Anti-Keap1 PAb

Background: 4,5-di-O-caffeoylquinic acid methyl ester (4,5-CQME) is a caffeoylquinic acid (CQA) isolated from Lonicera japonica Thunb., a traditional Chinese medicine. To date, the biological activity of 4,5-CQME has not been fully investigated. Purpose: The aim of the current study was to explore the anti-oxidative activity and the underlying mechanism of 4,5-CQME. Methods: MIT assay was used to evaluate the cytoprotective effect of 4,5-CQME. DCFH-DA was used as a fluorescence probe to detect intracellular ROS. The mitochondrial membrane potential was detected using the fluorescent probe JC-1. MDA and GSH levels were measured using MDA and GSH commercial kits, respectively. Apoptosis assay was performed using the Annexin V-FITC/PI method. The functional mechanism of 4,5-CQME was investigated by analyzing relative signaling pathways through immunofluorescent staining, quantitative PCR and western blot analysis. Results: HepG2 cells were incubated with different concentrations of 4,5-CQME for 12 h before exposure to 500 mu M H2O2 for 3 h. 4,5-CQME attenuated H2O2-induced oxidative damage and had a higher cytoprotective effect than 3-caffeoylquinic acid, 3-caffeoylquinic acid methyl ester, or 4,5-di-O-caffeoylquinic acid. 4,5-CQME also reduced ROS and MDA levels and rescued GSH depletion. Western blots demonstrated that 4,5-CQME decreased Bax/Bcl-2 and Bak levels. A mechanistic study confirmed that 4,5-CQME significantly suppressed H2O2-induced MAPKs phosphorylation but had little effect on MAPKs phosphorylation under normal conditions. By contrast, 4,5-CQME induced AKT phosphorylation in the presence or absence of H2O2. 4,5-CQME also regulated the Keap1/Nrf2 signaling pathway and enhanced both the mRNA and protein expressions of HO-1 and NQO1. The anti-oxidative effect of 4,5-CQME was greatly abolished by co-incubation with the Nrf2 inhibitor ML385 or PI3K inhibitor wortmannin. Conclusions: Taken together, these results showed that 4,5-CQME offered significant protection against H2O2-induced oxidative stress, and its effect was in part due to the modulation of the Keap1/Nrf2 pathway.

This study investigated whether resveratrol could improve meat quality, muscular antioxidant capacity, lipid metabolism and fiber type composition of intrauterine growth retarded pigs. Thirty-six pairs of male normal birth weight and intrauterine growth retardation (IUGR) piglets were orally fed with 80 mg resveratrol/kg body weight/d or vehicle during the sucking period (7-21 d). Then the offspring were fed with a basal diet containing 300 mg resveratrol/kg or a basal diet from weaning to slaughter (150 d). The IUGR-impaired meat quality (luminance and yellowness) was associated with muscular oxidative stress via increased Keap1 protein level, fat accumulation, and higher MyHC IIb gene expression. Expectedly, resveratrol increased glutathione peroxidase activity and MyHC I gene expression, reduced protein carbonyl and malondialdehyde contents, enhanced fatty acid oxidation via upregulated PPAR alpha and targeted genes expression, and thereby improving drip loss and yellowness. Results indicate that resveratrol improved meat quality of IUGR pigs through enhancing antioxidant capacity, increasing oxidative fiber composition, and suppressing lipid accumulation.