Guan Shi-He, Lu Mengji, Grünewald Petra, Roggendorf Michael, Gerken Guido, Schlaak Jörg F
Department of Gastroenterology and Hepatology, University Hospital of Essen, Hufelandstr. 55, Essen 45122, Germany.
World J Gastroenterol. 2007 Jan 14;13(2):228-35. doi: 10.3748/wjg.v13.i2.228.
To characterize the IFN-response and its modulation by the antiviral compound lamivudine in HBV-transfected HepG2.2.15 cells.
HepG2.2.15 and HepG2 cells were stimulated with various concentrations of IFN-alpha2a in the presence or absence of lamivudine. Then, total RNA was extracted and analysed by customised cDNA arrays and northern blot for interferon-inducible genes (ISGs). In addition, cellular proteins were extracted for EMSA and western blot. HBV replication was assessed by southern blot or ELISAs for HBsAg and HBeAg.
Two genes (MxA, Cig5) with completely abolished and 4 genes (IFITM1, -2, -3, and 6-16) with partially reduced IFN-responses were identified in HepG2.2.15 cells. In 2 genes (IFITM1, 6-16), the response to IFN-alpha could be restored by treatment with lamivudine. This effect could not be explained by a direct modulation of the Jak/Stat signalling pathway since EMSA and western blot experiments revealed no suppression of Stat1 activation and ISGF3 formation after stimulation with IFN-alpha in HepG2.2.15 compared to HepG2 cells.
These results are consistent with the assumption that chronic hepatitis B may specifically modulate the cellular response to IFN by a selective blockage of some ISGs. Antiviral treatment with lamivudine may partially restore ISG expression by reducing HBV gene expression and replication.
在转染乙肝病毒的HepG2.2.15细胞中，对干扰素反应及其受抗病毒化合物拉米夫定的调节作用进行特性分析。
在有或无拉米夫定存在的情况下，用不同浓度的干扰素α2a刺激HepG2.2.15和HepG2细胞。然后，提取总RNA，通过定制的cDNA阵列和针对干扰素诱导基因（ISG）的Northern印迹法进行分析。此外，提取细胞蛋白用于电泳迁移率变动分析（EMSA）和蛋白质免疫印迹法。通过Southern印迹法或针对乙肝表面抗原（HBsAg）和乙肝e抗原（HBeAg）的酶联免疫吸附测定（ELISA）评估乙肝病毒复制情况。
在HepG2.2.15细胞中鉴定出两个干扰素反应完全消失的基因（MxA、Cig5）和四个干扰素反应部分降低的基因（IFITM1、-2、-3和6-16）。在两个基因（IFITM1、6-16）中，拉米夫定处理可恢复对干扰素α的反应。由于EMSA和蛋白质免疫印迹实验显示，与HepG2细胞相比，HepG2.2.15细胞在用干扰素α刺激后，Stat1激活和ISGF3形成未受抑制，因此这种效应无法通过Jak/Stat信号通路的直接调节来解释。
这些结果与慢性乙型肝炎可能通过选择性阻断某些ISG来特异性调节细胞对干扰素反应的假设一致。拉米夫定抗病毒治疗可能通过降低乙肝病毒基因表达和复制来部分恢复ISG表达。
