干扰素λ1对Huh7细胞自噬相关基因ATG3、ATG5及Bcl-xl表达的影响

The effects of IFN-λ1 on the expression of autophagy-releated genes ATG3,ATG5 and Bcl-xl in Huh7 Cells

目的研究干扰素λ1(interferonλ1,IFN-λ1)对肝细胞内自噬相关基因表达的影响,探讨IFN-λ1抗HCV的新机制。方法感染HCV JFH-1毒株的Huh7细胞加入IFN-λ1处理,采用实时荧光逆转录定量PCR法分析细胞自噬相关基因LC3、Beclin、ATG3、ATG5、ATG7、Bcl-xl、Bad mRNA表达以及HCV RNA水平的变化,采用Western Blot检测自噬相关基因和HCV Core在蛋白水平的变化。结果 IFN-λ1抑制HCV在Huh7细胞中的复制,与对照组相比,IFN-λ1可以下调HCV RNA的水平(t=-4.513,P=0.011)和HCV Core蛋白相对表达量(t=6.943,P=0.020);IFN-λ1可下调细胞自噬基因ATG3、ATG5的mRNA表达(t=10.311,P=0.007;t=-14.246,P=0.004),同时上调细胞自噬抑制因子Bcl-xl的mRNA表达(t=-5.246,P=0.034)。IFN-λ1对ATG3、ATG5的蛋白表达同样具有抑制作用(t=6.668,P=0.022;t=13.343,P=0.006)。结论干扰素λ1可能通过抑制肝细胞内自噬基因ATG3、ATG5的表达以及上调细胞自噬抑制因子Bcl-xl的表达,破坏细胞自噬过程,从而达到抗HCV的作用。

Objective This study aims to investigate the effects of interferon -λ1 （IFN-λ1） on the expression of autophagy-related genes in Huh7 cells and illuminate novel mechanism（s） invovled in anti-HCV activity of IFN-λ1. Meth- ods HCV JFH-1-infected Huh7 cells were treated by IFN-λ1. The expression of autophagy-related genes LC3, Beclin, ATG3, ATGS, ATG7, Bcl-xl, Bad at mRNA level in Huh7 cells and the levels of HCV RNA were determined by RT- PCR. The protein expression of cellular autophagy-related genes and HCV Core were determined by Western blotting. Re- suits IFN-λ1 could significantly inhibit HCV replication in Huh7 cells. Compared to control group, IFN-λ1 treatment sig- nificantly decreased the level of HCV RNA （t = -4. 513 ,P =0. 011 ） and the relative expression of HCV Core protein （ t = 6. 943 ,P =0. 020） ; IFN-λ1 could also decrease the mRNA expression of cellular autophagy genes ATG3, ATG5 （ t = 10. 311, P =0. 007 ; t = - 14. 246, P =0. 004）, and significantly increase the mRNA expression of Bcl-xl, which is a inh- biroty factor of cellular autophagy （ t = - 5. 246, P = 0. 034 ）. The Western blotting examination at protein level confirmed the inhibitory effects of IFN-λ1 on ATG3, ATG5 （t=6.668,P=0.022; t=13.343,P=0.006）. Conclusions IFN-λ1 could inhibit the expression of autophagy-related genes ATG3, ATG5 and increase expression of Bcl-xl, an inhbiroty factor of cellular autophagy in human hepatocytes, leading to interrupting cellular autophagy and inhibiting HCV infection and rep- lication.