基于GADD153-LUC的重组人肺上皮细胞快速检测痕量PM_(2.5)致DNA损伤效应  

Rapid detection of DNA damage caused by trace PM_(2.5)using recombinant human lung epithelial cells based on GADD153-LUC

作　　者：姚炜 詹劲基 姚云英 汪东篱 李燕 杜海荣 YAO Wei;ZHAN Jin-ji;YAO Yun-ying(Guangming District Center For Disease Control and Prevention of Shenzheng,Guangdong Shenzheng 518106)

机构地区：[1]深圳市光明区疾病预防控制中心,广东深圳518106

出　　处：《医学检验与临床》2024年第11期1-5,共5页Medical Laboratory Science and Clinics

基　　金：深圳市科技计划基金资助项目,项目名称:基于GADD基因的快速筛选系统检测PM_(2.5)对细胞DNA损伤效应研究,项目编号:JCYJ20180301170716407。

摘　　要：目的:利用重组人体肺上皮细胞(BEAS-2B)快速检测细颗粒物(PM_(2.5))导致的DNA损伤效应。方法:制备不同浓度的PM_(2.5)并作用于重组肺上皮细胞,观察不同浓度条件下MTS(内盐法,3-(4,5-二甲基噻唑-2)-5-(3-羧甲基苯基)-2-(4-磺苯基)-2氢-四唑内盐法)检测重组人肺上皮细胞细胞生存率、彗星实验检测DNA损伤、RT-PCR检测GADD153-luc基因表达变化、荧光素酶发光检测DNA损伤情况。结果:相比于阴性对照,25、125μg/mLPM_(2.5)可诱导重组细胞活性显著下降和DNA损伤显著增加,相比于阴性对照,5、25、125μg/mL PM_(2.5)可诱导重组细胞内源性GADD153mRNA基因表达、荧光素酶活性显著增加,且GADD153mRNA基因表达、荧光素酶活性具有良好的相关性(r^(2)=0.992,P<0.01)。结论:相比传统细胞DNA损伤的方法,基于GADD153-LUC的重组人肺上皮细胞可快速检测更痕量PM_(2.5)所致DNA损伤效应。Objective:Rapid detection of DNA damage caused by trace fine particulate matter(PM_(2.5))using recombinant human lung epithelial cells(BEAS-2B).Methods:Different concentrations of PM_(2.5)were prepared and applied to recombinant lung epithelial cells.The survival rate of recombinant human lung epithelial cells was detected by MTS(Internal salt method,3-(4,5-dimethylthiazole-2)-5-(3-carboxymethylphenyl)-2-(4-sulfophenyl)-2H-tetrazole internal salt method),DNA damage was detected by comet assay,GADD153 mRNA gene expression was detected by RT-PCR,and DNA damage was detected by luciferase activity under different concentrations.Results:Compared with negative control,25,125μg/mL PM_(2.5)significantly decreased the activity of recombinant cells and significantly increased the DNA damage(comet assay).Compared with negative control,5,25,125μg/mL PM_(2.5)significantly increased the endogenous GADD153 mRNA gene expression and luciferase activity of recombinant cells.GADD153 mRNA�gene expression and luciferase activity had a good correlation(r^(2)=0.992,P<0.01).Conclusion:Compared with the traditional methods for DNA damage in cells,recombinant human lung epithelial cells based on GADD153-LUC can rapidly detect DNA damage caused by lower dose PM_(2.5).

关 键 词：细颗粒物 人肺上皮细胞 DNA损伤 Gadd153-luc 

分 类 号：R446[医药卫生—诊断学]