香烟烟雾提取物对巨噬细胞线粒体功能的影响*

doi:10.12047/j.cjap.6370.2022.114

中国应用生理学杂志 ›› 2022, Vol. 38 ›› Issue (6): 628-632.doi: 10.12047/j.cjap.6370.2022.114

香烟烟雾提取物对巨噬细胞线粒体功能的影响^*

魏金淑, 田园, 于晓雅, 管美琪, 蔚京京^△

山西医科大学儿科医学系, 太原 030001

收稿日期:2022-10-18 修回日期:2022-11-25 出版日期:2022-11-28 发布日期:2023-06-12
通讯作者: ^△Tel: 13934228746; E-mail: weijj82@126.com
基金资助:
^*山西省科技厅自然科学研究面上项目基金资助(202203021211242)

Effects of cigarette smoke extract on mitochondrial function of macrophages

WEI Jin-shu, TIAN Yuan, YU Xiao-ya, GUAN Mei-qi, WEI Jing-jing^△

Department of Pediatrics, Shanxi Medical University, Taiyuan 030001, China

Received:2022-10-18 Revised:2022-11-25 Online:2022-11-28 Published:2023-06-12

摘要/Abstract

摘要： 目的: 研究香烟烟雾提取物(CSE)对巨噬细胞线粒体功能的影响。 方法: 本研究选取RAW264.7巨噬细胞进行实验。当细胞融合度达70%左右,弃旧培养液,将100% CSE原液用无血清的DMEM和FBS稀释成1%、5%、15%、25%和90%CSE加入到孔板中,采用CCK-8法检测不同浓度CSE处理RAW264.7细胞24 h的细胞活性;再选取最佳CSE浓度分别处理细胞0 h、24 h、48 h、72 h,CCK-8法检测CSE处理细胞不同时间组细胞活性。0%、5%和25 % CSE处理24 h后,Annexin V-FITC /PI染色检测细胞坏死和凋亡情况;JC-1线粒体膜电位检测试剂盒检测细胞线粒体膜受损情况;ROS特异性染料DCFH-DA对小鼠巨噬细胞进行染色后,流式细胞计数法测定荧光度及ROS阳性巨噬细胞数目比例;增强型ATP检测试剂盒检测细胞内ATP浓度。结果: ①和0% CSE组比较,1% CSE处理组细胞活性显著增加(P＜0.01),当CSE浓度为5%以上时细胞活性显著下降(P＜0.05);使用5% CSE处理巨噬细胞,随着处理时间的增加细胞活性显著下降(P＜0.01)。②和0% CSE组比较,5% CSE和25% CSE处理主要导致巨噬细胞坏死,引起巨噬细胞线粒体膜电位显著下降、ROS生成显著增加(P＜0.01)、ATP浓度显著减少(P＜0.05或P＜0.01),且25% CSE处理组的变化更为显著(P＜0.05或P＜0.01)。结论: CSE处理可能影响巨噬细胞线粒体功能,进而导致细胞活性下降并出现坏死。

关键词: 香烟烟雾, 巨噬细胞, 线粒体功能, 细胞坏死

Abstract: Objective: To investigate the effects of cigarette smoke extract (CSE) on the mitochondrial function of macrophages. Methods: RAW264.7 macrophages were used for the experiment in this study. When the cell density was about 70%, the old culture medium was abandoned, and the 100% CSE stock solution was diluted with serum-free DMEM and FBS into 1%, 5%, 15%, 25% and 90% CSE and added to the well plate. The cell activity of RAW264.7 cells treated with CSE at different concentrations for 24 h was detected by CCK-8 method. Then the optimal CSE concentration was selected to treat cells for 0 h, 24 h, 48 h or 72 h respectively, and CCK-8 assay was used to detect the cell activity of CSE treated cells at different time groups. After the cells were treated with 0%, 5% and 25% CSE for 24 hours, cell necrosis and apoptosis was detected by Annexin V-FITC /PI staining; Mitochondrial membrane damage of RAW 264.7 was detected by mitochondrial membrane potential assay kit with JC-1; Macrophages were stained with ROS-specific dye DCFH-DA, and then Flow cytometer was used to determine the fluorescence and the proportion of ROS-positive macrophages; the enhanced ATP assay kit was used to detect the intracellular ATP concentration. Results: ①Compared with 0% CSE, cell viability was increased significantly in 1% CSE group (P＜0.01), cell viability was decreased significantly when CSE concentration was above 5% (P＜0.05); Macrophages were treated with 5% CSE, and cell viability was decreased significantly with the increase of treatment time (P＜0.01). ②Compared with 0% CSE, 5% CSE and 25% CSE mainly caused macrophage necrosis, decreased mitochondrial membrane potential, increased ROS production and decreased ATP significantly (P＜0.05 or P＜0.01), and the changes were more significant in 25% CSE treatment group(P＜0.05 or P＜0.01). Conclusion: CSE may affect mitochondrial function of macrophages, leading to decreased cell viability and necrosis.

Key words: cigarette smoke, macrophages, mitochondrial function, necrosis

中图分类号:

R392

魏金淑, 田园, 于晓雅, 管美琪, 蔚京京. 香烟烟雾提取物对巨噬细胞线粒体功能的影响^*[J]. 中国应用生理学杂志, 2022, 38(6): 628-632.

WEI Jin-shu, TIAN Yuan, YU Xiao-ya, GUAN Mei-qi, WEI Jing-jing. Effects of cigarette smoke extract on mitochondrial function of macrophages[J]. CJAP, 2022, 38(6): 628-632.

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香烟烟雾提取物对巨噬细胞线粒体功能的影响^*

Effects of cigarette smoke extract on mitochondrial function of macrophages

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