Figure 4.

Effect of GSH treatment 1 h before topical CEES exposure on H2A.X ser139 phosphorylation in female SKH-1 hairless mouse skin. Mice were treated either with 300 mg/kg/mouse GSH by oral gavage 1 h before 2 mg CEES exposure, GSH alone, CEES alone, acetone alone as CEES vehicle control or left untreated. Mice were sacrificed after 24 h of CEES exposure, dorsal skin was collected, and western immunoblotting or IHC analysis for H2A.X ser 139 phosphorylation was carried out as detailed in the Material and Methods. Representative pictures of H2A.X ser139 IHC staining are shown in panel A. In IHC stained skin tissue, the H2A.X ser139 positive cells were counted in five randomly selected fields per tissue section (400X magnification); data presented are mean ± SEM of five animals in each group (B). After western immunoblotting for H2A.X ser139 phosphorylation, membranes were reprobed with β-actin as a loading control (C). Data are presented as fold increase in comparison with their respective vehicle control and values of band intensities were adjusted with β-actin loading control. Data are representative of the results from randomly chosen two animals taken at each time point of the study for CEES exposed and GSH treated groups (C). UC, untreated control; VC, vehicle (acetone) control; G, Glutathione alone; GC, Glutathione + CEES; red arrows, H2A.X ser139 positive cells.