Figure 2. Nox4 deficiency confers long-term neuroprotection and reduces mortality after acute ischemic stroke in young adult and aged mice of either sex.
(A) Upper panel shows representative TTC staining of three corresponding coronal brain sections of 6- to 8-wk-old male and female wild-type (WT) mice, male Nox1 y/− mice, male Nox2 y/− mice, and male and female Nox4 −/− mice, as well as 18- to 20-wk-old male wild-type and Nox4 −/− mice on day 1 after tMCAO. The ischemic infarcts (white) appear smallest in the Nox4 −/− mice of either age or sex (arrows), and this result was confirmed by infarct volumetry (lower panel). ***, p<0.0001, and **, p<0.001, one-way ANOVA, Bonferroni post-hoc test compared with wild-type mice (n = 8–19 per group). (B) Neurological Bederson score (upper panel) and motor score (lower panel) on day 1 after tMCAO in the eight mouse groups indicated above. (C) Serial magnetic resonance images of cerebral infarcts 1 d and 6 d after tMCAO in wild-type and Nox4 −/− mice (lower panel). The broken white lines show hyperintense ischemic lesions on day 1 after tMCAO in wild-type and Nox4 −/− mice. Infarcts on day 1 are smaller in Nox4 −/− mice than in wild-type mice and remain restricted to the basal ganglia on day 6. Hematoxylin and eosin staining confirmed neuronal damage in the cortex of wild-type mice 24 h after tMCAO (top panel, left), whereas cortical integrity was preserved in Nox4 −/− mice (top panel, right). (D) Mortality (upper panel) and long-term functional outcome (Bederson score, lower panel) in 6- to 8-wk-old male Nox4 −/− mice and wild-type controls. Survival curve (upper panel): **, p = 0.0039, log-rank test compared with wild-type mice (n = 10–15 per group). Long-term outcome (lower panel): ***, p<0.0001, and *, p<0.05, one-way ANOVA, Bonferroni post-hoc test compared with wild-type mice (n = 10–15 per group). (E) Upper panel shows representative TTC staining of three corresponding coronal brain sections of 6- to 8-wk-old male wild-type mice (left) and matching Nox4 −/− mice (right) on day 1 after pMCAO. Lower panel: Infarct volumes as measured by infarct volumetry (left) and Neurological Bederson score (right). Nox4 deficiency also protects the brain from permanent ischemia. **, p<0.001, and *, p<0.05, two-tailed Student's t-test compared with wild-type mice (n = 7–11 per group). (F) Representative coronal brain sections of wild-type and Nox4 −/− mice stained with TTC on day 1 after permanent cortical photothrombosis (PT) (upper panel). Cortical infarctions are smaller in the absence of NOX4 (arrow). The lower panel shows infarct volumes in wild-type and Nox4 −/− mice on day 1 after cortical photothrombosis. **, p<0.001, two-tailed Student's t-test compared with wild-type mice (n = 7 per group). All scale bars represent 100 µm.