Oxidative stress after subarachnoid hemorrhage in gp91phox knockout mice

Abstract

Background: Oxidative stress largely contributes to early brain injury after subarachnoid hemorrhage (SAH). One of the major sources of reactive oxygen species is NADPH oxidase, upregulated after SAH. We hypothesized that NADPH oxidase-induced oxidative stress plays a major causative role in early brain injury after SAH.

Methods: Using gp91phox knockout (ko) and wild-type (wt) mice, we studied early brain injury in the endovascular perforation model of SAH. Mortality rate, cerebral edema, oxidative stress, and superoxide production were measured at 24 h after SAH. Neurological evaluation was done at 23 h after SAH surgery.

Results: Genotyping confirmed the existence of a nonfunctional gp91phox gene in the ko mice. CBF measurements did not show differences in SAH-induced acute ischemia between ko and wt mice. SAH caused a significant increase of water content in the ipsilateral hemisphere as well as an increase of Malondialdehyde (MDA) levels and superoxide production. There were no significant differences in post-SAH mortality rate, brain water content and the intensity of the oxidative stress between knockout and wild type groups of mice.

Conclusions: Our results suggest that gp91phox is not critically important to the early brain injury after SAH. An adaptive compensatory mechanism for free radical production in knockout mice is discussed.

Figure 1

Genotyping, neurological score, and brain water content. A. Genotyping. NADPH oxidase gp91phox subunit shown as a 240 bp band in wt mice (Group B) but shifted to 195 bp in ko mice (Group A). B. Neurological score at 23 h post-surgery in SAH and SHAM groups of ko and wt mice; expressed as median scores. Neurological scores in SAH groups were significantly lower than that of SHAM groups, however they did not differ between ko and wt mice. C. Brain water content in SAH and SHAM groups did not show a difference between ko and wt mice. Asterisks, p < 0.05 compared with corresponding SHAM group. NS, not significant (SAH-ko vs. SAH-wt). N = 20 in each SAH group, but N = 5 in each SHAM group.

Figure 2

Malondialdehyde (MDA) levels and superoxide production in cerebral cortex. A. MDA levels significantly increased at 24 h after SAH in ko and wt mice with no difference found between these two groups. Asterisks, p < 0.05 compared with corresponding SHAM group. N = 12 in each SAH group, but N = 6 in each SHAM group. B. SAH animals showed similarly high Et-like signals in ko and wt groups at 24 h post-surgery. The Et-signal-positive areas in SAH groups were significantly larger than that of SHAM groups indicating a prominent superoxide production (B). Asterisks, p < 0.05 compared with corresponding SHAM group. NS, not significant (SAH-ko vs. SAH-wt). N = 8 in each SAH group, but N = 4 in each SHAM group.