Abstract: Caspase-8 plays an important role in the mediation of inflammation and the effect of its role in subarachnoid hemorrhage remains elusive. The nucleotide-binding oligomerization domain-like receptor protein 3 inflammasome has been postulated to mediate inflammation during SAH. The aim of the present study was to investigate the effects of caspase-8 inhibition on SAH injury and further elucidate the molecular mechanisms. In this study, a subarachnoid hemorrhage model was established by endovascular perforation process in adult male Sprague-Dawley rats. Z-IETD-FMK (0.5, 1, 2 mg/kg; an inhibitor of caspase-8) was delivered via intravenous (tail vein) injection immediately after subarachnoid hemorrhage. After 12 hours of subarachnoid hemorrhage, western blot assay showed that the expression of cleaved caspase-8 was significantly increased at 12 hours, peaked at 24 hours, and then decreased at 72 hours after subarachnoid hemorrhage. Immunofluorescence staining demonstrated that caspase-8 was expressed in microglia after subarachnoid hemorrhage. Z-IETDFMK significantly improved neurological deficits and reduced brain water content 24 hours after subarachnoid hemorrhage. The Morris water maze and rotarod test confirmed that Z-IETD-FMK significantly improved spatial learning and memory abilities and motor coordination at 21–27 days after subarachnoid hemorrhage. Furthermore, inhibition of caspase-8 activation reduced the expression of pyrin domain-containing 3, caspase-1, and interleukin-1β after subarachnoid hemorrhage. In conclusion, our findings suggest that caspase-8 inhibition alleviates subarachnoid hemorrhage-induced brain injuries by suppressing inflammation. The study was approved by the Institutional Animal Ethics Committee of the First Affiliated Hospital, School of Medicine, Zhejiang University, China (approval No. 2016- 193) on February 25, 2016.

Key words: brain water content, caspase-8, inflammation, Morris water maze, neurological function, neuroprotection, pyrin domain-containing 3, rotarod test, subarachnoid hemorrhage, Z-IETD-FMK