Protective, Antioxidative and Antiapoptotic Effects of 2-Methoxy-6-Acetyl-7-Methyljuglone from Polygonum cuspidatum in PC12 Cells

Much correlative evidence indicates that the oxidative modification of protein by reactive oxygen species (ROS) is involved in normal aging as well as the pathogenesis of neurodegenerative diseases such as Alzheimer's disease. In this study, we explored the antioxidative and neuroprotective effects of a naphthoquinone, 2-methoxy-6acetyl-7-methyljuglone (MAM), purified from the dried rhizome of Polygonum cuspidatum (Chinese name Hu-Zhang). Pretreatments with MAM (24 h) were investigated for their protective effects against apoptosis induced by the oxidizing agent tert-butyl hydroperoxide (t-BHP) in PC12 cells. The results indicated that MAM pretreatments could effectively protect PC12 cells against cytotoxicity induced by t-BHP in a dose-dependent manner. Cell viability was determined by both MTT and LDH assays. Increasing concentrations of MAM enhanced cell viability significantly and completely prevented cell death induced by t-BHP at 2.5 mu M. The corresponding extracellular lactate dehydrogenase (LDH) levels were also attenuated significantly by various concentrations of MAM. In addition, it was found that the antioxidative effect of MAM was stronger than those of resveratrol and lipoic acid. The antiapoptotic property of MAM was further investigated with Hoechst 33342 nuclear staining and TUNEL assay. Pretreatments of MAM were able to prevent the t-BHP-induced nucleus fragmentation and accumulation of apoptotic bodies (commonly accepted as markers of apoptosis) inside the cells in a dose-dependent manner. t-BHP induced the phosphorylation of ERK 1/2, JNK and p38 MAPK, which were all impeded by pretreatments with MAM, indicating that MAM may act as a potent antioxidant which significantly interferes with the MAPK apoptotic cascades, probably rescuing cells by inhibiting the death pathways.