Dihydromyricetin alleviated the damage of hypoxia-induced mouse neurons by reducing ROS levels and inhibiting the expression of PAR and γH2AX

Abstract

AbstractObjectiveTo investigate the effect of dihydromyricetin on hypoxia-induced neurons, to understand the effect of dihydromyricetin on hypoxic-ischemic encephalopathy (HIE).MethodsCortical neurons were isolated from C57BL/6j mice (24 hour-year old), cultured, and subjected to 4h hypoxia and 20h reoxygenation to mimic the neonatal hypoxic-ischemic encephalopathy. After dihydromyricetin (20μmol/L) treatment of hypoxia-induced neurons for 2h, CCK-8 assay was used to analyze the neuronal viability, Hoechst33342/PI double staining assay was used to analyze the neuronal death, Western blotting was used to analyze the expression of Poly ADP-ribose (PAR) polymer protein andγH2AX, comet assay was used to detect DNA damage, immunofluorescence staining was used to observe the nuclear translocation of apoptosis inducing factor, and 2’,7’-dichlorodihydrofluorescein diacetate was used to detect the expression of reactive oxygen species (ROS).ResultsCompared with the control groups, hypoxia-treated neurons exhibited significantly lower activity, higher neuronal death rate and the high expressions of PAR andγH2AX, hypoxia could also induce AIF nuclear translocation, increase tail DNA content and tail length, increase the expression of ROS in neurons; after dihydromyricetin treatment, neuronal activity were significantly increased, neuronal death rate, ROS levels, and the expressions of PAR andγH2AX were also decreased, AIF nuclear translocation was inhibited, the tail DNA content and tail length were also decreased.ConclusionDihydromyricetin could alleviate the damage of hypoxia-induced neurons through decreasing the levels of ROS and inhibiting the expressions of PAR andγH2AX, suggesting that dihydromyricetin may have the protective effect on HIE.