N-乙酰半胱氨酸对香烟烟雾提取物诱导的气道上皮细胞线粒体过氧化损伤的保护作用  

The protective effect of N-acetylcysteine on mitochondrial oxidative damage of airway epithelial cells induced by cigarette smoke extract

作　　者：张明[1] 张莉 杨侠[1] 单虎[1] 张秋红[1] 张红妮 张洁[1] 李雅莉[1] ZHANG Ming;ZHANG Li;YANG Xia;SHAN Hu;ZHANG Qiuhong;ZHANG Hongni;ZHANG Jie;LI Yali(Department of Respiratory and Critical Care Medicine,The Second Affiliated Hospital of Xi’an Jiaotong University,Xi’an 710004;Department of Urology,the Ninth Hospital of Xi’an,Xi’an 710054,China)

机构地区：[1]西安交通大学第二附属医院呼吸与危重症医学科,陕西西安710004 [2]西安市第九医院泌尿外科,陕西西安710054

出　　处：《西安交通大学学报（医学版）》2021年第6期837-842,共6页Journal of Xi’an Jiaotong University（Medical Sciences）

基　　金：陕西省创新能力支撑计划项目(2019KJXX-045);陕西省重点研发计划项目(2020SF-108);国家自然科学基金项目(No.81600030)。

摘　　要：目的探讨N-乙酰半胱氨酸(N-acetylcysteine, NAC)对香烟烟雾提取物(cigarette smoke extract, CSE)诱导的气道上皮细胞线粒体损伤的保护作用及机制。方法体外培养人气道上皮细胞BEAS-2B,分为正常对照组、7.5%(75 mL/L)CSE组、7.5%CSE+NAC组,干预24 h后应用MTT法检测细胞活性,荧光显微镜下观察线粒体内活性氧(reactive oxygen species, ROS)和线粒体膜电位(mitochondrial membrane potential, MMP)强度,流式细胞仪检测MMP水平,Western blotting检测细胞内Sirt3和锰超氧化物歧化酶(manganese superoxide dismutase, MnSOD)的蛋白表达,比色法测定细胞内MnSOD活性。结果 7.5%CSE+NAC组的气道上皮细胞活性较7.5%CSE组显著升高,差异具有统计学意义(P<0.05)。荧光显微镜和流式细胞仪检测结果显示,7.5%CSE+NAC组的气道上皮细胞MMP水平显著高于7.5%CSE组(P<0.05)。与7.5%CSE组相比,7.5%CSE+NAC组气道上皮细胞线粒体内ROS显著降低,Sirt3和MnSOD蛋白表达以及MnSOD活性均显著增加(P<0.05)。结论 NAC通过调控Sirt3-MnSOD信号通路减轻CSE所致的气道上皮细胞线粒体过氧化损伤,有助于阐明NAC对慢性阻塞性肺疾病的作用机制。Objective To investigate the effect of N-acetylcysteine(NAC) on mitochondrial damage of airway epithelial cells induced by cigarette smoke extract(CSE). Methods Human airway epithelial cells(BEAS-2B) were cultured and divided into three groups as follows:normal control group, 7. 5%(75 mL/L)CSE-treated group and 7. 5%CSE plus NAC group. After stimulation for 24 hours, cell viability was determined by MTT, and the levels of mitochondrial reactive oxygen species(ROS) and mitochondrial membrane potential(MMP) were observed under the fluorescence microscope. MMP was also measured by flow cytometry, the protein expressions of Sirt3 and manganese superoxide dismutase(MnSOD)were detected by Western blotting, and MnSOD activity was measured by colorimetry.Results Pretreatment with NAC significantly improved the viability of airway epithelial cells(P<0. 05). The results of fluorescence microscopy and flow cytometry showed that NAC pretreatment significantly attenuated MMP decline in airway epithelial cells exposed to 7. 5% CSE(P<0. 05). Compared with 7. 5% CSE-treated group, mitochondrial ROS in airway epithelial cells was significantly decreased in 7. 5% CSE plus NAC group(P<0. 05). In addition,pretreatment with NAC significantly inhibited the decrease of Sirt3 and MnSOD protein expression and improved MnSOD activity in airway epithelial cells exposed to 7. 5% CSE(P<0. 05). Conclusion NAC attenuates CSE-induced airway epithelial mitochondrial damage through the regulation of Sirt3-MnSOD signaling pathway,which reveals a new mechanism of NAC treatment for chronic obstructive pulmonary disease.

关 键 词：N-乙酰半胱氨酸 气道上皮细胞 线粒体 Sirt3 锰超氧化物歧化酶(MnSOD) 慢性阻塞性肺疾病 

分 类 号：R562.2[医药卫生—呼吸系统]