原花青素B2对H2O2诱导的星形胶质细胞氧化损伤和凋亡的保护作用及机制研究[①]

摘要：目的 本研究旨在探讨原花青素B2（proanthocyanidin B2, PC-B2）对过氧化氢（H2O2）诱导的小鼠星形胶质细胞（astrocytes, AS）氧化损伤和凋亡的保护作用及其机制。方法 利用C57BL/6新生小鼠（1-3 d）分离、培养AS，通过随机数字表法分为正常组、正常+PC-B2组（100 μg·mL-1的PC-B2处理24 h）、H2O2模型组（200 μmol·L-1的H2O2处理24 h）、H2O2+PC-B2组（200 μmol·L-1的H2O2与100 μg·mL-1的PC-B2共同处理24 h）；CCK-8法检测各组细胞存活率，LDH法进行细胞毒性检测；ELISA试剂盒检测各组细胞中MDA含量以及SOD、CAT和GSH-Px活力；TUNEL染色法检测各组细胞凋亡情况；RT-PCR和Western Blot分别检测AS中Bax、Bcl-2、Caspase-3、Akt/Stat3、p-Akt、p-stat3、Nrf2/HO-1的mRNA和蛋白表达水平。结果 PC-B2能够明显增强细胞活力，抑制AS凋亡。并且与H2O2模型组相比，PC-B2干预能够显著降低AS中LDH、MDA含量，提高SOD、CAT和GSH-Px活力，抑制Bax，caspase-3的mRNA和蛋白表达，上调Akt/Stat3、Bcl-2、Nrf2/HO-1的mRNA和蛋白表达。结论 PC-B2能够通过Akt/Stat3和Nrf2/HO-1途径增强AS抗氧化能力，减轻H2O2诱导的AS氧化损伤和凋亡。

Protective mechanism of proanthocyanidin B2 against H2O2-induced oxidative damage and apoptosis of astrocytes

ABSTRACT: OBJECTIVE The purpose of this study was to investigate the protective effect of proanthocyanidin B2 (PC-B2) on oxidative damage and apoptosis of mouse astrocytes (AS) induced by hydrogen peroxide (H2O2) and its mechanism. METHODS AS were isolated and cultured from neonatal C57BL/6 mice (1-3 d). They were randomly divided into normal group, normal+PC-B2 group (100 μg·mL?1 PC-B2 treatment for 24 h), H2O2 model group (200 μmol·L?1 H2O2 treatment for 24 h), H2O2+PC-B2 group (200 μmol·L?1 H2O2 and 100 μg·mL?1 PC-B2 treatment for 24 h). The cell viability of each group was detected by CCK-8 method. Cytotoxicity was detected by LDH method. The content of MDA and the activity of SOD, CAT and GSH-Px were detected by ELISA kit. Detection of apoptosis in each group was done by TUNEL staining. The mRNA and protein expression levels of Bax, Bcl-2, Caspase-3, Akt/Stat3, p-Akt, p-stat3 and Nrf2/HO-1 in AS were detected by RT-PCR and Western Blot, respectively. RESULTS PC-B2 can significantly enhance cell viability and inhibit AS apoptosis. Compared with the H2O2 model group, PC-B2 intervention could significantly reduce the content of LDH and MDA in AS, and increase the activity of SOD, CAT and GSH-Px. PC-B2 intervention could inhibit the mRNA and protein expression of Bax and caspase-3, and up-regulate the mRNA and protein expression of Akt/Stat3, Bcl-2, Nrf2/HO-1. CONCLUSION PC-B2 can enhance the antioxidant capacity of AS through Akt/Stat3 and Nrf2/HO-1 pathways, therefore reduce H2O2-induced AS oxidative damage and apoptosis.