Figure 3
CCI-TBI results in increased hydrogen peroxide production and 3-nitrotyrosine detection in the ipsilateral cortex area of impact. (A) OxiSelect™ Hydrogen Peroxide/Peroxidase Assay Kit was used to measure hydrogen peroxide levels in fresh mouse cortical tissue ipsilateral to the area of impact 4 hrs following CCI-TBI, CCI-TBI+anti-ICAM-1/catalase, catalase only, and antibody only administration or from naive and sham controls. The naive condition provides basal hydrogen peroxide levels in uninjured cortical tissue which are not significantly different from sham. CCI-TBI results in a nearly 3-fold increase in hydrogen peroxide level detection. Anti-ICAM-1/catalase quenches hydrogen peroxide levels in the area of injury to physiological levels comparable to that of naive and sham, an effect not achieved with anti-ICAM-1 antibody and catalase alone. Immunohistochemical chomogen staining of 3-NT demonstrates increased tyrosine nitration following CCI-TBI (D) compared to naive and sham controls (B and C, respectively). Anti-ICAM-1/catalase significantly decreased 3-NT detection (E). 3-NT levels in anti-ICAM-1 antibody and catalase only groups did not significantly differ from CCI-TBI alone (F and G, respectively). Positive signal detection was pseudo-colored red for better visualization. All images taken at 20X. Scale bar equals 50 microns. (H) Quantification of 3-NT signal detection was analyzed by percent area and normalized to total tissue area per image. Data are presented as mean ± SD. (Ordinary one-way ANOVA with multiple comparisons for hydrogen peroxide data F = 25.95, P < 0.0001 and 3-NT data F = 21.07, P < 0.0001).
