TRIM28磷酸化对人肺上皮细胞高致病性禽流感病毒感染过程中IFN-β和促炎细胞因子的表达

TRIM28 phosphorylation enhances the expression of IFN-β and inflammatory cytokines in human lung epithelial cells infected with highly pathogenic avian influenza virus

目的探究TRIM28磷酸化对人肺上皮细胞高致病性禽流感病毒感染过程中IFN-β和促炎细胞因子表达的影响。方法以人肺癌上皮细胞(human lung carcinoma cells A549,ATCC)及人肺腺癌细胞系PAa为研究对象。将细胞进行高致病性禽流感病毒ⅣAnhui/112005(H5N1)转染。将病毒转染后的细胞进行siRNA转染,对转染后的细胞的总RNA进行提取,检测各组细胞中mRNA的表达量,细胞内的蛋白表达量,细胞活性及细胞凋亡率。结果转染均siRNA2与siRNA3后能有效抑制TRIM28蛋白的磷酸化;TRIM28的磷酸化可促进IFN-β、IL-1β在细胞内的表达;与转染TRIM28 siRNA2、TRIM28 siRNA3后的对照组GAPDH和对照组scrambled siRNA中PAa细胞的G0/G1期比例(即50.02%和53.46%)相比,转染后的PAa细胞在G0/G1期的比例(60.13%和59.38%)增加(P<0.05)。但转染TRIM28 siRNA2、TRIM28 siRNA3的PAa细胞的G2/M期与S期的比例均下降。结论 TRIM28磷酸化增强了人肺上皮细胞高致病性禽流感病毒感染过程中IFN-β和促炎细胞因子的表达。

Objective To explore the effect of TRIM28 phosphorylation on the expression of IFN-beta and inflammatory cytokines in human lung epithelial cells infected with highly pathogenic avian influenza virus.Methods Human lung carcinoma cells A549(ATCC)and human lung adenocarcinoma cell line PAa were infected with highly pathogenic avian influenza virus IV Anhui/112005(H5 N1)followed by siRNA transfection.Cells were harvested and total RNA of the transfected cells was extracted.The expression of mRNA in each group was measured by real-time PCR and the proteins were determined by Western-blot.The activity of cells and the apoptotic rate of cells were determined using flow cytometry.Results The results showed that the phosphorylation of TRIM28 protein could be effectively inhibited by transfection of both siRNA 2 and siRNA 3.The percentages of G0/G1 phase of PAa cells transfected with TRIM28 siRNA 2 and TRIM28 siRNA 3(60.13%and 59.38%)were significantly higher than that of control cells transfected with GAPDH and scrambled siRNA(50.02%and 53.46%)(P<0.05),while the percentages of G2/M phase and S phase of PAa cells transfected with TRIM28 siRNA2 and TRIM28 siRNA3 decreased.Conclusions TRIM28 phosphorylation enhances the expression of IFN-βand pro-inflammatory cytokines in human lung epithelial cells infected with highly pathogenic avian influenza virus.