Dietary grape seed proanthocyanidins suppress UVB-induced oxidative stress and activation of MAPK proteins and nuclear factor-κB through restoration of MAP kinase phosphatases in SKH-1 hairless mice

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We have shown previously that dietary grape seed proanthocyanidins (GSPs) inhibit ultraviolet (UV) B-induced photocarcinogenesis in mice. As UVB-induced oxidative stress and oxidative stress-mediated signaling have been implicated in photocarcinogenesis, this study was designed to investigate the effect of dietary GSPs on UVB-induced oxidative stress in in vivo SKH-1 hairless mouse model. Here, we report that provision of dietary GSPs (0.2 and 0.5%, w/w) with control AIN76A diet to mice exposed to either acute UVB irradiation (120 mJ/cm²) or chronic irradiation of UVB (for one month on alternate days) inhibited UVB-induced: (i) depletion of antioxidant defense enzymes, such as, glutathione peroxidase, catalase and glutathione, and (ii) the levels of H₂O₂, lipid peroxidation, protein oxidation and nitric oxide in mouse skin. As UV-induced oxidative stress mediates activation of mitogen-activated protein kinases (MAPK) and nuclear factor-kappa B (NF-κB) signaling pathways, we determined the effect of dietary GSPs on these pathways. Employing western blot and immunohistochemical analysis, we found that treatment of mice with GSPs inhibited UVB-induced phosphorylation of extracellular signal-regulated kinase (ERK1/2), c-Jun N-terminal kinase (JNK) and p38 proteins of MAPK family. To determine the negative regulation of MAP kinase phosphatases on UVB-induced increases in the phosphorylation of MAPK proteins, we determined the levels of ERK1/2, JNK and p38-specific MAP kinase phosphatases in the skin samples using the antibodies against MAP kinase phosphatases having dual-specificity directed to phosphothreonine and phosphotyrosine residues within MAP kinases. Western blot analysis revealed that chronic UVB exposure suppressed the levels of MAP kinase phosphatases in the mouse skin compared to non-UVB-exposed control mice; however, the UVB-induced suppression or inactivation of MAP kinase phosphatases was reversed or restored in mice which were treated with dietary GSPs. GSPs also inhibited UVB-induced activation of NF-κB/p65 through inhibition of degradation of IκBα and activation of IKKα. As NF-κB-targeted genes play critical roles in inflammation and cellular proliferation, we assessed the effect of GSPs on proteins encoded by these genes. We found that dietary GSPs resulted in inhibition of UVB-induced expression of PCNA, cyclin D1, iNOS and COX-2 in the skin. Collectively, our data demonstrate that GSPs have the ability to protect the skin from UVB radiation-induced oxidative stress and activation of MAPK and NF-κB signaling pathways and thus provide a molecular basis for the photoprotective effects of GSPs in an in vivo animal model.