茶多酚对甲基汞致大鼠大脑皮质神经元氧化损伤的防护作用

Protective effect of tea polyphenols against methylmercury-induced oxidative damage in rat cortical neurons

目的探讨茶多酚(tea polypheonols,TP)对甲基汞(methylmercury,MeHg)所致大鼠大脑皮质神经元氧化损伤的防护作用及机制。方法进行大鼠大脑皮质神经元原代培养,细胞成熟后给予0.01、0.1、1、2μmol/L MeHg分别处理0.5、1、3、6、12 h,通过测定培养液中乳酸脱氢酶(lactate dehydrogenase,LDH)活力来进行MeHg细胞毒性分析;根据测定结果选择最具代表性的1μmol/L MeHg暴露6 h作为MeHg染毒组。应用同样方法进行TP预处理组选定,向培养液中分别加入终浓度为5、10、20及40μmol/L TP,分别预处理0.5、1、3及6 h后,再加入终浓度为1μmol/L MeHg,继续培养6 h后测定培养液LDH漏出,根据实验结果选定5、10、20μmol/L预处理3 h作为TP预处理剂量及时间;细胞经各剂量TP预处理后,再暴露于1μmol/L MeHg 6 h,测定神经元细胞凋亡率、非蛋白巯基(non-protein sulfhydryl,NPSH)含量、活性氧簇(reactive oxygen species,ROS)水平及Na+-K+-ATPase和Ca2+-ATPase活力。结果与对照组比较,随着染MeHg剂量的升高,培养液中LDH活力逐渐升高,呈现剂量和时间依赖性的效应关系。TP预处理后,LDH活力逐渐降低,在10、20μmol/L TP预处理组显著降低(P<0.05或P<0.01);1μmol/L MeHg导致神经元凋亡率显著升高,NPSH含量显著降低,ROS水平显著升高,Na+-K+-ATPase和Ca2+-ATPase活力显著降低,差异均有统计学意义(P<0.01),TP预处理对上述指标的拮抗作用呈现剂量-效应关系,差异均有统计学意义(P<0.05或P<0.01)。结论 TP对MeHg所致大鼠大脑皮质神经元氧化损伤具有一定的防护作用。

Objective To explore the protective effect of tea polyphenols （ TP ） against methylmercury （MeHg） -induced neuronal oxidative damage in rat cortical neurons. Methods The primary cultured cortical neurons were exposed to 0. 01, 0. 1, 1, and 2 μmol/L MeHg for 0. 5, 1, 3, 6, and 12 h, respectively. According to the leakage situation of LDH in the medium to judge the cytotoxicity of MeHg, 1 μmol/L MeHg for 6 h was intended as treatment group. Same method was used for TP pretreatment and 5, 10, 20μmol/L TP pre-treated for 3 h were intended as TP pre-treatment groups for evaluation the protective effect of TP on neuronal apoptosis, NPSH content, ROS formation, Na＋-K＋-ATPase and Ca2＋-ATPase activities. Results The resuhs showed that comparing with control group, the LDH activities increased in a dose-and time-dependent manner in MeHg exposure groups, while the TP pre-treatment resulted in LDH activity dose-and time-dependently lower, especially in 10 and 20 μmol/L TP pre-treatment for 3 h groups （P〈0.05 or P〈0. 01 ）. In addition, the obvious elevations of neuronal apoptosis rates, of ROS formation, as well as decrease in NPSH content, in activities of Na＋-K＋-ATPase and Ca2＋-ATPase （ P〈0. 01 ） caused by 1 μmol/L of MeHg could partially antagonize by TP pre-treatment （P〈0. 05 or P〈0. 01 ）. Conclusion It is suggested that TP has the ability to prevent the oxidative damage caused by MeHg in cortical neurons of rat.