The
complement cascade is a central component of innate immunity which plays a critical role in
brain inflammation.
Complement C3a receptor (C3aR) is a key mediator of post-ischemic cerebral injury, and pharmacological antagonism of the C3a receptor is neuroprotective in
stroke.
Cerebral ischemia injures brain endothelial cells, causing blood brain barrier (BBB) disruption which further exacerbates ischemic neuronal injury. In this study, we used an in vitro model of
ischemia (
oxygen glucose deprivation; OGD) to investigate the protective effect of a C3aR antagonist (C3aRA,
SB290157) on brain endothelial cells (bEnd.3). Following 24 hours of reperfusion, OGD-induced cell death was assessed by TUNEL and
Caspase-3 staining. Western blot and immunocytochemistry were utilized to demonstrate that OGD upregulates inflammatory, oxidative stress and
antioxidant markers (ICAM-1, Cox-2, Nox-2 and MnSOD) in endothelial cells and that C3aRA treatment significantly attenuate these markers. We also found that C3aRA administration restored the expression level of the
tight junction protein occludin in endothelial cells following OGD. Interestingly, OGD/
reperfusion injury increased the phosphorylation of ERK1/2 and C3aR inhibition significantly reduced the activation of ERK suggesting that endothelial C3aR may act via ERK signaling. Furthermore, exogenous C3a administration stimulates these same inflammatory mechanisms both with and without OGD, and C3aRA suppresses these C3a-mediated responses, supporting an antagonist role for C3aRA. Based on these results, we conclude that C3aRA administration attenuates
inflammation, oxidative stress, ERK activation, and protects brain endothelial cells following experimental
brain ischemia.