泛素特异性蛋白酶18抑制IFNα抗HBV活性但并不抑制IFNλ抗HBV的活性

干扰素α（IFN-α）是临床最常用的抗乙型肝炎病毒（HBV）药物之一。泛素特异性蛋白酶18（USP18）被证实是抑制IFN-α抗HBV活性的因子，但USP18是否对干扰素λ(IFN-λ)抗HBV有影响还尚未可知。为了明确USP18对IFNλ抗HBV活性的影响，本研究以HepG2.2.15细胞作为乙肝体外模型，采用脂质体转染法分别向细胞转染pEGFP-USP18、PEGFP-N1经 48 h，再经IFN-α和IFN-λ处理24 h，分为阴性对照组﹑USP18过表达+IFN-α组﹑空载组+IFN-α组﹑USP18过表达+IFN-λ组﹑空载组+IFN-λ组。采用Western印迹、RT-qPCR和ELISA检测各组的乙肝病毒标志物、STAT1/pSTAT1和下游的干扰素刺激基因(ISGs)的表达。结果显示，与阴性对照组和空载组相比，USP18蛋白在过表达组明显升高（P<0.05），过表达细胞模型构建成功;在IFN-α处理的两组中，空载组中HBsAg、HBeAg、HBcAg及HBV-DNA的表达均低于USP18过表达组，差异有统计学意义（P<0.05）。而IFN-λ处理组中，乙肝病毒标志物的差异不明显。在IFN-α处理组中，空载组的ISG15、MxA、IFIT1和pSTAT1表达均高于USP18过表达组，差异有统计学意义（P<0.05），而在IFN-λ处理组中ISGs和 pSTAT1的表达无明显差异。上述结果证实，USP18可通过抑制JAK/STAT信号通路的激活来减弱IFN-α抗HBV的活性。研究还证实，IFN-λ可发挥抗HBV的作用，USP18不通过JAK/STAT信号通路抑制其抗HBV活性。

Ubiquitin Specific Protease 18 Inhibits the Anti-Hepatitis B Virus Activity of Interferon-Alpha,but Does Not Inhibit Interferon-Lambda in HepG2.2.15 Cells

Interferon alpha (IFN-α) is one of the most widely used anti-hepatitis B virus (HBV) medicine. It has been reported that ubiquitin specific protease 18 (USP18) inhibits the anti-HBV activity of IFN-α, but whether USP18 has an effect on the anti-HBV activity of IFN-λ was unclear. Here we aim to detect the anti-HBV effect of USP18 on IFN-λ in an in vitro model. HepG2.2.15 cells were transfected with two different plasmids: the empty vector (pEGFP-N1) and USP18 overexpressed plasmid (pEGFP-USP18), and then were treated with IFN-α and IFN-λ for 24 hours, respectively. Untreated groups served as the negative control. The expression of HBV markers, STAT1/pSTAT1 protein expression and interferon stimulated genes(ISGs) were tested on HepG2.2.15 by Western blotting, quantitative real-time PCR (RT-qPCR) and enzyme-linked immunosorbent assays (ELISA). The results showed that USP18 was successfully overexpressed in the overexpression group compared with both negative control and empty vector control groups (P<0.05), which demonstrated the successful establishment of vitro models. In IFN-α treated groups, the expression of hepatitis B surface antigen (HBsAg), hepatitis B e antigen (HBeAg), hepatitis B core antigen (HBcAg) and HBV-DNA in overexpression groups were significantly higher than empty vectors (P< 0.05), while there was little difference in the IFN-λ treatment group. In addition, the expression of ISG15, MxA, IFIT1 and pSTAT1 of the empty vector were obviously higher than those in overexpression groups in the treatment group of IFN-α (P<0.05), but there was also no significant difference in ISGs and pSTAT1expression in IFN-λ treatment groups. USP18 inhibits the anti-HBV activity of IFN-α by suppressing the activation of the JAK/STAT signaling pathway. In contrast, USP18 has no effect on the anti-HBV activity of IFN-λ via the JAK/STAT signaling pathway.