Figure 6. NADPH oxidase is increased in MG/MΦ in the aged TBI mice and is associated with reduced antioxidant enzyme expression.

Quantitative real-time PCR was used to assess the expression levels of SOD1 (A), GPX1 (B) p22^phox (C) and gp91^phox (D) in cortical tissue from young (3 month) and aged (24 month) mice at 24 h post-injury. SOD1 (A) expression levels were significantly reduced in aged sham mice when compared to young sham mice (p<0.01), and TBI in aged mice resulted in significantly reduced SOD1 levels when compared to young TBI mice (p<0.01). GPX1 (B) expression levels were significantly increased in the young TBI mice when compared to young sham mice (p<0.05), and TBI in aged mice resulted in significantly reduced GPX1 levels when compared to young TBI mice (p<0.05). p22^phox (C) and gp91^phox (D) expression levels were significantly increased in aged sham mice when compared to young sham mice (p<0.01), and TBI increased the expression of both subunits (p<0.001). The levels of p22^phox and gp91^phox were significantly higher in the aged TBI mice when compared to young TBI mice (p<0.001). Statistical analysis was by two-way ANOVA (injury x aging), followed by post-hoc adjustments using Student-Newman-Keuls Multiple Comparison test; +p<0.5, +++p<0.001, #p<0.05, ##p<0.01, ###p<0.001, ^^p<0.01, and ***p<0.001, where + = TBI 3 month vs. sham 3 month, * = TBI 24 month vs. sham 24 month, # = TBI 3 month vs. TBI 24 month, ^ = sham 3 month vs. sham 24 month. Bars represent mean ± s.e.m. (E) Triple immunofluorescence staining for gp91^phox (red), Iba-1 (green) and ED1 (magenta) in the cortex of young and aged TBI mice at 7 d post injury. gp91^phox expression was strongly up-regulated in ED1- and Iba-1-positive MG/MΦ that displayed a hypertrophic/bushy cellular morphology in the aged TBI mice when compared to young TBI mice. Bar = 200μm, inset bar = 25μm.