旨在探究组蛋白去乙酰化酶(HDACs)选择性抑制剂MGCD0103(Mocetinostat)对小反刍兽疫病毒(peste des petits ruminants virus,PPRV)在山羊子宫内膜上皮细胞(EEC细胞)复制的影响.本研究通过RT-qPCR法测定PPRV感染后宿主细胞HDACs和抑制剂处理细胞中PPRV N基因的转录水平,通过Time of Addition Assay确定MGCD0103在PPRV复制过程的作用阶段,用Western blot检测PPRV N蛋白的相对表达量,并采用TCID56方法测定了病毒滴度.结果表明,PPRV感染引起EEC细胞HDAC1(P＜0.001)和HDAC2(P＜0.002)转录水平明显上升;MGCD0103在病毒入侵和复制阶段均发挥了抑制作用,且在复制阶段抑制作用更明显;MGCD0103显著降低PPRV N基因转录水平和表达水平及病毒滴度(P＜0.002),且其抑制PPRV在EEC中的半数有效浓度(EC50)和药物选择指数(SI)分别是0.43 μmol·L-1和4.35.本研究确定MGCD0103显著抑制PPRV的复制,这对研发有效抗PPRV药物具有重要意义,为高效率产毒细胞株的构建提供新思路.
Abstract
In this study,the effect of MGCD0103(Mocetinostat),a selective HDACi,on the rep-lication of peste des petits ruminants virus(PPRV)in goat endometrial epithelial cells(EEC cells)was investigated,aiming to clarify the role and activity of MGCD0103 in PPRV multiplica-tion.The transcription levels of HDACs and PPRV N gene in different treated cells were deter-mined by RT-qPCR,and the action stage of MGCD0103 in the viral proliferation cycle of PPRV was determined by the time of addition assay.The effect of MGCD0103 on the expression level of PPRV N protein and viral titer were further analyzed by Western blot and TCID50,respectively.The results showed that the mRNA expression levels of HDAC1(P＜0.001)and HDAC2(P＜0.002)were increased significantly in PPRV-infected EEC cells.Although MGCD0103 has an ex-cellent inhibitory effect on PPRV multiplication,its inhibitory effect in the virus entry is limited.MGCD0103 significantly reduced the transcription and expression levels of PPRV N gene,as well as the viral titer(P＜0.002).The half-maximal effective concentration(EC50)and selectivity in-dex(SI)of MGCD0103 against PPRV were 0.43 μmol·L-1 and 4.35,respectively.These results indicated that MGCD0103 significantly inhibited PPRV replication,which is of great significance for the development of anti-PPRV drugs and provides a new idea for constructing sensitive cell lines for PPRV proliferation.
关键词
组蛋白去乙酰化酶抑制剂/MGCD0103/抑制/小反刍兽疫病毒/病毒复制
Key words
histone deacetylase inhibitor/MGCD0103/inhibit/peste des petits ruminants virus/replication
维普
万方数据