温石棉对人支气管上皮细胞BEAS-2B凋亡的影响  

Effects of Chrysotile Fibers on Apoptosis of Human Bronchial Epithelial Cell Line (BEAS-2B) in vitro

作　　者：张敏[1] 陈钧强[1] 贾振宇[1] 周华仕[1] 张幸[1] 

机构地区：[1]浙江省医学科学院卫生学研究所,浙江杭州310013

出　　处：《环境与职业医学》2009年第2期155-158,共4页Journal of Environmental and Occupational Medicine

基　　金：浙江省科技厅重大与高发疾病防治技术专项(编号:2008C13029-2)

摘　　要：[目的]研究温石棉对永生化人支气管上皮细胞(BEAS-2B)凋亡的影响,比较单细胞凝胶电泳试验(SCGE)与流式细胞术(FCM)检测细胞凋亡结果的异同。[方法]选用BEAS-2B细胞为受试物,以5、10、20、40、100、200μg/cm2不同剂量的温石棉,分别刺激BEAS-2B细胞24、48、72h,阳性对照用石英(SiO2),阴性对照用硅灰石(WO1),用四甲基偶氮唑盐(MTT)比色法检测温石棉对BEAS-2B活力的影响。用终剂量10μg/cm2温石棉、5μg/cm2SiO2、20μg/cm2WO1分别刺激细胞24、48和72h后,采用SCGE和FCM检测温石棉对BEAS-2B凋亡指数和凋亡率的影响。采用逆转录聚合酶链反应(RT-PCR)技术检测凋亡相关半胱氨酸蛋白酶-3基因caspase-3、caspase-9的表达。[结果]10μg/cm2温石棉可诱导BEAS-2B细胞凋亡,呈时间-效应关系(P<0.05),采用SCGE和FCM方法检测细胞凋亡率结果一致。温石棉刺激BEAS-2B细胞24、48、72h后凋亡指数为11.7%、21.8%和34.5%,凋亡率为9.6%、17.9%和31.2%;温石棉诱导细胞caspase-3、caspase-9表达水平随时间增加而增加。[结论]温石棉可诱导BEAS-2B凋亡;SCGE和FCM均可以灵敏、快捷地检测温石棉对BEAS-2B的凋亡作用。[Objective] To investigate the influence on apoptosis of human bronchial epithelial cell line （ BEAS-2B ） induced by chrysotile fibers and to compare the methods of single cell gel electrophoresis assay（ SCGE ）with flow cytometry analysis （FCM）for detecting apoptosis. [ Mothods ] BEAS-2B cell line was cultured together with 5, 10, 20, 40, 100, and 200μg/cm^2 chrysotile fibers separately for 24, 48, and 72h, using 5μg/cm^2 quartz（ SiO2 ）as postive control and 20μg/cm^2wollastonite（ WO1 ） as negtive control. The viability of BEAS-2B cells was detected by MTr assay, and the apoptotic effects were observed by SCGE and FCM. The mRNA levels of caspases-3 and caspases-9 were determined using reverse transtription polymerase chain reaction （ RT-PCR ）. [ Results ]10μg/cm^2 ehrysotile fibers could induce a time-dependent apoptosis in BEAS-2B cells. The results of SCGE and FCM were consisitent. The apoptosis indices were 11.7%, 21.8% and 34.5% and apoptosis rate were 9.6%, 17.9% and 31.2% by stimulating BEAS-2B for 24, 48, and 72h respectively. The mRNA level of caspases-3 and caspases-9 increased with time in BEAS-2B cells treated with chrysotile fibers. [ Conclusion ] Chrysotile fibers could induce apoptosis of BEAS-2B cells. Both SCGE and FCM could be two good methods to detect apoptotic effects induced by ehrysotile fibers.

关 键 词：温石棉 人支气管上皮细胞 凋亡 单细胞凝胶电泳试验 流式细胞术 

分 类 号：R114[医药卫生—卫生毒理学]