Vam3, a resveratrol dimer, inhibits cigarette smoke-induced cell apoptosis in lungs by improving mitochondrial function
Vam3, a resveratrol dimer, inhibits cigarette smoke-induced cell apoptosis in lungs by improving mitochondrial function
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DOI：
10.1038/aps.2014.17
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发表时间：
2014-06-01
影响因子：
8.2
通讯作者：
Hou, Qi
中科院分区：
文献类型：
--
作者：
Xuan, Ling-ling;Shi, Ji;Hou, Qi
目的：研究Vam3（从山葡萄中提取的白藜芦醇二聚体）对香烟烟雾（CS）诱导的肺细胞凋亡的体外和体内作用及其潜在机制。方法：将人支气管上皮细胞系BEAS-2B暴露于香烟烟雾冷凝液（CSC，300 mg/L），并观察细胞凋亡情况。 使用流式细胞术和Hoechst染色测定细胞凋亡。用 TMRE 染色检查线粒体膜电位。分别用 DCFH-DA 荧光和 HPLC-MS/MS 检测 ROS 和神经酰胺水平。使用免疫荧光检测细胞色素c的释放。使用蛋白质印迹法测量 Caspase-9 和中性鞘磷脂酶 2 的表达。稳定表达 GFP 标记的 Bax 的乳腺癌细胞系 MCF7 用于阐明线粒体在 CS 诱导的细胞凋亡中的作用。对于体内研究，雄性小鼠每天两次接触 CS，每次 5 分钟，持续 4 周。小鼠在第一次CS暴露前1小时每天口服Vam3（50 mg center dot kg(-1)center dot d(-1)）或白藜芦醇（30 mg center dot kg(-1)center dot d(-1)）。结果：用Vam3（5 mu mol/L）或白藜芦醇（5 pmol/L）预处理BEAS-2B细胞显着抑制 CSC 诱导细胞凋亡，并阻止 CSC 诱导的线粒体 Bax 水平增加、线粒体膜电位丧失、细胞色素 c 释放和 caspase-9 激活。此外，用 Vam3 或白藜芦醇预处理 BEAS-2B 细胞可显着抑制 CSC 刺激的细胞内神经酰胺产生，以及 CSC 诱导的中性鞘磷脂酶 2（负责支气管上皮细胞中神经酰胺产生的酶）的上调。在体外 C6-吡啶神经酰胺诱导的 GFP-Bax 稳定的 MCF7 细胞凋亡中，以及在口服 Vam3 或白藜芦醇治疗的 CS 暴露小鼠的肺部中，也获得了类似的结果。结论：Vam3 通过阻止线粒体在体外和体内保护支气管上皮细胞免受 CS 诱导的细胞凋亡。 功能障碍。
Aim: To investigate the effects of Vam3 (a resveratrol dimer extracted from Vitis amurensis Rupr) on cigarette smoke (CS)-induced cell apoptosis in lungs in vitro and in vivo and the underlying mechanisms of action.Methods: Human bronchial epithelial cell line BEAS-2B was exposed to cigarette smoke condensate (CSC, 300 mg/L), and cell apoptosis was determined using flow cytometry and Hoechst staining. Mitochondrial membrane potential was examined with TMRE staining. ROS and ceramide levels were detected with DCFH-DA fluorescence and HPLC-MS/MS, respectively. Cytochrome c release was detected using immunofluorescence. Caspase-9 and neutral sphingomyelinase 2 expression was measured with Western blotting. The breast carcinoma cell line MCF7 stably expressing GFP-tagged Bax was used to elucidate the role of mitochondria in CS-induced apoptosis. For in vivo study, male mice were exposed to CS for 5 min twice a day for 4 weeks. The mice were orally administered Vam3 (50 mg center dot kg(-1)center dot d(-1)) or resveratrol (30 mg center dot kg(-1)center dot d(-1)) each day 1 h before the first CS exposure.Results: Pretreatment of BEAS-2B cells with Vam3 (5 mu mol/L) or resveratrol (5 pmol/L) significantly suppressed CSC-induced apoptosis, and prevented CSC-induced Bax level increase in the mitochondria, mitochondrial membrane potential loss, cytochrome c release and caspase-9 activation. Furthermore, pretreatment of BEAS-2B cells with Vam3 or resveratrol significantly suppressed CSC-stimulated intracellular ceramide production, and CSC-induced upregulation of neutral sphingomyelinase 2, the enzyme responsible for ceramide production in bronchial epithelial cells. Similar results were obtained in C6-pyridinium ceramide-induced apoptosis of GFP-Bax-stable MCF7 cells in vitro, and in the lungs of CS-exposed mice that were treated with oral administration of Vam3 or resveratrol.Conclusion: Vam3 protects bronchial epithelial cells from CS-induced apoptosis in vitro and in vivo by preventing mitochondrial dysfunction.