was not altered when serum used in the buffers was heat inactivated suggesting a non-complement mediated mechanism. Conclusion: In this in vitro simulated I/R injury model IgG purified from patients with SLE significantly enhanced I/R injury as assessed by caspase-3 cleavage and TUNEL. This novel pathogenic role of these antibodies will now be tested in vivo to validate this finding and explore potential mechanisms of action. Disclosure statement: The authors have declared no conflicts of interest.