Baicalein inhibition of oxidative-stress-induced apoptosis via modulation of ERKs activation and induction of HO-1 gene expression in rat glioma cells C6
Baicalein inhibition of oxidative-stress-induced apoptosis via modulation of ERKs activation and induction of HO-1 gene expression in rat glioma cells C6
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DOI：
10.1016/j.taap.2006.05.008
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发表时间：
2006-10-15
影响因子：
3.8
通讯作者：
Shen, Shing-Chuan
中科院分区：
文献类型：
--
作者：
Chen, Yen-Chou;Chow, Jyh-Ming;Shen, Shing-Chuan
本研究探讨了黄芩素（BE）及其苷类黄芩苷（BI）对过氧化氢（H2O2）诱导的大鼠胶质瘤C6细胞死亡的保护机制。MTT实验、LDH释放实验和形态学观察结果表明，H2O2的加入降低了C6细胞的活力，BE的加入对这种影响有抑制作用，而BI没有。以DCHF-DA为荧光底物的流式细胞术分析显示，BE孵育C6细胞可显著降低H2O2诱导的细胞内过氧化物水平。在C6细胞中，BE而非BI抑制h2o2诱导的凋亡事件，包括DNA阶梯、次二倍体细胞和caspase 3,8和9的激活。ERK抑制剂PD98059可阻断H2O2诱导的ERK蛋白的细胞毒性和磷酸化作用。过氧化氢酶能抑制h2o2诱导的ROS生成、ERKs蛋白磷酸化和细胞死亡，BE能剂量依赖性地抑制h2o2诱导的C6细胞ERK蛋白磷酸化。这些数据表明，ros清除活性通过阻断ERKs激活参与BE预防h2o2诱导的细胞死亡。此外，BE而非BI诱导热休克蛋白32 （HSP32; HO-1）蛋白表达呈时间和剂量依赖性，但不诱导血红素加氧酶2 （HO-2）、热休克蛋白70 （HSP70）或热休克蛋白90 （HSP90）蛋白表达。在没有H2O2的情况下，BE诱导ERKs蛋白磷酸化，并通过添加环己亚胺、放线菌素D和ERK抑制剂PD98059阻断BE诱导的HO-I蛋白表达。MTT实验和显微镜下观察的细胞凋亡形态学显示，加入HO抑制剂ZnPP抑制了BE对h2o2诱导的C6细胞毒性的保护作用，同时阻断了BE对C6细胞ros的清除活性。然而，BE处理不能保护C6细胞免受c2 -神经酰胺诱导的细胞死亡。这些数据表明，BE具有通过调节ERKs激活和诱导HO-I蛋白表达来抑制ros介导的细胞毒性作用的能力。提出了ho - 1在BE清除ros活性中的作用。(c) 2006爱思唯尔公司版权所有。
In the present study, we examined the protective mechanism of baicalein (BE) and its glycoside, baicalin (BI), on hydrogen-peroxide (H2O2)-induced cell death in rat glioma C6 cells. Results of the MTT assay, LDH release assay, and morphological observation showed that H2O2 addition reduced the viability of C6 cells, and this was prevented by the addition of BE but not BI. Incubation of C6 cells with BE significantly decreased the intracellular peroxide level induced by H2O2 according to flow cytometric analysis using DCHF-DA as a fluorescent substrate. Suppression of H2O2-induced apoptotic events including DNA ladders, hypodiploid cells, and activation of caspases 3, 8, and, 9 by BE but not BI was identified in C6 cells. The cytotoxicity and phosphorylation of ERK proteins induced by H2O2 were blocked by the ERK inhibitor PD98059. Catalase addition prevented H2O2-induced ROS production, ERKs protein phosphorylation, and cell death, and BE dose-dependently inhibited H2O2-induced ERK protein phosphorylatiou in C6 cells. These data suggest that ROS-scavenging activity is involved in BE prevention of H2O2-induced cell death via blocking ERKs activation. Additionally, BE but not BI induced heat shock protein 32 (HSP32; HO-1) protein expression in both time- and dose-dependent manners, but not heme oxygenase 2 (HO-2), heat shock protein 70 (HSP70), or heat shock protein 90 (HSP90) protein expression. In the absence of H2O2, BE induces ERKs protein phosphorylation, and HO-I protein expression induced by BE was blocked by the addition of cycloheximide, actinomycin D, and the ERK inhibitor PD98059. The addition of the HO inhibitor ZnPP inhibited the protective effect of BE against H2O2-induced cytotoxicity in C6 cells according to the MTT assay and apoptotic morphology under microscopic observation, accompanied by blocking the ROS-scavenging activity of BE in C6 cells. However, BE treatment was unable to protect C6 cells from C2-ceramide-induced cell death. These data indicate that BE possesses abilities to inhibit ROS-mediated cytotoxic effects through modulation of ERKs activation and induction of HO-I protein expression. The role of HO-I in ROS-scavenging activity of BE is proposed. (c) 2006 Elsevier Inc. All rights reserved.