PM2.5对L02肝细胞部分癌基因和凋亡相关基因表达的影响

PM2.5 exposure on expression of oncogenes and apoptosis genes in hepatocytes

目的:探讨细颗粒物(PM2.5)对人正常肝细胞(L02肝细胞)癌基因和凋亡相关基因表达的影响。方法:以L02肝细胞为研究对象,设10和50μg/mL两个PM2.5混悬液染毒组,以未染毒的L02肝细胞为阴性对照,10μmol/L的K2CrO4(下用Cr^6+表示)染毒细胞为阳性对照,均处理24 h,应用实时荧光定量聚合酶链式反应(qPCR)和Western blot技术分别检测促癌基因c-myc、c-fos、抑癌基因p53和促凋亡相关基因Caspase-3、Caspase-8,抑凋亡基因Bcl-2的mRNA以及相应的蛋白表达变化。结果:qPCR法分析结果显示,与阴性对照组比较,10、50μg/mL PM2.5混悬液染毒组以及阳性对照组L02肝细胞癌基因c-myc的mRNA表达水平分别升高69.5%、118.0%、51.1%,c-fos表达水平分别升高50.3%、64.4%、23.3%,p53表达水平分别下降4.0%、22.0%、18.0%;凋亡相关基因Caspase-3的mRNA表达水平分别升高31.0%、30.5%、42.0%,Caspase-8表达分别升高25.3%、40.3%、64.5%,Bcl-2表达分别下降40.5%、46.9%、41.4%,差异均有统计学意义(P<0.05)。Western blot法分析结果显示,在10、50μg/mL PM2.5混悬液和Cr6+染毒后L02肝细胞c-myc蛋白表达水平分别升高32.3%、49.3%、70.6%,c-fos蛋白表达水平分别升高11.3%、50.2%、45.2%,p53蛋白表达水平分别下降17.5%、40.0%、42.9%;Caspase-3蛋白表达水平分别升高23.1%、33.9%、43.1%,Caspase-8蛋白表达水平分别升高31.4%、52.1%、80.0%,Bcl-2蛋白表达水平分别下降14.3%、23.3%、36.9%,差异均有统计学意义(P<0.05)。结论:PM2.5染毒引起L02肝细胞中促癌基因和促凋亡相关基因表达升高,抑癌基因和抑凋亡相关基因表达下降,提示PM2.5对L02肝细胞的恶性转化和凋亡可能有促进与激活作用。

OBJECTIVE:To study the effect of particulate matter 2.5(PM2.5)exposure on expression of oncogenes and apoptosis-related genes in normal hepatocytes(L02 cells).METHODS:L02 cells were exposed to PM2.5(10 and 50μg/mL),as the experimental group,to 10μmol/L Cr^6+as the positive control group,and nothing as the negative control group,for 24 h.After the treatment,mRNA expression of oncogenes including promoting oncogenes including c-myc,c-fos and suppressing oncogenes p53 and promoting apoptosis-related genes including Caspase-3,Caspase-8 and suppressing apoptosis gene Bcl-2 were detected by fluorescent quantitative real-time PCR(qPCR),the protein expression of oncogenes and apoptosis genes were detected with Western blot.RESULTS:The qPCR showed that compared with the negative control group,significantly different mRNA expressions in the PM2.510μg/mL,50μg/mL and positive control groups were:c-myc gene increased by 69.5%,118.0%,51.1%,c-fos gene increased by 50.3%,64.4%,23.3%;p53 gene decreased by 4.0%,22.0%,18.0%;Caspase-3 gene increased by 31.0%,30.5%and 42.0%;Caspase-8 expression level increased by 25.3%,40.3%and 64.5%,Bcl-2 gene expression level decreased by 40.5%,46.9%and 41.4%,respectively(P<0.05).Western blot analyses showed that expression levels of c-myc protein increased by 32.2%,49.3%,70.6%;c-fos protein increased by 11.3%,50.2%,45.2%;p53 protein decreased by 17.5%,40.0%,42.5%;Caspase-3 protein increased by 23.1%,33.9%,43.1%;Caspase-8 protein increased by 31.4%,52.1%,80.0%;Bcl-2 protein decreased by 14.3%,23.3%,36.9%,respectively,(P<0.05).CONCLUSION:PM2.5 exposure increased expression of promoting oncogenes and promoting apoptosis genes as well as decreased expression of suppressing oncogenes and suppressing apoptosis genes in L02 hepatocytes.The results indicate that PM2.5 exposure can promote and activate malignant transformation and apoptosis of L02 hepatocytes.