六价铬通过lncRNA TERC对BEAS-2B细胞KLHL20与EHBP1L基因表达的影响

Effect of hexavalent chromium on KLHL20 and EHBP1L gene expression in BEAS-2B cells through lncRNA TERC

目的探究六价铬暴露对BEAS-2B细胞长链非编码RNA TERC及下游靶基因KLHL20和EHBP1L基因表达的影响,为六价铬致癌机制研究提供新思路。方法采用CCK8法检测不同浓度六价铬(K2Cr2O7)对BEAS-2B细胞活力的影响,选择不同浓度六价铬处理细胞24、48和72 h,或连续染毒7 d,采用实时荧光定量PCR法(RT-q PCR法)检测lncRNA TERC及其靶基因(KLHL20和EHBP1L1)表达水平变化。通过转染实验敲减或过表达BEAS-2B细胞lncRNA TERC后,选用1.00μmol/L六价铬染毒24 h,检测lncRNA TERC及其靶基因表达水平的变化。结果染毒24、48、72 h,六价铬处理组的细胞存活率随着染毒浓度的升高而逐渐降低(F=7.88、340.26、655.78,P<0.05)。不同浓度六价铬处理细胞24 h,lncRNA TERC与其靶基因KLHL20和EHBP1L1的表达水平随着六价铬浓度的升高而升高,呈现一定的浓度-效应关系(F=28.99、10.82、39.11,P<0.05)。0.50μmol/L六价铬处理BEAS-2B细胞1、3、5、7 d,lncRNA TERC的表达水平随着染毒时间的增加而升高(F=7.16,P<0.05),EHBP1L1的转录水平先升高后降低(F=37.34,P<0.05)。敲减lncRNA TERC后,KLHL20和EHBP1L1的表达水平为(0.90±0.01,0.83±0.01)低于对照组(1.00±0.04,1.00±0.05,F=176.40,74.40,P<0.05),而过表达lncRNA TERC后,KLHL20和EHBP1L1的表达水平为(1.20±0.06,1.67±0.15)均高于对照组(1.00±0.04,1.00±0.05,F=176.40、74.40,P<0.05)。结论六价铬可能通过lncRNA TERC调控BEAS-2B细胞下游靶基因KLHL20和EHBP1L1的表达。

Objective To investigate the effect of hexavalent chromium exposure on the expression of long noncoding RNA TERC and downstream target genes KLHL20 and EHBP1L in BEAS-2B cells,and to provide new ideas for the study of the carcinogenic mechanism of hexavalent chromium.Methods CCK8 method was used to detect the effect of different concentrations of hexavalent chromium(K_2Cr_2O_7)on the viability of BEAS-2B cells.The cells were treated with different concentrations of hexavalent chromium for 24,48 and 72 h,or continuously for 7 days,and the expression levels of lncRNA TERC and its target genes(KLHL20 and EHBP1L1)were detected by real-time fluorescence quantitative PCR(RT-q PCR).After knocking down or overexpressing lncRNA TERC in BEAS-2B cells by transfection experiment,1.00μmol/L hexavalent chromium was used to treat for 24 h to detect the changes in the expression levels of lncRNA TERC and its target genes.Results The cell survival rate of the hexavalent chromium-treated group decreased gradually with the increase of the hexavalent chromium concentration at 24,48,and 72 h(F=7.88,340.26,655.78,P<0.05).The expression levels of lncRNA TERC and its target genes KLHL20 and EHBP1L1 increased with the increase of hexavalent chromium concentration after the cells were treated with different hexavalent chromium concentrations for 24 h,showing a certain concentration-effect relationship(F=28.99,10.82,39.11,P<0.05).When BEAS-2B cells were treated with 0.50μmol/L hexavalent chromium for 1,3,5,and 7 days,the expression level of lncRNA TERC increased with the increase of the exposure time(F=7.16,P<0.05),and the transcription level of EHBP1L1 increased first and then decreased(F=37.34,P<0.05).After knocking down lncRNA TERC,the expression levels of KLHL20 and EHBP1L1 were(0.90±0.01,0.83±0.01)lower than those of the control group(1.00±0.04,1.00±0.05,F=176.40,74.40,P<0.05),while after overexpressing lncRNA TERC,the expression levels of KLHL20 and EHBP1L1 were(1.20±0.06,1.67±0.15)higher than those of the control group(1.00±0.04,1.00±0.05,F=176.40,74.40,P<0.05).Conclusion Hexavalent chromium may regulate the expression of downstream target genes KLHL20 and EHBP1L1 in BEAS-2B cells through lncRNA TERC.