N-乙酰半胱氨酸对PM2.5致支气管上皮细胞损伤的保护作用体外实验

郭淼; 闫鹏; 韩国鑫; 朱明; 金珊; 肖坤; 解立新

doi:10.3969/j.issn.2095-5227.2021.01.021

N-乙酰半胱氨酸对PM2.5致支气管上皮细胞损伤的保护作用体外实验

doi: 10.3969/j.issn.2095-5227.2021.01.021

郭淼^{1, 2, 3,},
闫鹏²,
韩国鑫³,
朱明³,
金珊⁴,
肖坤²,
解立新^2, ,

1.
解放军医学院，北京　100853
2.
解放军总医院呼吸与危重症医学部，北京　100853
3.
战略支援部队特色医学中心急诊科，北京　100101
4.
美中宜和妇儿医院内科，北京　100101

详细信息

作者简介:
郭淼，男，本科，主治医师。研究方向：气道炎症和损伤，危重症诊治。Email: guom306em@163.com

通讯作者:
解立新，男，博士，主任医师，解放军总医院呼吸与危重症医学部主任。Email: xielx301@126.com

中图分类号: R 965
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出版历程
- 收稿日期: 2020-09-08
- 网络出版日期: 2021-02-10
- 刊出日期: 2021-01-28

Protective effect of N-acetylcysteine on PM2.5-induced injury of bronchial epithelial cells in vitro

GUO Miao^{1, 2, 3
,},
YAN Peng²,
HAN Guoxin³,
ZHU Ming³,
JIN Shan⁴,
XIAO Kun²,
XIE Lixin^{2
, ,}

1.
Chinese PLA Medical School, Beijing 100853, China
2.
College of Pulmonary and Critical Care Medicine, Chinese PLA General Hospital, Beijing 100853, China
3.
Emergency Department, PLA Strategic Support Force Characteristic Medical Center, Beijing 100101, China
4.
Internal Medicine, Amcare Women's and Children's Hospital, Beijing 100101, China

More Information

Corresponding author: XIE Lixin. Email: xielx301@126.com

摘要

摘要: 背景大气污染物中的PM2.5可以引起气道上皮细胞炎症反应，N-乙酰半胱氨酸(N-acetyl-L-cysteine，NAC)可以有效减轻这种炎症反应，但相关的自噬机制研究较少。通过对自噬机制的研究，可为抑制PM2.5诱导的气道炎症反应提供新思路。目的探讨N-乙酰半胱氨酸对PM2.5引起的支气管上皮细胞损伤的保护机制。方法将人支气管上皮细胞(BEAS-2B)按照不同暴露因素设为控制对照组、PM2.5组、PM2.5+NAC组。对照组使用普通培养基处理16 h；PM2.5组用普通培养基预处理4 h后再用含有PM2.5的培养基处理12 h；PM2.5+NAC组用含有NAC的培养基预处理4 h再用含有PM2.5的培养基处理12 h。流式细胞仪分析细胞凋亡情况，使用荧光共聚焦显微镜观察LC3荧光量，Western blot法观察LC3Ⅱ/LC3Ⅰ及P62的情况，分析三组细胞发生自噬的情况。结果与对照组和PM2.5+NAC组比较，PM2.5组细胞凋亡明显增加；而PM2.5+NAC组由PM2.5(100 μg/mL)引起的BEAS-2B细胞凋亡明显被抑制。免疫荧光结果显示，与对照组比较，PM2.5组LC3荧光量明显增加；与PM2.5组比较，PM2.5+NAC组LC3荧光量明显减少。Western blot结果显示，与对照组比较，PM2.5组LC3Ⅱ/LC3Ⅰ明显增加，P62明显减少；与PM2.5组比较，PM2.5+NAC组LC3Ⅱ/LC3Ⅰ明显减少，P62明显增加。结论 NAC可以抑制PM2.5介导的人支气管上皮细胞的凋亡和自噬，从而减少细胞的损伤。
- N-乙酰半胱氨酸 /
- PM2.5 /
- 自噬 /
- 凋亡 /
- BEAS-2B细胞
Abstract: Background PM2.5 in air pollutants can cause inflammation in airway epithelial cells, and N-acetylcysteine (NAC) can effectively reduce PM2.5-induced inflammation, but the related autophagy mechanism has been rarely studied. The study on autophagy mechanism will provide a new method for inhibiting PM2.5-induced airway inflammation. Objective To investigate the protective effect of NAC on PM2.5-induced injury of bronchial epithelial cells. Methods Bronchial epithelial cells (BEAS-2B) were divided into control group, PM2.5 group, PM2.5+NAC group according to different exposure factors. The control group was treated with ordinary medium for 16 hours. The PM2.5 group was pretreated with ordinary medium for 4 hours and then treated with PM2.5 medium for 12 hours. The PM2.5+NAC group was pretreated with NAC medium for 4 hours and then treated with PM2.5 medium for 12 hours. Flow cytometry was used to analyze cell apoptosis. LC3 fluorescence was observed by fluorescence confocal microscope, LC3Ⅱ/LC3Ⅰ and P62 were observed by Western Blot. The occurrence of autophagy in the three groups was analyzed. Results Compared with control group and PM2.5+NAC group, the apoptosis of BEAS-2B cells in PM2.5 group increased significantly, while the apoptosis of BEAS-2B cells induced by PM2.5 (100 μg/mL) in PM2.5+NAC group was significantly inhibited by NAC. The immunofluorescence result showed that compared with the control group, the fluorescence of LC3 in PM2.5 group significantly increased. However, it decreased significantly in PM2.5+NAC group when compared with PM2.5 group. Conclusion NAC can inhibit PM2.5-induced apoptosis and autophagy of human bronchial epithelial cells, thereby reducing cell damage.
- N-acetylcysteine /
- PM2.5 /
- autophagy /
- apoptosis /
- BEAS-2B cells

HTML全文

图 1 流式细胞仪测定细胞凋亡及定量分析 (^aP＜0.05，vs PM2.5+NAC；^bP＜0.05，vs PM2.5

Figure 1. Flow cytometry results of apoptosis and quantification analysis (^aP＜0.05, vs PM2.5+NAC; ^bP＜0.05, vs PM2.5

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图 2 细胞自噬荧光图

A：细胞核的DAPI染色(蓝色)、细胞骨架的鬼笔环肽染色(红色)作为背景染色，LC3荧光呈绿色；B：荧光定量分析(^aP＜0.05，vs PM2.5；^bP＜0.05，vs PM2.5+NAC)

Figure 2. Measurement of autophagy by fluorescent microscopy

A: DAPI staining of nuclei (blue) and Phalloidin staining of cytoskeleton (red) were set as the background stain, and LC3 fluorescence was green; B: Quantification analysis (^aP＜0.05, vs PM2.5; ^bP＜0.05, vs PM2.5+NAC)

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图 3 LC3Ⅰ、LC3Ⅱ、P62、GAPDH定量分析(A。^aP＜0.05，vs PM2.5；^bP＜0.05，vs PM2.5+NAC)及化学发光结果(B)

Figure 3. Quantification analysis (A. ^aP＜0.05, vs PM2.5; ^bP＜0.05, vs PM2.5+NAC) and chemiluminescence results of LC3Ⅰ, LC3Ⅱ, P62 (B)

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