慢性丙型肝炎患者外周血单个核细胞干扰素及相关基因表达与干扰素疗效关系的研究
|
摘要
慢性丙型肝炎患者外周血单个核细胞干扰素及相关基因表达与干扰素疗效关系的研究
丙型肝炎病毒(Hepatitis C Virus，HCV)感染引起的丙型肝炎仍是严重危害人民健康的重要病毒性疾病。丙型肝炎的主要危害性在于患者易转变为慢性肝炎，并可进一步发展为肝癌(HCC)而死亡。目前Ⅰ型干扰素(下简称干扰素)已被临床广泛用于慢性丙型肝炎的治疗，但尚存在治疗效果有限等缺点。虽然国内外对HCV基因结构、功能、致病性及机体免疫应答等已进行了大量研究，但迄今仍不十分清楚HCV与干扰素在机体和细胞内如何相互作用以影响机体对干扰素的应答。本室曾对丙型肝炎病毒的有关基因序列进行研究，结果未发现相关氨基酸序列与中国丙型肝炎病人的干扰素疗效有关。因此，有必要进一步探讨机体因素与干扰素疗效的相关性。
HCV感染机体后可诱生机体产生内源性干扰素，同时又可抵抗干扰素的作用。当使用外源性干扰素对病人进行治疗时，机体对干扰素的应答实际上是病毒，内源性干扰素系统与外源性干扰素在体内相互作用、平衡的结果。理论推测，HCV可能通过基因复制和蛋白表达对干扰素通路上的关键环节进行干预，使内源性干扰素产生不足、信号通路受阻或干扰素的作用被拮抗，从而导致对干扰素的治疗不敏感；而由于宿主遗传背景因素差异引起的丙肝病毒感染者体内干扰素产生不足、干扰素诱导的信号转导通路缺陷或干扰素效应蛋白表达水平低下可能是其另一重要原因。
为此，本研究以34例慢性丙型肝炎(chronic hepatitis C，CHC)病人为研究对象，在接受干扰素联合利巴韦林治疗前、治疗1个月、3个月、6个月、12个月和停药6个月时分别收集外周血标本，分离血清、血浆和外周血单个核细胞(PBMCs)后冻存于-80℃一并检测。根据疗程结束时和停药6个月时血清HCV RNA的检测结果，6例病人对干扰素非持续应答(包括无应答和复发的病人，简称NR)，28例病人则对干扰素持续应答(简称SR)。在此基础上采用实时定量聚合酶链式反应(Real-time PCR)方法分别对不同干扰素疗效病人PBMCs中Ⅰ型干扰素合成、信号转导以及抗病毒效应相关的基因的表达水平进行研究，并在丙肝病毒亚基因组复制细胞系统中验证有关结果；同时用基因芯片检测不同干扰素疗效病人PBMCs中全基因表达水平，以搜寻与疗效相关的基因表达特征(signature)或模式(pattern)。
本研究首先探讨Ⅰ型干扰素及其合成相关基因的表达与HCV感染及干扰素疗效的关系。先检测了HCV慢性感染情况下PBMCs中IFN-α/β及其合成调节因子(IRF-1，IRF-3，IRF-5和IRF-7)的表达。结果发现，IFN-α和IFN-β的表达水平在病人和正常人(donor)之间没有显著的差异(P值分别为0.927和0.937)，而IRF家族成员的表达各不相同，其中IRF-3表达不受慢性感染的影响(P=0.980)，IRF-7表达在病人与正常人的差异未达到显著水平(P=0.071)，但在病人中IRF-1表达水平明显增强(P＜0.001)，IRF-5表达水平却显著低于正常人(P=0.002)。基因表达相关性分析显示，IRF-1的表达与IFN-α/β的表达无相关性；IRF-7的表达与IFN-α的表达显著相关(相关系数0.453；P值0.007)；IRF-5表达与IFN-α和IFN-β的表达则高度相关(相关系数0.686，0.621；P值＜0.001，0.001)。进一步分析Ⅰ型干扰素及其合成相关基因的表达与干扰素疗效的关系，发现上述相关基因在治疗前的表达水平与干扰素疗效无相关性，不能用于干扰素疗效的预测。为了确认HCV持续感染对IRF和Ⅰ型干扰素的影响，进一步在无HCV复制的Huh7细胞和HCV持续复制的Replicon细胞中研究IRF-7和IFN-α的表达水平。结果发现，在NDV攻击下，Replicon细胞的IRF-7 promoter活性、内源性IRF-7和IFN-α表达水平均较Huh7细胞低(IRF-7 promoter活性上调倍数：2.54 vs 4.20，P＜0.001；内源性IRF-7mRNA表达水平：0.137 vs 1.671，P＜0.001；内源性IFN-αmRNA表达水平：0.246 vs 2.207，P＜0.001)。但当Replicon细胞中的HCV复制被ribavirin和IFN-α降低或基本抑制后，IRF-7 promoter活性和内源性IRF-7、IFN-α表达水平均有不同程度的恢复。上述研究结果提示，在HCV慢性感染情况下，宿主Ⅰ型干扰素表达并无明显激活，部分干扰素合成调节因子的表达水平低下，可能为HCV易形成慢性感染的原因之一。此外，Ⅰ型干扰素及其合成相关基因在治疗前的表达水平与干扰素疗效无相关性，不能用于干扰素疗效的预测。
干扰素诱导细胞内抗病毒的效应有赖于细胞内干扰素信号通路的效率，为此本研究进一步探讨了HCV慢性感染对病人PBMCs中干扰素信号通路正负调节基因表达的影响，并分析正负调节基因的平衡与干扰素疗效的关系。结果发现，HCV慢性感染者与正常人相比，JAK-STAT信号通路的正向调节因子STAT1和P48的表达和负向调节因子SOCS1和SOCS3的表达均显著增强(STAT1：CHC vs Donors 96.5 vs 69，P=0.039；P48：CHC vs Donors 121.5 vs 66.9，P=0.013；SOCS1：CHC vs Donors 5.96 vs 2.13，P=0.005；SOCS3：CHC vs Donors，78.3 vs 18.9，P＜0.001)；干扰素治疗前的SOCS1和SOCS5表达水平与HCV的复制水平呈负相关(R：-0.427和-0.385；P：0.013和0.027)。鉴于SOCS1和SOCS5主要表达于Th1细胞中，以上结果提示HCV复制可能会抑制Th1免疫的形成。进一步分析JAK-STAT信号通路正负调节因子的基因表达水平与干扰素疗效的相关性，发现治疗前负调因子SOCS3的高表达与干扰素的疗效不佳有关(NR vs SR，203.0 vs 64.1，P=0.007)，推测SOCS3的高表达削弱了细胞对外源性干扰素的反应能力。鉴于SOCS3主要在Th2细胞中表达，非持续应答病人治疗前的PBMCs中SOCS3的高水平表达也提示在Th2类免疫状态下机体对干扰素反应低下。利用流式细胞仪分析应答病人在干扰素治疗前后PBMCs中γ-干扰素和IL-4分泌细胞的水平，结果与SOCS的水平相一致：在干扰素治疗过程中，随着血清中HCV RNA的下降，SOCS1和SOCS5表达上调而SOCS3表达下调，Th1和Th2细胞百分含量也分别呈现增加和减少(Th1：治疗前vs治疗3个月，4.28+2.4％vs 7.63±3.5％P＜0.001；Th2：治疗前vs治疗3个月，0.66±0.52％vs 0.37±0.53％，P=0.013)，提示其可用于干扰素免疫调节效果的监测。进一步在体外肝细胞系中确认HCV蛋白表达和基因复制对肝细胞内SOCS1和SOCS3表达的影响，发现HCV编码的Core蛋白可诱导Huh7细胞表达SOCS1和SOCS3，尤其以SOCS1表达最为显著，其它HCV的非结构蛋白则无此诱导能力。
据于外源性干扰素治疗丙型肝炎的效果主要取决于能否诱导足够的抗病毒效应分子的产生，本研究进一步检测了抗病毒效应分子MxA，PKR和2’-5’OAS在干扰素治疗前及治疗过程中的表达，并分析其基因表达与疗效的关系。结果发现，病人PBMCs中MxA，PKR和2’5’OAS的表达在干扰素治疗前与对照相比已有明显增强(MxA：CHC vs Donors 141.4 vs 5.5，P＜0.001；PKR：CHC vs Donors 75.7 vs 12.3，P＜0.001；2’-5’OAS：CHC vs Donors 72.6 vs 4.18，P＜0.001)，干扰素治疗过程中其表达水平进一步升高，但其治疗前的基础表达水平和治疗中的诱导表达水平均与干扰素疗效无关，提示MxA，PKR和2’-5’OAS的基因表达不能用于疗效的预测。进一步用基因芯片(U-133A，Affymetrix公司)的方法检测了干扰素持续应答者和非持续应答者的PBMCs在体外用干扰素诱导后的全基因表达，发现不同疗效组的基因表达大部分相似，仅有16个基因的诱导表达倍数在不同疗效组中有显著性差异，包括已知的干扰素刺激基因(STAT1，GBP1，IFIT1，GIP3和IFIT2)，趋化因子基因(CXCL10和CCL8)，信号通路相关蛋白基因(STK3，RASGRP3，RIN2和LEPR)，代谢相关蛋白基因(APOBEC3A，MANEA和ZC3HAV1)和功能尚不明确的基因(REC8L1和C6orf62)。
综上所述，本研究的结果表明，HCV慢性感染对PBMCs中Ⅰ型干扰素及其
相关基因的表达产生影响。HCV慢性感染情况下，Ⅰ型干扰素及其合成关键因子的表达受到抑制，治疗前的表达水平与疗效无关；干扰素信号通路的正负调节因子的表达均被增强，并且负调因子SOCS3的表达水平与疗效相关；抗病毒效应分子MxA，PKR和2’-5’OAS的基础表达增强，基础表达和诱导表达水平均与疗效无关，同时基因组学研究发现PBMCs中16个基因在干扰素诱导下的表达变化与疗效相关。HCV慢性感染干预宿主细胞基因表达的有关机制有待进一步利用细胞系统进行深入研究；宿主基因表达与临床干扰素疗效的相关性研究及其在指导临床用药等方面的应用，需要在更大的病人样本中探索和实践。本课题的研究结果一方面能丰富人们对干扰素诱导抗病毒及HCV拮抗干扰素机制的认识，另一方面可指导临床预测疗效、筛选病人和制定治疗方案，最终实现个性化的干扰素治疗，达到提高丙型肝炎治愈率的目的。
Hepatitis C virus (HCV) is estimated to infect more than 170 million people worldwide. HCV infection usually causes chronic hepatitis in approximately 80% of infected individuals with 20% progressing to cirrhosis and 1-2% developing hepatocellular carcinoma. Currently, Interferon-α (IFN-α) is one of the most efficient drugs for the treatment of chronic hepatitis C and the standard treatment for HCV infection is IFN-α in combination with ribavirin. Unfortunately, more than 60% of patients with chronic HCV infection will experience either no control of viral replication by therapy (nonresponders) or a relapse when therapy is stopped (relapsers). The molecular mechanisms underlying failure of IFN-α treatment are not well understood. As our previous research found no correlation between viral genomic sequence and outcome of IFN-α treatment, host factors associated with IFN therapy, were explored in this study.
It has been proposed that HCV has the ability of interfering with type I interferon (IFN-α/β) production, blocking JAK-STAT signaling transduction, inhibiting expression or activity of ISGs (interferon -stimulated genes), which may support HCV replication and persistence as well as resistance to IFN-α therapy. Host gene expression is the outcome of host reaction to infection and viral interference with host, and may correlate with efficiency of IFN-α treatment. To identify signature or pattern of host gene expression which can distinguish different outcome of therapy, expression of type I interferon, interferon regulated factors (IRFs), positive and negative regulatory factors of IFN signaling, classical antiviral effectors and whole cellular genes were analyzed in peripheral blood mononuclear cells (PBMCs) from 34 chronic hepatitis C (CHC) patients before, during and after IFN/ribavirin treatment by
real-time quantitative RT-PCR and microarray. Among them, 6 of 34 were nonresponders (NR) and 28 of 34 were sustained responders (SR) according to serum HCV RNA test at the end of treatment and 6 months post-treatment. Gene expression was compared between these two groups.
To identify the correlation between gene expression of type I interferon, IRFs and outcome of IFN therapy, gene expression level of IFN-α/β, IRF-1, IRF-3, IRF-5 and IRF-7 were quantified in CHC patients. Result showed that, IFN-α and IFN-β expression level in the patients and donors were not significantly different (P: 0.927 and 0.937). However, IRF family members were differentially expressed in the two patient groups. IRF-3 expression was not influenced by chronic infection (P=0.980) and IRF-7 expression was not significantly different between patients and donors(P=0.071); IRF-1 expression levels were obviously enhanced in patients(P<0.001) and IRF-5 expression level was remarkably lower than that of the donors (P=0.002). The relevance analysis of gene expression showed that, while IRF-1 expression and the IFN-α/β expression did not have relevance, IRF-7 expression and IFN-α expression were correlated (R= 0.453; P= 0.007) and IRF-5 expression correlated with expression of both IFN-α and IFNβ (R=0.686, 0.621; P <0.001, 0.001). This result may reflect the mechanism of low expression of host type I interferon, which might be associated with low expression of key factors that regulated interferon's synthesis. However, further analysis did not reveal any correlation between gene expression of IFN-α/β, IRF-1, IRF-3, IRF-5 and IRF-7 before treatment and responses of host to interferon treatment. In order to confirm the influence of HCV persistent infection on IRF and type I interferon, HCV replicon cell (Huh7 cell line harboring subgenomic HCV replicons) was used. Result showed that, under NDV attack, IRF-7 promoter activity or the endogenous IRF-7 and IFN-α expression level of HCV replicon cells was lower than that of Huh7 cell (fold activity of IRF-7 promoter: 2.54 vs. 4.20, P<0.001; endogenous IRF-7mRNA expression level: 0.137 vs. 1.671, P<0.001; endogenous IFN-α mRNA expression level: 0.246 vs 2.207, P<0.001). On inhibition of HCV replication by ribavirin or IFN-α treatment, both IRF-7 promoter activity and expression of endogenous IRF-7and IFN-α were restored. This result may suggest a new mechanism of inhibition of type I interferon by HCV.
As the interferon-induced intracellular antiviral effect depends largely on efficiency of signal transduction (JAK-STAT pathway), the effect of HCV chronic
infection on expression of positive and negative regulatory genes of JAK-STAT pathway in patients' PBMCs was further analyzed. Results showed that, compared to healthy donors, the expressions of positive regulatory genes (STAT1 and P48) and negative regulatory genes (SOCS1 and SOCS3) were markedly enhanced in CHC patients (STAT1: CHC vs Donors 96.5 vs. 69, P=0.039; P48: CHC vs. Donors 121.5 vs. 66.9, P=0.013; SOCS1: CHC vs. Donors 5.96 vs. 2.13, P=0.005; SOCS3: CHC vs. Donors 78.3 vs. 18.9, P<0.001). Expression level of SOCS1 and SOCS5 before treatment was negatively correlated with serum HCV RNA level (R= -0.427 and -0.385; P= 0.013 and 0.027). As SOCS1 and the SOCS5 are mainly expressed in the Thl cell, the above result indicated that HCV replication might be able to suppress the Thl immunity. Further analysis of relevance between expression of these key factors of signaling pathway and outcome of IFN therapy showed high SOCS3 expression was predictive of no response to IFN therapy (NR vs. SR 203.0 vs. 64.1, P=0.007). It indicates that SOCS3 expression level may be used as marker for prediction of IFN-α therapy. Using flow cytometer to analyze the cytokine secreting cells, we found a switch-over of cytokine synthesis from Th2 to Th1 (IFN-γ for Th1, IL-4 for Th2) (Th1: 0 week vs. 12 weeks 4.28±2.4% vs. 7.63±3.5%, P<0.01; Th2: 0 week vs. 12 weeks 0.66±0.52% vs. 0.37±0.53%, P= 0.013) during IFN-α therapy, which was paralleled to up-regulation of SOCS1 and SOCS5 level and down-regulation of SOCS3 level. This result indicated that SOCS expression level in PBMC might be used to evaluate immune regulatory effect of IFN-α treatment. In vitro assay in cell line further showed that the expression of SOCS1 and SOCS3 in Huh7 cell line were induced by HCV Core protein, but not by other HCV non-structural proteins.
As antiviral effect of interferon mainly depends on induction of antiviral effectors molecule, gene expression of classical antiviral effectors MxA, PKR and 2 ' 5 ' OAS before and during treatment and its correlation with outcome of therapy were further analyzed. Results showed that, compared with healthy donors, MxA, PKR and 2 ' 5 ' OAS expression in patients were obviously enhanced (MxA: CHC vs. Donors 141.4 vs. 5.5, P<0.001; PKR: CHC vs. Donors 75.7 vs. 12.3, P<0.001; 2'5'OAS: CHC vs. Donors 72.6 vs. 4.18, P<0.001) and were further elevated by IFN-α treatment. However, there was no correlation of both basal expression levels and fold induction during treatment of the above genes with outcome of IFN treatment, which suggest that MxA, PKR and 2'5' OAS gene expression cannot be used as predictive marker for IFN therapy. DNA microarrays(Affymetrix Com, HG U-133A, containing 13000
cellular genes and EST) were further used to analyze the global gene expression of PBMCs in NR and SR groups. Comparing the fold of induction of genes from these two groups, we found that 16 genes were differentially expressed between NR and SR. These genes included ISGs (STAT1, GBP1, IFIT1, GIP3 and IFIT2), chemokines (CXCL10 and CCL8), genes related with signaling pathway (STK3, RASGRP3, RIN2 and LEPR), metabolism-correlated gene (APOBEC3A, MANEA and ZC3HAV1) and several genes with uncertain function (REC8L1 and C6orf62).
In summary, our research results indicated that HCV chronic infection has the influence on expression of type I interferon and its related genes in PBMCs. Under HCV chronic infection situation, while expression of type I interferon and its key regulatory factor was suppressed, the expression of the positive and negative regulatory factors of interferon signaling pathway were strengthened. Furthermore, while the expression of type I interferon, its key regulatory factor and MxA, PKR, 2'5'OAS before treatment have no relationship with outcome of IFN-α treatment, there was correlation of expression of negative factor SOCS3 with outcome of IFN-α treatment. In addition, 16 cellular genes in PBMCs were differentially expressed between NR and SR. Further study with HCV replicon cells is needed to understand the molecular mechanism underlying intervention of host cell gene expression by HCV chronic infection. The possible application of host gene expression pattern in guiding clinical interferon treatment such as predicting responses to IFN treatment need to be further investigated in a larger cohort of CHC patients. These findings, on the one hand, can enrich our knowledge about interaction between HCV and host defense system; on the other hand, may help to optimize clinical treatment and improve efficacy of IFN, and finally realize the individualized interferon treatment for HCV patients.
丙型肝炎病毒(Hepatitis C Virus，HCV)感染引起的丙型肝炎仍是严重危害人民健康的重要病毒性疾病。丙型肝炎的主要危害性在于患者易转变为慢性肝炎，并可进一步发展为肝癌(HCC)而死亡。目前Ⅰ型干扰素(下简称干扰素)已被临床广泛用于慢性丙型肝炎的治疗，但尚存在治疗效果有限等缺点。虽然国内外对HCV基因结构、功能、致病性及机体免疫应答等已进行了大量研究，但迄今仍不十分清楚HCV与干扰素在机体和细胞内如何相互作用以影响机体对干扰素的应答。本室曾对丙型肝炎病毒的有关基因序列进行研究，结果未发现相关氨基酸序列与中国丙型肝炎病人的干扰素疗效有关。因此，有必要进一步探讨机体因素与干扰素疗效的相关性。
HCV感染机体后可诱生机体产生内源性干扰素，同时又可抵抗干扰素的作用。当使用外源性干扰素对病人进行治疗时，机体对干扰素的应答实际上是病毒，内源性干扰素系统与外源性干扰素在体内相互作用、平衡的结果。理论推测，HCV可能通过基因复制和蛋白表达对干扰素通路上的关键环节进行干预，使内源性干扰素产生不足、信号通路受阻或干扰素的作用被拮抗，从而导致对干扰素的治疗不敏感；而由于宿主遗传背景因素差异引起的丙肝病毒感染者体内干扰素产生不足、干扰素诱导的信号转导通路缺陷或干扰素效应蛋白表达水平低下可能是其另一重要原因。
为此，本研究以34例慢性丙型肝炎(chronic hepatitis C，CHC)病人为研究对象，在接受干扰素联合利巴韦林治疗前、治疗1个月、3个月、6个月、12个月和停药6个月时分别收集外周血标本，分离血清、血浆和外周血单个核细胞(PBMCs)后冻存于-80℃一并检测。根据疗程结束时和停药6个月时血清HCV RNA的检测结果，6例病人对干扰素非持续应答(包括无应答和复发的病人，简称NR)，28例病人则对干扰素持续应答(简称SR)。在此基础上采用实时定量聚合酶链式反应(Real-time PCR)方法分别对不同干扰素疗效病人PBMCs中Ⅰ型干扰素合成、信号转导以及抗病毒效应相关的基因的表达水平进行研究，并在丙肝病毒亚基因组复制细胞系统中验证有关结果；同时用基因芯片检测不同干扰素疗效病人PBMCs中全基因表达水平，以搜寻与疗效相关的基因表达特征(signature)或模式(pattern)。
本研究首先探讨Ⅰ型干扰素及其合成相关基因的表达与HCV感染及干扰素疗效的关系。先检测了HCV慢性感染情况下PBMCs中IFN-α/β及其合成调节因子(IRF-1，IRF-3，IRF-5和IRF-7)的表达。结果发现，IFN-α和IFN-β的表达水平在病人和正常人(donor)之间没有显著的差异(P值分别为0.927和0.937)，而IRF家族成员的表达各不相同，其中IRF-3表达不受慢性感染的影响(P=0.980)，IRF-7表达在病人与正常人的差异未达到显著水平(P=0.071)，但在病人中IRF-1表达水平明显增强(P＜0.001)，IRF-5表达水平却显著低于正常人(P=0.002)。基因表达相关性分析显示，IRF-1的表达与IFN-α/β的表达无相关性；IRF-7的表达与IFN-α的表达显著相关(相关系数0.453；P值0.007)；IRF-5表达与IFN-α和IFN-β的表达则高度相关(相关系数0.686，0.621；P值＜0.001，0.001)。进一步分析Ⅰ型干扰素及其合成相关基因的表达与干扰素疗效的关系，发现上述相关基因在治疗前的表达水平与干扰素疗效无相关性，不能用于干扰素疗效的预测。为了确认HCV持续感染对IRF和Ⅰ型干扰素的影响，进一步在无HCV复制的Huh7细胞和HCV持续复制的Replicon细胞中研究IRF-7和IFN-α的表达水平。结果发现，在NDV攻击下，Replicon细胞的IRF-7 promoter活性、内源性IRF-7和IFN-α表达水平均较Huh7细胞低(IRF-7 promoter活性上调倍数：2.54 vs 4.20，P＜0.001；内源性IRF-7mRNA表达水平：0.137 vs 1.671，P＜0.001；内源性IFN-αmRNA表达水平：0.246 vs 2.207，P＜0.001)。但当Replicon细胞中的HCV复制被ribavirin和IFN-α降低或基本抑制后，IRF-7 promoter活性和内源性IRF-7、IFN-α表达水平均有不同程度的恢复。上述研究结果提示，在HCV慢性感染情况下，宿主Ⅰ型干扰素表达并无明显激活，部分干扰素合成调节因子的表达水平低下，可能为HCV易形成慢性感染的原因之一。此外，Ⅰ型干扰素及其合成相关基因在治疗前的表达水平与干扰素疗效无相关性，不能用于干扰素疗效的预测。
干扰素诱导细胞内抗病毒的效应有赖于细胞内干扰素信号通路的效率，为此本研究进一步探讨了HCV慢性感染对病人PBMCs中干扰素信号通路正负调节基因表达的影响，并分析正负调节基因的平衡与干扰素疗效的关系。结果发现，HCV慢性感染者与正常人相比，JAK-STAT信号通路的正向调节因子STAT1和P48的表达和负向调节因子SOCS1和SOCS3的表达均显著增强(STAT1：CHC vs Donors 96.5 vs 69，P=0.039；P48：CHC vs Donors 121.5 vs 66.9，P=0.013；SOCS1：CHC vs Donors 5.96 vs 2.13，P=0.005；SOCS3：CHC vs Donors，78.3 vs 18.9，P＜0.001)；干扰素治疗前的SOCS1和SOCS5表达水平与HCV的复制水平呈负相关(R：-0.427和-0.385；P：0.013和0.027)。鉴于SOCS1和SOCS5主要表达于Th1细胞中，以上结果提示HCV复制可能会抑制Th1免疫的形成。进一步分析JAK-STAT信号通路正负调节因子的基因表达水平与干扰素疗效的相关性，发现治疗前负调因子SOCS3的高表达与干扰素的疗效不佳有关(NR vs SR，203.0 vs 64.1，P=0.007)，推测SOCS3的高表达削弱了细胞对外源性干扰素的反应能力。鉴于SOCS3主要在Th2细胞中表达，非持续应答病人治疗前的PBMCs中SOCS3的高水平表达也提示在Th2类免疫状态下机体对干扰素反应低下。利用流式细胞仪分析应答病人在干扰素治疗前后PBMCs中γ-干扰素和IL-4分泌细胞的水平，结果与SOCS的水平相一致：在干扰素治疗过程中，随着血清中HCV RNA的下降，SOCS1和SOCS5表达上调而SOCS3表达下调，Th1和Th2细胞百分含量也分别呈现增加和减少(Th1：治疗前vs治疗3个月，4.28+2.4％vs 7.63±3.5％P＜0.001；Th2：治疗前vs治疗3个月，0.66±0.52％vs 0.37±0.53％，P=0.013)，提示其可用于干扰素免疫调节效果的监测。进一步在体外肝细胞系中确认HCV蛋白表达和基因复制对肝细胞内SOCS1和SOCS3表达的影响，发现HCV编码的Core蛋白可诱导Huh7细胞表达SOCS1和SOCS3，尤其以SOCS1表达最为显著，其它HCV的非结构蛋白则无此诱导能力。
据于外源性干扰素治疗丙型肝炎的效果主要取决于能否诱导足够的抗病毒效应分子的产生，本研究进一步检测了抗病毒效应分子MxA，PKR和2’-5’OAS在干扰素治疗前及治疗过程中的表达，并分析其基因表达与疗效的关系。结果发现，病人PBMCs中MxA，PKR和2’5’OAS的表达在干扰素治疗前与对照相比已有明显增强(MxA：CHC vs Donors 141.4 vs 5.5，P＜0.001；PKR：CHC vs Donors 75.7 vs 12.3，P＜0.001；2’-5’OAS：CHC vs Donors 72.6 vs 4.18，P＜0.001)，干扰素治疗过程中其表达水平进一步升高，但其治疗前的基础表达水平和治疗中的诱导表达水平均与干扰素疗效无关，提示MxA，PKR和2’-5’OAS的基因表达不能用于疗效的预测。进一步用基因芯片(U-133A，Affymetrix公司)的方法检测了干扰素持续应答者和非持续应答者的PBMCs在体外用干扰素诱导后的全基因表达，发现不同疗效组的基因表达大部分相似，仅有16个基因的诱导表达倍数在不同疗效组中有显著性差异，包括已知的干扰素刺激基因(STAT1，GBP1，IFIT1，GIP3和IFIT2)，趋化因子基因(CXCL10和CCL8)，信号通路相关蛋白基因(STK3，RASGRP3，RIN2和LEPR)，代谢相关蛋白基因(APOBEC3A，MANEA和ZC3HAV1)和功能尚不明确的基因(REC8L1和C6orf62)。
综上所述，本研究的结果表明，HCV慢性感染对PBMCs中Ⅰ型干扰素及其
相关基因的表达产生影响。HCV慢性感染情况下，Ⅰ型干扰素及其合成关键因子的表达受到抑制，治疗前的表达水平与疗效无关；干扰素信号通路的正负调节因子的表达均被增强，并且负调因子SOCS3的表达水平与疗效相关；抗病毒效应分子MxA，PKR和2’-5’OAS的基础表达增强，基础表达和诱导表达水平均与疗效无关，同时基因组学研究发现PBMCs中16个基因在干扰素诱导下的表达变化与疗效相关。HCV慢性感染干预宿主细胞基因表达的有关机制有待进一步利用细胞系统进行深入研究；宿主基因表达与临床干扰素疗效的相关性研究及其在指导临床用药等方面的应用，需要在更大的病人样本中探索和实践。本课题的研究结果一方面能丰富人们对干扰素诱导抗病毒及HCV拮抗干扰素机制的认识，另一方面可指导临床预测疗效、筛选病人和制定治疗方案，最终实现个性化的干扰素治疗，达到提高丙型肝炎治愈率的目的。
Hepatitis C virus (HCV) is estimated to infect more than 170 million people worldwide. HCV infection usually causes chronic hepatitis in approximately 80% of infected individuals with 20% progressing to cirrhosis and 1-2% developing hepatocellular carcinoma. Currently, Interferon-α (IFN-α) is one of the most efficient drugs for the treatment of chronic hepatitis C and the standard treatment for HCV infection is IFN-α in combination with ribavirin. Unfortunately, more than 60% of patients with chronic HCV infection will experience either no control of viral replication by therapy (nonresponders) or a relapse when therapy is stopped (relapsers). The molecular mechanisms underlying failure of IFN-α treatment are not well understood. As our previous research found no correlation between viral genomic sequence and outcome of IFN-α treatment, host factors associated with IFN therapy, were explored in this study.
It has been proposed that HCV has the ability of interfering with type I interferon (IFN-α/β) production, blocking JAK-STAT signaling transduction, inhibiting expression or activity of ISGs (interferon -stimulated genes), which may support HCV replication and persistence as well as resistance to IFN-α therapy. Host gene expression is the outcome of host reaction to infection and viral interference with host, and may correlate with efficiency of IFN-α treatment. To identify signature or pattern of host gene expression which can distinguish different outcome of therapy, expression of type I interferon, interferon regulated factors (IRFs), positive and negative regulatory factors of IFN signaling, classical antiviral effectors and whole cellular genes were analyzed in peripheral blood mononuclear cells (PBMCs) from 34 chronic hepatitis C (CHC) patients before, during and after IFN/ribavirin treatment by
real-time quantitative RT-PCR and microarray. Among them, 6 of 34 were nonresponders (NR) and 28 of 34 were sustained responders (SR) according to serum HCV RNA test at the end of treatment and 6 months post-treatment. Gene expression was compared between these two groups.
To identify the correlation between gene expression of type I interferon, IRFs and outcome of IFN therapy, gene expression level of IFN-α/β, IRF-1, IRF-3, IRF-5 and IRF-7 were quantified in CHC patients. Result showed that, IFN-α and IFN-β expression level in the patients and donors were not significantly different (P: 0.927 and 0.937). However, IRF family members were differentially expressed in the two patient groups. IRF-3 expression was not influenced by chronic infection (P=0.980) and IRF-7 expression was not significantly different between patients and donors(P=0.071); IRF-1 expression levels were obviously enhanced in patients(P<0.001) and IRF-5 expression level was remarkably lower than that of the donors (P=0.002). The relevance analysis of gene expression showed that, while IRF-1 expression and the IFN-α/β expression did not have relevance, IRF-7 expression and IFN-α expression were correlated (R= 0.453; P= 0.007) and IRF-5 expression correlated with expression of both IFN-α and IFNβ (R=0.686, 0.621; P <0.001, 0.001). This result may reflect the mechanism of low expression of host type I interferon, which might be associated with low expression of key factors that regulated interferon's synthesis. However, further analysis did not reveal any correlation between gene expression of IFN-α/β, IRF-1, IRF-3, IRF-5 and IRF-7 before treatment and responses of host to interferon treatment. In order to confirm the influence of HCV persistent infection on IRF and type I interferon, HCV replicon cell (Huh7 cell line harboring subgenomic HCV replicons) was used. Result showed that, under NDV attack, IRF-7 promoter activity or the endogenous IRF-7 and IFN-α expression level of HCV replicon cells was lower than that of Huh7 cell (fold activity of IRF-7 promoter: 2.54 vs. 4.20, P<0.001; endogenous IRF-7mRNA expression level: 0.137 vs. 1.671, P<0.001; endogenous IFN-α mRNA expression level: 0.246 vs 2.207, P<0.001). On inhibition of HCV replication by ribavirin or IFN-α treatment, both IRF-7 promoter activity and expression of endogenous IRF-7and IFN-α were restored. This result may suggest a new mechanism of inhibition of type I interferon by HCV.
As the interferon-induced intracellular antiviral effect depends largely on efficiency of signal transduction (JAK-STAT pathway), the effect of HCV chronic
infection on expression of positive and negative regulatory genes of JAK-STAT pathway in patients' PBMCs was further analyzed. Results showed that, compared to healthy donors, the expressions of positive regulatory genes (STAT1 and P48) and negative regulatory genes (SOCS1 and SOCS3) were markedly enhanced in CHC patients (STAT1: CHC vs Donors 96.5 vs. 69, P=0.039; P48: CHC vs. Donors 121.5 vs. 66.9, P=0.013; SOCS1: CHC vs. Donors 5.96 vs. 2.13, P=0.005; SOCS3: CHC vs. Donors 78.3 vs. 18.9, P<0.001). Expression level of SOCS1 and SOCS5 before treatment was negatively correlated with serum HCV RNA level (R= -0.427 and -0.385; P= 0.013 and 0.027). As SOCS1 and the SOCS5 are mainly expressed in the Thl cell, the above result indicated that HCV replication might be able to suppress the Thl immunity. Further analysis of relevance between expression of these key factors of signaling pathway and outcome of IFN therapy showed high SOCS3 expression was predictive of no response to IFN therapy (NR vs. SR 203.0 vs. 64.1, P=0.007). It indicates that SOCS3 expression level may be used as marker for prediction of IFN-α therapy. Using flow cytometer to analyze the cytokine secreting cells, we found a switch-over of cytokine synthesis from Th2 to Th1 (IFN-γ for Th1, IL-4 for Th2) (Th1: 0 week vs. 12 weeks 4.28±2.4% vs. 7.63±3.5%, P<0.01; Th2: 0 week vs. 12 weeks 0.66±0.52% vs. 0.37±0.53%, P= 0.013) during IFN-α therapy, which was paralleled to up-regulation of SOCS1 and SOCS5 level and down-regulation of SOCS3 level. This result indicated that SOCS expression level in PBMC might be used to evaluate immune regulatory effect of IFN-α treatment. In vitro assay in cell line further showed that the expression of SOCS1 and SOCS3 in Huh7 cell line were induced by HCV Core protein, but not by other HCV non-structural proteins.
As antiviral effect of interferon mainly depends on induction of antiviral effectors molecule, gene expression of classical antiviral effectors MxA, PKR and 2 ' 5 ' OAS before and during treatment and its correlation with outcome of therapy were further analyzed. Results showed that, compared with healthy donors, MxA, PKR and 2 ' 5 ' OAS expression in patients were obviously enhanced (MxA: CHC vs. Donors 141.4 vs. 5.5, P<0.001; PKR: CHC vs. Donors 75.7 vs. 12.3, P<0.001; 2'5'OAS: CHC vs. Donors 72.6 vs. 4.18, P<0.001) and were further elevated by IFN-α treatment. However, there was no correlation of both basal expression levels and fold induction during treatment of the above genes with outcome of IFN treatment, which suggest that MxA, PKR and 2'5' OAS gene expression cannot be used as predictive marker for IFN therapy. DNA microarrays(Affymetrix Com, HG U-133A, containing 13000
cellular genes and EST) were further used to analyze the global gene expression of PBMCs in NR and SR groups. Comparing the fold of induction of genes from these two groups, we found that 16 genes were differentially expressed between NR and SR. These genes included ISGs (STAT1, GBP1, IFIT1, GIP3 and IFIT2), chemokines (CXCL10 and CCL8), genes related with signaling pathway (STK3, RASGRP3, RIN2 and LEPR), metabolism-correlated gene (APOBEC3A, MANEA and ZC3HAV1) and several genes with uncertain function (REC8L1 and C6orf62).
In summary, our research results indicated that HCV chronic infection has the influence on expression of type I interferon and its related genes in PBMCs. Under HCV chronic infection situation, while expression of type I interferon and its key regulatory factor was suppressed, the expression of the positive and negative regulatory factors of interferon signaling pathway were strengthened. Furthermore, while the expression of type I interferon, its key regulatory factor and MxA, PKR, 2'5'OAS before treatment have no relationship with outcome of IFN-α treatment, there was correlation of expression of negative factor SOCS3 with outcome of IFN-α treatment. In addition, 16 cellular genes in PBMCs were differentially expressed between NR and SR. Further study with HCV replicon cells is needed to understand the molecular mechanism underlying intervention of host cell gene expression by HCV chronic infection. The possible application of host gene expression pattern in guiding clinical interferon treatment such as predicting responses to IFN treatment need to be further investigated in a larger cohort of CHC patients. These findings, on the one hand, can enrich our knowledge about interaction between HCV and host defense system; on the other hand, may help to optimize clinical treatment and improve efficacy of IFN, and finally realize the individualized interferon treatment for HCV patients.
引文
1. WHO. Hepatitis C. Fact Sheet 164, 2000. http://www.who.int.
2. Di Bisceglie AM. Hepatitis C and hepatocellular carcinoma. HEPATOLOGY 1997; 26: 34S-38S.
3. Di Bisceglie AM, Martin P, Kassianides C, Lisker-Melman M, Murray L, Waggoner J, Goodman Z, Banks SM, Hoofnagle JH. Recombinant interferon alfa therapy for chronic hepatitis C. A randomized, double-blind, placebo-controlled trial. N Engl J Med. 1989 Nov 30; 321(22): 1506-10.
4. McHutchison JG, Gordon SC, Schiff ER, Shiffman ML, Lee WM, Rustgi VK, Goodman ZD, Ling MH, Cort S, Albrecht JK. Interferon alfa-2b alone or in combination with ribavirin as initial treatment for chronic hepatitis C. Hepatitis Interventional Therapy Group. N Engl J Med. 1998 Nov 19; 339(21): 1485-92.
5. Manns MP, McHutchison JG, Gordon SC, Rustgi VK, Shiffman M, Reindollar R, Goodman ZD, Koury K, Ling M, Albrecht JK. Peginterferon alfa-2b plus ribavirin compared with interferon alfa-2b plus ribavirin for initial treatment of chronic hepatitis C: a randomised trial. Lancet. 2001 Sep 22; 358(9286): 958-65.
6. Liang T J, Rehermann B, Seeff LB, Hoofnagle JH Pathogenesis, natural history, treatment, and prevention of hepatitis C. Ann Intern Med 2000; 132: 296-305.
7. Trepo C. Genotype and viral load as prognostic indicators in the treatment of hepatitis C. J Viral Hepat 2000; 7: 250-257
8. Enomoto N, Sakuma I, Asahina Yet al. Mutations in the nonstractural protein 5A gene and response to interferon in patients with chronic hepatitis C virus 1b infection. N Engl J Med 1996, 334(2): 77-81
9. Gale MJ Jr, Korth MJ, Tang NM, Tan SL, Hopkins DA, Dever TE, Polyak SJ, Gretch DR, Katze MG. Evidence that hepatitis C virus resistance to interferon is mediated through repression of the PKR protein kinase by the nonstructural 5A protein. Virology. 1997 Apr 14; 230(2): 217-27
10. Taylor DR, Shi ST, Romano PR et al. Inhibition of the interferon-inducible protein kinase PKR by HCV E2 protein. Science 1999, 285: 107-110
11．胡芸文，唐美芳，蒋伟伦和袁正宏慢性丙型肝炎病毒NS5A区序列与干扰素 疗效的关系 中华实验与临床病毒学杂志2002 16(2)：114-118
12．胡芸文，王健，张继明，谢怡和袁正宏慢性丙型肝炎患者外周血单个核细胞中趋化因子水平与干扰素治疗的关系中华微生物学和免疫学杂志200525 (6)：457-462
13. Kuzushita N, Hayashi N, Katayama K, Kanto T, Oshita M, Hagiwara H, Kasahara A, et al. High levels of serum interleukin-10 are associated with a poor response to interferon treatment in patients with chronic hepatitis C. Scand J Oastroenterol 1997; 32: 169-174
14. Larrea E, Garcia N, Qian C, Civeira MP, Prieto J. Tumor necrosis factor alpha gene expression and the response to interferon in chronic hepatitisC. HEPATOLOGY 1996; 23: 210-217
15. Kishihara Y, Hayashi J, Yoshimura E, Yamaji K, Nakashima K, Kashiwagi S. IL-1 beta and TNF-alpha produced by peripheral blood mononuclear cells before and during interferon therapy in patients with chronic hepatitis C. Dig Dis Sci 1996; 41: 315-321.
16. Polyak SJ, Khabar KS, Rezeiq M, Gretch DR. Elevated levels of interleukin-8 in serum are associated with hepatitis C virus infection and resistance to interferon therapy. J Virol 2001; 75: 6209-6211
17. Soardo G, Pirisi M, Fonda M, Fabris C, Falleti E, Toniutto P, Vitulli D, et al. Changes in blood lipid composition and response to interferon treatment in chronic hepatitis C. J Interferon Cytoldne Res 1995; 15: 705-712
18. Barrett S, Collins M, Kermy C, Ryan E, Keane CO, Crowe J Polymorphisms in tumour necrosis factor-alpha, transforming growth factor-beta, interleukin-10, interleukin-6, interferon-gamma, and outcome of hepatitis C vires infection. J Med Virol. 2003; 71(2): 212-8.
19. Sim H, Wojcik J, Margulies M, Wade JA, Heathcote J. Response to interferon therapy: influence of human leucocyte antigen alleles in patients with chronic hepatitis C. J Viral Hepat 1998; 5: 249-253.
20. Wawrzynowicz-Syczewska M, Underhill JA, Clare MA, Boron-Kaczmarska A, McFarlane IG, Donaldson PT. HLA class Ⅱ genotypes associated with chronic hepatitis C virus infection and response to alpha-interferon treatment in Poland. Liver 2000; 20:234-239
21. Romero-Gomez M, Gonzalez-Escribano MF, Torres B, Barroso N, Montes-Cano MA, Sanchez-Munoz D, Nunez-Roldan A, et al. HLA class I B44 is associated with sustained response to interferona ribavirin therapy in patients with chronic hepatitis C. Am J Gastroenterol 2003; 98:1621-1626.
22. Ishii K, Takamura N, Shinohara E, Shin HY, Ikehara T, Hata S, Kawafune T,Sumino Y, Ohmoto Y. Intracellular cytokine analysis of CD4-positive T cells predictive of sustained response to interferon therapy for patients with chronic hepatitis C. Dig Dis Sci. 2002; 47(4):778-83.
23. Barnes BJ, Field AE, Pitha-Rowe PM. Virus-induced heterodimer formation between IRF-5 and IRF-7 modulates assembly of the IFNA enhanceosome in vivo and transcriptional activity of IFNA genes. J Biol Chem. 2003; 278(19):16630-41.
24. Keskinen P, Nyqvist M, Sareneva T, Pirhonen J, Melen K, Julkunen I. Impaired antiviral response in human hepatoma cells. Virology. 1999; 263(2):364-75.
25. Colonna M, Krug A, Cella M. Interferon-producing cells: on the front line in immune responses against pathogens.Curr Opin Immunol. 2002; 14(3):373-9.
26. Izaguirre A, Barnes BJ, Amrute S, Yeow WS, Megjugorac N, Dai J, Feng.D, Chung E, Pitha PM, Fitzgerald-Bocarsly P. Comparative analysis of IRF and IFN-alpha expression in human plasmacytoid and monocyte-derived dendritic cells. J Leukoc Biol. 2003;74(6):1 125-38
27. Zhou S, Liu R, Baroudy BM, Malcolm BA, Reyes GR. The effect of ribavirin and IMPDH inhibitors on hepatitis C virus subgenomic replicon RNA. Virology. 2003;310(2):333-42.
28. Blight KJ, McKeating JA, Rice CM. Highly permissive cell lines for subgenomic and genomic hepatitis C virus RNA replication. Virol. 2002 ;76(24):13001-14
29. Foy E, Li K, Sumpter R Jr, Loo YM, Johnson CL, Wang C, Fish PM, Yoneyama M,Fujita T, Lemon SM, Gale M Jr. Control of antiviral defenses through hepatitis C virus disruption of retinoicacid-inducible gene-I signaling. Proc Natl Acad Sci U S A. 2005;102(8):2986-91
30. Li K, Foy E, Ferreon JC, Nakamura M, Ferreon AC, Ikeda M, Ray SC, Gale M Jr, Lemon SM. Immune evasion by hepatitis C virus NS3/4A protease-mediated cleavage of the Toll-like receptor 3 adaptor protein TRIF. Proc Natl Acad Sci U S A. 2005;102(8):2992-7
31.Abbate I, Romano M, Longo R, Cappiello G, Lo Iacono O, Di Marco V, PaparellaC, Spano A, Capobianchi MR. Endogenous levels of mRNA for IFNs and IFN-related genes in hepatic biopsies of chronic HCV-infected and non-alcoholic steatohepatitis patients. J Med Virol. 2003; 70(4): 581-7
32. Castelruiz Y, Larrea E, Boya P, Civeira MP, Prieto J. Interferon alfa subtypes and levels of type I interferons in the liver andperipheral mononuclear cells in patients with chronic hepatitis C and controls. Hepatology. 1999; 29(6):1900-4.
33. Mihm S, Frese M, Meier V, Wietzke-Braun P, Scharf JG, Bartenschlager R, Ramadori G. Interferon type I gene expression in chronic hepatitis C. Lab Invest. 2004; 84(9): 1148-59.
34. Bacon BR, McHutchison JG.Treatment issues with chronic hepatitis C: special populations and pharmacy strategies. Am J Manag Care. 2005 ; 11(10 Suppl):S296-306.
35. Barnes B, Lubyova B, Pitha PM. On the role of IRF in host defense.J Interferon Cytokine Res. 2002; 22(1):59-71.
36. Taniguchi T, Ogasawara K, Takaoka A, Tanaka N. IRF family of transcription factors as regulators of host defense. Annu Rev Immunol. 2001;19:623-55.
37. Fujita, T., Reis, L. F., Watanabe, N., Kimura, Y. & Taniguchi, T. Induction of the transcription factor IRF-1 and interferon- mRNAs by cytokines and activators of second-messenger pathways. Proc. Natl Acad. Sci. USA 1986;86:9936-9940 .
38. Barnes BJ, Richards J, Mancl M, Hanash S, Beretta L, Pitha PM. Global and distinct targets of IRF-5 and IRF-7 during innate response to viral infection. J Biol Chem. 2004; 279(43):45194-207.
39. Castillo I, Rodriguez-Inigo E, Bartolome J, de Lucas S, Ortiz-Movilla N,Lopez-Alcorocho JM, Pardo M, Carreno V. Hepatitis C virus replicates in peripheral blood mononuclear cells of patients with occult hepatitis C virus infection. Gut. 2005;54(5):682-5
40. Goutagny N, Fatmi A, De Ledinghen V, Penin F, Couzigou P, Inchauspe G, Bain C. Evidence of viral replication in circulating dendritic cells during hepatitis C virus infection. J Infect Dis. 2003; 187(12):1951-8.
41. Zhang T, Lin RT, Li Y, Douglas SD, Maxcey C, Ho C, Lai JP, Wang YJ, Wan Q, Ho WZ. Hepatitis C virus inhibits intracellular interferon alpha expression in human hepatic cell lines. Hepatology. 2005; 42(4):819-27.
42. Alexander WS, Hilton DJ. The role of suppressors of cytokine signaling (SOCS) proteins in regulation of the immune response. Annu Rev Immunol. 2004; 22: 503-29.
43. Wormald S, Hilton DJ. Inhibitors of cytokine signal transduction. J Bio1 Chem. 2004; 279(2):821-4.
44. Lin W, Choe WH, Hiasa Y, Kamegaya Y, Blackard JT, Schmidt EV, Chung RT. Hepatitis C virus expression suppresses interferon signaling by degrading STATl.Gastroenterology. 2005; 128(4):1034-41.
45. Melen K, Fagerlund R, Nyqvist M, Keskinen P, Julkunen I Expression of hepatitis C virus core protein inhibits interferon-induced nuclear import of STATs. J Med Virol. 2004; 73(4):536-47.
46. Geiss, G. K. et al. Gene expression profiling of the cellular transcriptional network regulated by α/β interferon and its partial attenuation by the hepatitis C virus nonstructural 5A protein. J Virol 2003;77: 6367-6375.
47. Bode JG, Ludwig S, Ehrhardt C, Albrecht U, Erhardt A, Schaper F, Heinrich PC, Haussinger D. IFN-alpha antagonistic activity of HCV core protein involves induction of suppressor of cytokine signaling-3. FASEB J. 2003; 17(3):488-90.
48. Imanaka K, Tamura S, Fukui K, Ito N, Kiso S, Imai Y, Naka T, Kishimoto T, Kawata S, Shinomura Y; The Kansai Viral Hepatitis Research Group. Enhanced expression of suppressor of cytokine signalling-1 in the liver of chronic hepatitis C: possible involvement in resistance to interferon therapy. J Viral Hepat. 2005; 12(2):130-8.
49. Egwuagu CE, Yu CR, Zhang M, Mahdi RM, Kim SJ, Gery I. Suppressors of cytokine signaling proteins are differentially expressed in Th1 and Th2 cells: implications for Th cell lineage commitment and maintenance. J Immunol 2002; 168:3181-7.
50. Seki Y, Hayashi K, Matsumoto A, Seki N, Tsukada J, Ransom J,et al. Expression of the suppressor of cytokine signaling-5 (SOCS5) negatively regulates IL-4-dependent STAT6 activation and Th2 differentiation. Proc Natl Acad Sci U S A 2002; 99:13003-8.
51. Egwuagu CE, Yu CR, Li Z, Nussenblatt RB. SOCS5 mRNA levels in peripheral blood mononuclear cells (PBMC): a potential bio-marker for monitoring response of uveitis patients to Daclizumab therapy.J Autoimmun. 2005;24(l):39-46
52. Blumenstein M, Bowen-Shauver JM, Keelan JA, Mitchell MD. Identification of suppressors of cytokine signaling (SOCS) proteins in human gestational tissues: differential regulation is associated with the onset of labor. J Clin Endocrinol Metab. 2002;87(3):1094-7
53. Sun WH, Pabon C, Alsayed Y, Huang PP, Jandeska S, Uddin S, Platanias LC, Rosen ST. Interferon-alpha resistance in a cutaneous T-cell lymphoma cell line is associated with lack of STAT1 expression. Blood. 1998;91(2):570-6
54. Wong LH, Sim H, Chatterjee-Kishore M, Hatzinisiriou I, Devenish RJ, Stark G,Ralph SJ. Isolation and characterization of a human STAT1 gene regulatory element. Inducibility by interferon (IFN) types I and II and role of IFN regulatory factor- 1.J Biol Chem. 2002;277(22): 19408-17
55. Xiao W, Wang L, Yang X, Chen T, Hodge D, Johnson PF, Farrar W. CCAAT/enhancer-binding protein beta mediates interferon-gamma - induced p48(ISGF3-gamma ) gene transcription in human monocytic cells. J Biol Chem. 2001;276(26):23275-81.
56. Minoru Fujimoto and Tetsuji Naka Regulation of cytokine signaling by SOCS family molecules. TRENDS in Immunology 2003;.24(12): 659-666
57. Barth, H. et al. Uptake and presentation of hepatitis C virus-like particles by human dendritic cells. Blood 2005(105):3605-3614.
58. Meier V, Mihm S, Ramadori G. MxA gene expression in peripheral blood mononuclear cells from patients infected chronically with hepatitis C virus treated with interferon-alpha. J Med Virol. 2000; 62(3):318-26.
59. MacQuillan GC, Mamotte C, Reed WD, Jeffrey GP, Allan JE. Upregulation of endogenous intrahepatic interferon stimulated genes during chronic hepatitis C virus infection. J Med Virol. 2003; 70(2):219-27.
60. Antonelli G, Simeoni E, Turriziani O, Tesoro R, Redaelli A, Roffi L, Antonelli L, Pistello M, Dianzani F. Correlation of interferon-induced expression of MxA mRNA in peripheral blood mononuclear cells with the response of patients with chronic active hepatitis C to IFN-alpha therapy. J Interferon Cytokine Res. 1999;19(3):243-51
61. Gerotto M, Dal Pero F, Bortoletto G, Realdon S, Ferrari A, Boccato S,Alberti A. PKR gene expression and response to pegylated interferon plus ribavirin therapy in chronic hepatitis C. Antivir Ther. 2004; 9(5):763-70.
62. Pawlotsky JM, Roudot-Thoraval F, Bastie A, Darthuy F, Remire J, Metreau JM, Zafrani ES, Duval J, Dhumeaux D. Factors affecting treatment responses to interferon-alpha in chronic hepatitis C.J Infect Dis. 1996; 174(1): 1-7
63. Ji X, Cheung R, Cooper S, Li Q, Greenberg HB, He XS. Interferon alfa regulated gene expression in patients initiating interferon treatment for chronic hepatitis C. Hepatology. 2003;37(3):610-21
64. Chen L, Borozan I, Feld J, Sun J, Tannis LL, Coltescu C, Heathcote J,Edwards AM, McGilvray ID. Hepatic gene expression discriminates responders and nonresponders in treatment of chronic hepatitis C viral infection. Gastroenterology. 2005; 128(5): 1437-44.
65. Farrugia J, Cann AJ. Internally controlled quantitative assays for PKR mRNA and protein from liver biopsies and other finite clinical samples. J Immunol Methods. 1999 Aug 31;228(1-2):59-68
66. Fernandez M, Quiroga JA, Martin J, Herrero M, Pardo M, Horis berger MA, Carreno V. 1999. In vivo and in vitro induction of MxA protein in peripheral blood mononuclear cells from patients chronically infected with hepatitis C virus. J Infect Dis 180:262-267.
67. Goetschy JF, Zeller H, Content J, et al. Regulation of the interferon-inducible IFI-78K gene, the human equivalent of the murine Mx gene, by interferons, doublestranded RNA, certain cytokines, and viruses. J Virol 1989; 63: 2616-2622.
68. Memet S, Besancon F, Bourgeade MF, et al. Direct induction of interferon-gamma- and interferon-alpha/ beta-inducible genes by double-stranded RNA. J Interferon Res. 1991; 11:131-141.
69. Kuhen KL, Samuel CE. Isolation of the interferon-inducible RNA-dependent protein kinase Pkr promoter and identification of a novel DNA element within the 5'-flanking region of human and mouse Pkr genes. Virology. 1997; 227(l):119-30.
70. MacQuillan GC, de Boer WB, Platten MA, McCaul KA, Reed WD, Jeffrey GP, Allan JE. Intrahepatic MxA and PKR protein expression in chronic hepatitis C virus infection. J Med Virol. 2002; 68(2):197-205.
71. Frese M, Pietschmann T, Moradpour D, Haller O, Bartenschlager R. Interferon-alpha inhibits hepatitis C virus subgenomic RNA replication by an MxA-independent pathway. J Gen Virol. 2001; 82(4):723-33.
72. Der SD, Zhou A, Williams BR, Silverman RH. Identification of genes differentially regulated by interferon alpha, beta, or gamma using oligonucleotide arrays. Proc Natl Acad Sci U S A. 1998; 95(26): 15623-8.
73. de Veer MJ, Holko M, Frevel M, Walker E, Der S, Paranjape JM, Silverman RH, Williams BR. Functional classification of interferon-stimulated genes identified us ingmicroarrays. J Leukoc Biol 2001; 69:912-920.
74. Butera D, Marukian S, Iwamaye AE, Hembrador E, Chambers TJ, Di Bisceglie AM,Charles ED, Talal AH, Jacobson IM, Rice CM, Dustin LB. Plasma chemokine levels correlate with the outcome of antiviral therapy in patients with hepatitis C. Blood. 2005; 106(4): 1175-82.
75. Apolinario A, Majano PL, Lorente R, Nunez O, Clemente G, Garcia-Monzon C. Gene expression profile of T-cell-specific chemokines in human hepatocyte-derived cells: evidence for a synergistic inducer effect of cytokines and hepatitis C virus proteins. J Viral Hepat. 2005;12(1):27-37.
1. Liang TJ, Rehermann B, Seeff LB, Hoofnagle JH. Pathogenesis, natural history, treatment, and prevention of hepatitis C. Ann Intern Med 2000; 132: 296-305.
2. Di Bisceglie AM. Hepatitis C and hepatocellular carcinoma. HEPATOLOGY 1997; 26: 34S-38S.
3. Davis GL. Current therapy for chronic hepatitis C. Gastroenterology 2000; 118: S104-114.
4. Trepo C. Genotype and viral load as prognostic indicators in the treatment of hepatitis C. J Viral Hepat 2000; 7: 250-257.
5. Mabee CL, Crippin JS, Lee WM. Review article: interferon and hepatitis C—factors predicting therapeutic outcome. Aliment Pharmacol Ther 1998; 12: 509-518.
6. Pestka S. The human interferon alpha species and receptors. Biopolymers 2000; 55: 254-287.
7. Pfeffer LM, Basu L, Pfeffer SR, Yang CH, Murti A, Russell-Harde D, Croze E. The short form of the interferon alpha/beta receptor chain 2 acts as a dominant negative for type Ⅰ interferon action. J Biol Chem 1997; 272: 11002-11005.
8. Radaeva S, Jaruga B, Hong F, KimWH, Fan S, Cai H, Strom S, et al. Interferon-alpha activates multiple STAT signals and down-regulates c-Met in primary human hepatocytes. Gastroenterology 2002; 122: 1020-1034.
9. Ji X, Cheung R, Cooper S, Li Q, GreenbergHB, He XS. Interferon alfa regulated gene expression in patients initiating interferon treatment for chronic hepatitis C. HEPATOLOGY 2003; 37: 610-621.
10. Brassard DL, Grace MJ, Bordens RW. Interferon-alpha as an immunotherapeutic protein. J Leukoc Biol 2002; 71: 565-581.
11. Krebs DL, Hilton DJ. SOCS: physiological suppressors of cytokine signaling. J Cell Sci 2000; 113:2813-2819.
12. Hong F, Jaruga B, Kim WH, Radaeva S, El-Assal ON, Tian Z, Nguyen VA, et al. Opposing roles of STAT1 and STAT3 in T cell-mediated hepatitis: regulation by SOCS. J Clin Invest 2002; 110:1503-1513.
13. Campbell JS, Prichard L, Schaper F, Schmitz J, Stephenson-Famy A, Rosenfeld ME, Argast GM, et al. Expression of suppressors of cytokine signaling during liver regeneration. J Clin Invest 2001:107:1285-1292.
14. Song MM, Shuai K. The suppressor of cytokine signaling (SOCS) 1 and S0CS3 but not S0CS2 proteins inhibit interferon-mediated antiviral and antiproliferative activities. J Biol Chem 1998; 273:35056 - 35062.
15. David M, Chen HE, Goelz S, Larner AC, Neel BG. Differential regulation of the alpha/beta interferon-stimulated Jak/Stat pathway by the SH2 domain-containing tyrosine phosphatase SHPTPl. Mol Cell Biol 1995; 15:7050 -7058.
16. Morita K, Tanaka K, Saito S, Kitamura T, Kiba T, Fujii T, Numata K, et al. Expression of interferon receptor genes in the liver as a predictor of interferon response in patients with chronic hepatitis C. J Med Virol 1999; 58:359 -365.
17. Mizukoshi E, Kaneko S, Kaji K, Terasaki S, Matsushita E, Muraguchi M, Ohmoto Y, et al. Serum levels of soluble interferon Alfa/Beta receptor as an inhibitory factor of interferon in the patients with chronic hepatitis C. HEPATOLOGY 1999:30:1325-1331.
18. Larrea E, Garcia N, QianC, CiveiraMP, Prieto J. Tumor necrosis factor alpha gene expression and the response to interferon in chronic hepatitis C. HEPATOLOGY 1996:23:210 -217.
19. Neuman MG, Benhamou JP, Malkiewicz IM, Akremi R, Shear NH, Asselah T, Ibrahim A, et al. Cytokines as predictors for sustained response and as markers for immunomodulation in patients with chronic hepatitis C. Clin Biochem 2001:34:173-182.
20. Nelson DR, Tu Z, Soldevila-Pico C, Abdelmalek M, Zhu H, Xu YL, Cabrera R, et al. Long-term interleukin 10 therapy in chronic hepatitis C patients has a proviral and anti-inflammatory effect. HEPATOLOGY 2003; 38:859-868.
21. Edwards-Smith CJ, Jonsson JR, Purdie DM, Bansal A, Shorthouse C, Powell EE. Interleukin-10 promoter polymorphism predicts initial response of chronic hepatitis C to interferon alfa. HEPATOLOGY 1999;30: 526 - 530.
22. Yee LJ, Tang J, Gibson AW, Kimberly R, Van Leeuwen DJ, Kaslow RA. Interleukin 10 polymorphisms as predictors of sustained response in antiviral therapy for chronic hepatitis C infection. HEPATOLOGY 2001:33:708-712
23. Kishihara Y, Hayashi J, Yoshimura E, Yamaji K, Nakashima K, Kashiwagi S. IL—1 beta and TNF-alpha produced by peripheral blood mononuclear cells before and during interferon therapy in patients with chronic hepatitis C. Dig Dis Sci 1996:41:315-321.
24. Tian Z, Shen X, Feng H, Gao B. IL-1 beta attenuates IFN-alpha beta induced antiviral activity and STAT1 activation in the liver: involvement of proteasome-dependent pathway. J Immunol 2000; 165:3959 - 3965.
25. Thimme R, Bukh J, Spangenberg HC, Wieland S, Pemberton J, Steiger C, Govindarajan S, et al. Viral and immunological determinants of hepatitis C virus clearance, persistence, and disease. Proc Natl Acad Sci U S A 2002; 99:15661 -15668.
26. Saez-Royuela F, Porres JC, Moreno A, Castillo I, Martinez G, Galiana F, Carreno V. High doses of recombinant alpha-interferon or gamma-interferon for chronic hepatitis C: a randomized, controlled trial. HEPATOLOGY 1991; 13:327-331.
27. Katayama K, Kasahara A, Sasaki Y, Kashiwagi T, Naito M, Masuzawa M, Katoh M, et al. Immunological response to interferon-gamma priming prior to interferon- alpha treatment in refractory chronic hepatitis C in relation to viral clearance. J Viral Hepat 2001; 8:180 -185.
28. Dumoulin FL, Wennrich U, Nischalke HD, Leifeld L, Fischer HP, Sauerbruch T, Spengler U. Intrahepatic mRNA levels of interferon gamma and tumor necrosis factor alpha and response to antiviral treatment of chronic hepatitis C. J Hum Virol 2001;4:195 - 199.
29. Polyak SJ, Khabar KS, Rezeiq M, Gretch DR. Elevated levels of interleukin- 8 in serum are associated with hepatitis C virus infection and resistance to interferon therapy. J Virol 2001; 75:6209-6211.
30. 69. Khabar KS, Al-Zoghaibi F, Al-Ahdal MN, Murayama T, Dhalla M, Mukaida N, Taha M, et al. The alpha chemokine, interleukin 8, inhibits the antiviral action of interferon alpha. J Exp Med 1997:186:1077-1085.
31. Promrat K, McDermott DH, Gonzalez CM, Kleiner DE, Koziol DE, Lessie M, Merrell M, et al. Associations of chemokine system polymorphisms with clinical outcomes and treatment responses of chronic hepatitis C. Gastroenterology 2003:124:352-360.
32. Hayashi J, Kishihara Y, Ueno K, Yamaji K, Kawakami Y, Furusyo N, Sawayama Y, et al. Age-related response to interferon alfa treatment in women vs men with chronic hepatitis C virus infection. Arch Intern Med 1998:158:177-181.
33. Layden-Almer JE, Ribeiro RM, Wiley T, Perelson AS, Layden TJ. Viral dynamics and response differences in HCV-infected African American and white patients treated with IFN and ribavirin. HEPATOLOGY 2003; 37:1343-1350.
34. Sim H, Wojcik J, Margulies M, Wade JA, Heathcote J. Response to interferon therapy: influence of human leucocyte antigen alleles in patients with chronic hepatitis C. J Viral Hepat 1998; 5:249 -253.
35. Wawrzynowicz-Syczewska M, Underhill JA, Clare MA, Boron-Kaczmarska A, McFarlane IG, Donaldson PT. HLA class II genotypes associated with chronic hepatitis C virus infection and response to alpha-interferon treatment in Poland. Liver 2000; 20:234 -239.
36. Romero-Gomez M, Gonzalez-Escribano MF, Torres B, Barroso N, Montes- Cano MA, Sanchez-Munoz D, Nunez-Roldan A, et al. HLA class I B44 is associated with sustained response to interferon_ribavirin therapy in patients with chronic hepatitis C.Am J Gastroenterol 2003:98:1621-1626.
37. McCullough AJ. Obesity and its nurturing effect on hepatitis C. HEPATOLOGY 2003; 38:557-559.
38. Bressler BL, Guindi M, Tomlinson G, Heathcote J. High body mass index is an independent risk factor for nonresponse to antiviral treatment in chronic hepatitis C. HEPATOLOGY 2003:38:639-644.
39. Banner BF, Barton AL, Cable EE, Smith L, Bonkovsky HL. A detailed analysis of the Knodell score and other histologic parameters as predictors of response to interferon therapy in chronic hepatitis C. Mod Pathol 1995; 8:232-238.
40. Myers RP, Patel K, Pianko S, Poynard T, McHutchison JG. The rate of fibrosis progression is an independent predictor of the response to antiviral therapy in chronic hepatitis C. J Viral Hepat 2003; 10:16 -22.
41. Farci, P. Hepatitis C virus. The importance of viral heterogeneity. Clin Liver Dis 2001; 5, 895-916
42. Farci, P. et al. Early changes in hepatitis C viral quasispecies during interferon therapy predict the therapeutic outcome. Proc Natl Acad Sci USA 2002; 99, 3081 -3086
43. Farci, P. et al. The outcome of acute hepatitis C predicted by the evolution of the viral quasispecies. Science 2000-, 288, 339 - 344.
44. Enomoto, N. et al. Mutations in the nonstructural protein 5A gene and response to interferon in patients with chronic hepatitis C virus 1b infection. N Engl J Med 1996; 334, 77-81
45. Schinkel, J., Spoon, W. J. & Kroes, A. C. Meta-analysis of mutations in the NS5A gene and hepatitis C virus resistance to interferon therapy: uniting discordant conclusions. Antivir Ther 2004; 9, 275 - 286.
46. Geiss, G. K. et al. Gene expression profiling of the cellular transcriptional network regulated by α/β interferon and its partial attenuation by the hepatitis C virus nonstructural 5A protein. J Virol 2003; 77, 6367-6375.
47. Lin W, Choe WH, Hiasa Y, Kamegaya Y, Blackard JT, Schmidt EV, Chung RT. Hepatitis C virus expression suppresses interferon signaling by degrading STAT1.Gastroenterology. 2005; 128(4):1034-41.
48. Bode JG, Ludwig S, Ehrhardt C, Albrecht U, Erhardt A, Schaper F, Heinrich PC, Haussinger D. IFN-alpha antagonistic activity of HCV core protein involves induction of suppressor of cytokine signaling-3. FASEB J. 2003; 17(3):488-90.
49. Melen K, Fagerlund R, Nyqvist M, Keskinen P, Julkunen I Expression of hepatitis C virus core protein inhibits interferon-induced nuclear import of STATs. J Med Virol. 2004; 73(4):536-47.
50. Duong FH, Filipowicz M, Tripodi M, La Monica N, Heim MH. Hepatitis C virus inhibits interferon signaling through up-regulation of protein phosphatase 2A. Gastroenterology. 2004;126(1):263-77.
51. Gale, M. Jr et al. Control of PKR protein kinase by hepatitis C virus nonstructural 5A protein: molecular mechanisms of kinase regulation. Mol Cell Biol 1998; 18, 5208-5218
52. Taylor, D. R., Shi, S. T., Romano, P. R., Barber, G. N. & Lai, M. M. C. Inhibition of the interferon inducible protein kinase PKR by HCV E2 protein. Science 1999; 285, 107-110.
53. Au, W. C., Yeow, W. S. & Pitha, P. M. Analysis of functional domains of interferon regulatory factor 7 and its association with IRF-3. Virology 2001; 280, 273-282.
54. Breiman, A. et al. Inhibition of RIG-I-dependent signaling to the interferon pathway during hepatitis C virus expression and restoration of signaling by IKK_. J. Virol. 2005; 79, 3969-3978.
55. Han, J. Q., Wroblewski, G., Xu, Z., Silverman, R. H. & Barton, D. J. Sensitivity of hepatitis C virus RNA to the antiviral enzyme ribonuclease L is determined by a subset of efficient cleavage sites. J. Interferon Cytokine Res. 2004; 24, 664 - 676.
2. Di Bisceglie AM. Hepatitis C and hepatocellular carcinoma. HEPATOLOGY 1997; 26: 34S-38S.
3. Di Bisceglie AM, Martin P, Kassianides C, Lisker-Melman M, Murray L, Waggoner J, Goodman Z, Banks SM, Hoofnagle JH. Recombinant interferon alfa therapy for chronic hepatitis C. A randomized, double-blind, placebo-controlled trial. N Engl J Med. 1989 Nov 30; 321(22): 1506-10.
4. McHutchison JG, Gordon SC, Schiff ER, Shiffman ML, Lee WM, Rustgi VK, Goodman ZD, Ling MH, Cort S, Albrecht JK. Interferon alfa-2b alone or in combination with ribavirin as initial treatment for chronic hepatitis C. Hepatitis Interventional Therapy Group. N Engl J Med. 1998 Nov 19; 339(21): 1485-92.
5. Manns MP, McHutchison JG, Gordon SC, Rustgi VK, Shiffman M, Reindollar R, Goodman ZD, Koury K, Ling M, Albrecht JK. Peginterferon alfa-2b plus ribavirin compared with interferon alfa-2b plus ribavirin for initial treatment of chronic hepatitis C: a randomised trial. Lancet. 2001 Sep 22; 358(9286): 958-65.
6. Liang T J, Rehermann B, Seeff LB, Hoofnagle JH Pathogenesis, natural history, treatment, and prevention of hepatitis C. Ann Intern Med 2000; 132: 296-305.
7. Trepo C. Genotype and viral load as prognostic indicators in the treatment of hepatitis C. J Viral Hepat 2000; 7: 250-257
8. Enomoto N, Sakuma I, Asahina Yet al. Mutations in the nonstractural protein 5A gene and response to interferon in patients with chronic hepatitis C virus 1b infection. N Engl J Med 1996, 334(2): 77-81
9. Gale MJ Jr, Korth MJ, Tang NM, Tan SL, Hopkins DA, Dever TE, Polyak SJ, Gretch DR, Katze MG. Evidence that hepatitis C virus resistance to interferon is mediated through repression of the PKR protein kinase by the nonstructural 5A protein. Virology. 1997 Apr 14; 230(2): 217-27
10. Taylor DR, Shi ST, Romano PR et al. Inhibition of the interferon-inducible protein kinase PKR by HCV E2 protein. Science 1999, 285: 107-110
11．胡芸文，唐美芳，蒋伟伦和袁正宏慢性丙型肝炎病毒NS5A区序列与干扰素 疗效的关系 中华实验与临床病毒学杂志2002 16(2)：114-118
12．胡芸文，王健，张继明，谢怡和袁正宏慢性丙型肝炎患者外周血单个核细胞中趋化因子水平与干扰素治疗的关系中华微生物学和免疫学杂志200525 (6)：457-462
13. Kuzushita N, Hayashi N, Katayama K, Kanto T, Oshita M, Hagiwara H, Kasahara A, et al. High levels of serum interleukin-10 are associated with a poor response to interferon treatment in patients with chronic hepatitis C. Scand J Oastroenterol 1997; 32: 169-174
14. Larrea E, Garcia N, Qian C, Civeira MP, Prieto J. Tumor necrosis factor alpha gene expression and the response to interferon in chronic hepatitisC. HEPATOLOGY 1996; 23: 210-217
15. Kishihara Y, Hayashi J, Yoshimura E, Yamaji K, Nakashima K, Kashiwagi S. IL-1 beta and TNF-alpha produced by peripheral blood mononuclear cells before and during interferon therapy in patients with chronic hepatitis C. Dig Dis Sci 1996; 41: 315-321.
16. Polyak SJ, Khabar KS, Rezeiq M, Gretch DR. Elevated levels of interleukin-8 in serum are associated with hepatitis C virus infection and resistance to interferon therapy. J Virol 2001; 75: 6209-6211
17. Soardo G, Pirisi M, Fonda M, Fabris C, Falleti E, Toniutto P, Vitulli D, et al. Changes in blood lipid composition and response to interferon treatment in chronic hepatitis C. J Interferon Cytoldne Res 1995; 15: 705-712
18. Barrett S, Collins M, Kermy C, Ryan E, Keane CO, Crowe J Polymorphisms in tumour necrosis factor-alpha, transforming growth factor-beta, interleukin-10, interleukin-6, interferon-gamma, and outcome of hepatitis C vires infection. J Med Virol. 2003; 71(2): 212-8.
19. Sim H, Wojcik J, Margulies M, Wade JA, Heathcote J. Response to interferon therapy: influence of human leucocyte antigen alleles in patients with chronic hepatitis C. J Viral Hepat 1998; 5: 249-253.
20. Wawrzynowicz-Syczewska M, Underhill JA, Clare MA, Boron-Kaczmarska A, McFarlane IG, Donaldson PT. HLA class Ⅱ genotypes associated with chronic hepatitis C virus infection and response to alpha-interferon treatment in Poland. Liver 2000; 20:234-239
21. Romero-Gomez M, Gonzalez-Escribano MF, Torres B, Barroso N, Montes-Cano MA, Sanchez-Munoz D, Nunez-Roldan A, et al. HLA class I B44 is associated with sustained response to interferona ribavirin therapy in patients with chronic hepatitis C. Am J Gastroenterol 2003; 98:1621-1626.
22. Ishii K, Takamura N, Shinohara E, Shin HY, Ikehara T, Hata S, Kawafune T,Sumino Y, Ohmoto Y. Intracellular cytokine analysis of CD4-positive T cells predictive of sustained response to interferon therapy for patients with chronic hepatitis C. Dig Dis Sci. 2002; 47(4):778-83.
23. Barnes BJ, Field AE, Pitha-Rowe PM. Virus-induced heterodimer formation between IRF-5 and IRF-7 modulates assembly of the IFNA enhanceosome in vivo and transcriptional activity of IFNA genes. J Biol Chem. 2003; 278(19):16630-41.
24. Keskinen P, Nyqvist M, Sareneva T, Pirhonen J, Melen K, Julkunen I. Impaired antiviral response in human hepatoma cells. Virology. 1999; 263(2):364-75.
25. Colonna M, Krug A, Cella M. Interferon-producing cells: on the front line in immune responses against pathogens.Curr Opin Immunol. 2002; 14(3):373-9.
26. Izaguirre A, Barnes BJ, Amrute S, Yeow WS, Megjugorac N, Dai J, Feng.D, Chung E, Pitha PM, Fitzgerald-Bocarsly P. Comparative analysis of IRF and IFN-alpha expression in human plasmacytoid and monocyte-derived dendritic cells. J Leukoc Biol. 2003;74(6):1 125-38
27. Zhou S, Liu R, Baroudy BM, Malcolm BA, Reyes GR. The effect of ribavirin and IMPDH inhibitors on hepatitis C virus subgenomic replicon RNA. Virology. 2003;310(2):333-42.
28. Blight KJ, McKeating JA, Rice CM. Highly permissive cell lines for subgenomic and genomic hepatitis C virus RNA replication. Virol. 2002 ;76(24):13001-14
29. Foy E, Li K, Sumpter R Jr, Loo YM, Johnson CL, Wang C, Fish PM, Yoneyama M,Fujita T, Lemon SM, Gale M Jr. Control of antiviral defenses through hepatitis C virus disruption of retinoicacid-inducible gene-I signaling. Proc Natl Acad Sci U S A. 2005;102(8):2986-91
30. Li K, Foy E, Ferreon JC, Nakamura M, Ferreon AC, Ikeda M, Ray SC, Gale M Jr, Lemon SM. Immune evasion by hepatitis C virus NS3/4A protease-mediated cleavage of the Toll-like receptor 3 adaptor protein TRIF. Proc Natl Acad Sci U S A. 2005;102(8):2992-7
31.Abbate I, Romano M, Longo R, Cappiello G, Lo Iacono O, Di Marco V, PaparellaC, Spano A, Capobianchi MR. Endogenous levels of mRNA for IFNs and IFN-related genes in hepatic biopsies of chronic HCV-infected and non-alcoholic steatohepatitis patients. J Med Virol. 2003; 70(4): 581-7
32. Castelruiz Y, Larrea E, Boya P, Civeira MP, Prieto J. Interferon alfa subtypes and levels of type I interferons in the liver andperipheral mononuclear cells in patients with chronic hepatitis C and controls. Hepatology. 1999; 29(6):1900-4.
33. Mihm S, Frese M, Meier V, Wietzke-Braun P, Scharf JG, Bartenschlager R, Ramadori G. Interferon type I gene expression in chronic hepatitis C. Lab Invest. 2004; 84(9): 1148-59.
34. Bacon BR, McHutchison JG.Treatment issues with chronic hepatitis C: special populations and pharmacy strategies. Am J Manag Care. 2005 ; 11(10 Suppl):S296-306.
35. Barnes B, Lubyova B, Pitha PM. On the role of IRF in host defense.J Interferon Cytokine Res. 2002; 22(1):59-71.
36. Taniguchi T, Ogasawara K, Takaoka A, Tanaka N. IRF family of transcription factors as regulators of host defense. Annu Rev Immunol. 2001;19:623-55.
37. Fujita, T., Reis, L. F., Watanabe, N., Kimura, Y. & Taniguchi, T. Induction of the transcription factor IRF-1 and interferon- mRNAs by cytokines and activators of second-messenger pathways. Proc. Natl Acad. Sci. USA 1986;86:9936-9940 .
38. Barnes BJ, Richards J, Mancl M, Hanash S, Beretta L, Pitha PM. Global and distinct targets of IRF-5 and IRF-7 during innate response to viral infection. J Biol Chem. 2004; 279(43):45194-207.
39. Castillo I, Rodriguez-Inigo E, Bartolome J, de Lucas S, Ortiz-Movilla N,Lopez-Alcorocho JM, Pardo M, Carreno V. Hepatitis C virus replicates in peripheral blood mononuclear cells of patients with occult hepatitis C virus infection. Gut. 2005;54(5):682-5
40. Goutagny N, Fatmi A, De Ledinghen V, Penin F, Couzigou P, Inchauspe G, Bain C. Evidence of viral replication in circulating dendritic cells during hepatitis C virus infection. J Infect Dis. 2003; 187(12):1951-8.
41. Zhang T, Lin RT, Li Y, Douglas SD, Maxcey C, Ho C, Lai JP, Wang YJ, Wan Q, Ho WZ. Hepatitis C virus inhibits intracellular interferon alpha expression in human hepatic cell lines. Hepatology. 2005; 42(4):819-27.
42. Alexander WS, Hilton DJ. The role of suppressors of cytokine signaling (SOCS) proteins in regulation of the immune response. Annu Rev Immunol. 2004; 22: 503-29.
43. Wormald S, Hilton DJ. Inhibitors of cytokine signal transduction. J Bio1 Chem. 2004; 279(2):821-4.
44. Lin W, Choe WH, Hiasa Y, Kamegaya Y, Blackard JT, Schmidt EV, Chung RT. Hepatitis C virus expression suppresses interferon signaling by degrading STATl.Gastroenterology. 2005; 128(4):1034-41.
45. Melen K, Fagerlund R, Nyqvist M, Keskinen P, Julkunen I Expression of hepatitis C virus core protein inhibits interferon-induced nuclear import of STATs. J Med Virol. 2004; 73(4):536-47.
46. Geiss, G. K. et al. Gene expression profiling of the cellular transcriptional network regulated by α/β interferon and its partial attenuation by the hepatitis C virus nonstructural 5A protein. J Virol 2003;77: 6367-6375.
47. Bode JG, Ludwig S, Ehrhardt C, Albrecht U, Erhardt A, Schaper F, Heinrich PC, Haussinger D. IFN-alpha antagonistic activity of HCV core protein involves induction of suppressor of cytokine signaling-3. FASEB J. 2003; 17(3):488-90.
48. Imanaka K, Tamura S, Fukui K, Ito N, Kiso S, Imai Y, Naka T, Kishimoto T, Kawata S, Shinomura Y; The Kansai Viral Hepatitis Research Group. Enhanced expression of suppressor of cytokine signalling-1 in the liver of chronic hepatitis C: possible involvement in resistance to interferon therapy. J Viral Hepat. 2005; 12(2):130-8.
49. Egwuagu CE, Yu CR, Zhang M, Mahdi RM, Kim SJ, Gery I. Suppressors of cytokine signaling proteins are differentially expressed in Th1 and Th2 cells: implications for Th cell lineage commitment and maintenance. J Immunol 2002; 168:3181-7.
50. Seki Y, Hayashi K, Matsumoto A, Seki N, Tsukada J, Ransom J,et al. Expression of the suppressor of cytokine signaling-5 (SOCS5) negatively regulates IL-4-dependent STAT6 activation and Th2 differentiation. Proc Natl Acad Sci U S A 2002; 99:13003-8.
51. Egwuagu CE, Yu CR, Li Z, Nussenblatt RB. SOCS5 mRNA levels in peripheral blood mononuclear cells (PBMC): a potential bio-marker for monitoring response of uveitis patients to Daclizumab therapy.J Autoimmun. 2005;24(l):39-46
52. Blumenstein M, Bowen-Shauver JM, Keelan JA, Mitchell MD. Identification of suppressors of cytokine signaling (SOCS) proteins in human gestational tissues: differential regulation is associated with the onset of labor. J Clin Endocrinol Metab. 2002;87(3):1094-7
53. Sun WH, Pabon C, Alsayed Y, Huang PP, Jandeska S, Uddin S, Platanias LC, Rosen ST. Interferon-alpha resistance in a cutaneous T-cell lymphoma cell line is associated with lack of STAT1 expression. Blood. 1998;91(2):570-6
54. Wong LH, Sim H, Chatterjee-Kishore M, Hatzinisiriou I, Devenish RJ, Stark G,Ralph SJ. Isolation and characterization of a human STAT1 gene regulatory element. Inducibility by interferon (IFN) types I and II and role of IFN regulatory factor- 1.J Biol Chem. 2002;277(22): 19408-17
55. Xiao W, Wang L, Yang X, Chen T, Hodge D, Johnson PF, Farrar W. CCAAT/enhancer-binding protein beta mediates interferon-gamma - induced p48(ISGF3-gamma ) gene transcription in human monocytic cells. J Biol Chem. 2001;276(26):23275-81.
56. Minoru Fujimoto and Tetsuji Naka Regulation of cytokine signaling by SOCS family molecules. TRENDS in Immunology 2003;.24(12): 659-666
57. Barth, H. et al. Uptake and presentation of hepatitis C virus-like particles by human dendritic cells. Blood 2005(105):3605-3614.
58. Meier V, Mihm S, Ramadori G. MxA gene expression in peripheral blood mononuclear cells from patients infected chronically with hepatitis C virus treated with interferon-alpha. J Med Virol. 2000; 62(3):318-26.
59. MacQuillan GC, Mamotte C, Reed WD, Jeffrey GP, Allan JE. Upregulation of endogenous intrahepatic interferon stimulated genes during chronic hepatitis C virus infection. J Med Virol. 2003; 70(2):219-27.
60. Antonelli G, Simeoni E, Turriziani O, Tesoro R, Redaelli A, Roffi L, Antonelli L, Pistello M, Dianzani F. Correlation of interferon-induced expression of MxA mRNA in peripheral blood mononuclear cells with the response of patients with chronic active hepatitis C to IFN-alpha therapy. J Interferon Cytokine Res. 1999;19(3):243-51
61. Gerotto M, Dal Pero F, Bortoletto G, Realdon S, Ferrari A, Boccato S,Alberti A. PKR gene expression and response to pegylated interferon plus ribavirin therapy in chronic hepatitis C. Antivir Ther. 2004; 9(5):763-70.
62. Pawlotsky JM, Roudot-Thoraval F, Bastie A, Darthuy F, Remire J, Metreau JM, Zafrani ES, Duval J, Dhumeaux D. Factors affecting treatment responses to interferon-alpha in chronic hepatitis C.J Infect Dis. 1996; 174(1): 1-7
63. Ji X, Cheung R, Cooper S, Li Q, Greenberg HB, He XS. Interferon alfa regulated gene expression in patients initiating interferon treatment for chronic hepatitis C. Hepatology. 2003;37(3):610-21
64. Chen L, Borozan I, Feld J, Sun J, Tannis LL, Coltescu C, Heathcote J,Edwards AM, McGilvray ID. Hepatic gene expression discriminates responders and nonresponders in treatment of chronic hepatitis C viral infection. Gastroenterology. 2005; 128(5): 1437-44.
65. Farrugia J, Cann AJ. Internally controlled quantitative assays for PKR mRNA and protein from liver biopsies and other finite clinical samples. J Immunol Methods. 1999 Aug 31;228(1-2):59-68
66. Fernandez M, Quiroga JA, Martin J, Herrero M, Pardo M, Horis berger MA, Carreno V. 1999. In vivo and in vitro induction of MxA protein in peripheral blood mononuclear cells from patients chronically infected with hepatitis C virus. J Infect Dis 180:262-267.
67. Goetschy JF, Zeller H, Content J, et al. Regulation of the interferon-inducible IFI-78K gene, the human equivalent of the murine Mx gene, by interferons, doublestranded RNA, certain cytokines, and viruses. J Virol 1989; 63: 2616-2622.
68. Memet S, Besancon F, Bourgeade MF, et al. Direct induction of interferon-gamma- and interferon-alpha/ beta-inducible genes by double-stranded RNA. J Interferon Res. 1991; 11:131-141.
69. Kuhen KL, Samuel CE. Isolation of the interferon-inducible RNA-dependent protein kinase Pkr promoter and identification of a novel DNA element within the 5'-flanking region of human and mouse Pkr genes. Virology. 1997; 227(l):119-30.
70. MacQuillan GC, de Boer WB, Platten MA, McCaul KA, Reed WD, Jeffrey GP, Allan JE. Intrahepatic MxA and PKR protein expression in chronic hepatitis C virus infection. J Med Virol. 2002; 68(2):197-205.
71. Frese M, Pietschmann T, Moradpour D, Haller O, Bartenschlager R. Interferon-alpha inhibits hepatitis C virus subgenomic RNA replication by an MxA-independent pathway. J Gen Virol. 2001; 82(4):723-33.
72. Der SD, Zhou A, Williams BR, Silverman RH. Identification of genes differentially regulated by interferon alpha, beta, or gamma using oligonucleotide arrays. Proc Natl Acad Sci U S A. 1998; 95(26): 15623-8.
73. de Veer MJ, Holko M, Frevel M, Walker E, Der S, Paranjape JM, Silverman RH, Williams BR. Functional classification of interferon-stimulated genes identified us ingmicroarrays. J Leukoc Biol 2001; 69:912-920.
74. Butera D, Marukian S, Iwamaye AE, Hembrador E, Chambers TJ, Di Bisceglie AM,Charles ED, Talal AH, Jacobson IM, Rice CM, Dustin LB. Plasma chemokine levels correlate with the outcome of antiviral therapy in patients with hepatitis C. Blood. 2005; 106(4): 1175-82.
75. Apolinario A, Majano PL, Lorente R, Nunez O, Clemente G, Garcia-Monzon C. Gene expression profile of T-cell-specific chemokines in human hepatocyte-derived cells: evidence for a synergistic inducer effect of cytokines and hepatitis C virus proteins. J Viral Hepat. 2005;12(1):27-37.
1. Liang TJ, Rehermann B, Seeff LB, Hoofnagle JH. Pathogenesis, natural history, treatment, and prevention of hepatitis C. Ann Intern Med 2000; 132: 296-305.
2. Di Bisceglie AM. Hepatitis C and hepatocellular carcinoma. HEPATOLOGY 1997; 26: 34S-38S.
3. Davis GL. Current therapy for chronic hepatitis C. Gastroenterology 2000; 118: S104-114.
4. Trepo C. Genotype and viral load as prognostic indicators in the treatment of hepatitis C. J Viral Hepat 2000; 7: 250-257.
5. Mabee CL, Crippin JS, Lee WM. Review article: interferon and hepatitis C—factors predicting therapeutic outcome. Aliment Pharmacol Ther 1998; 12: 509-518.
6. Pestka S. The human interferon alpha species and receptors. Biopolymers 2000; 55: 254-287.
7. Pfeffer LM, Basu L, Pfeffer SR, Yang CH, Murti A, Russell-Harde D, Croze E. The short form of the interferon alpha/beta receptor chain 2 acts as a dominant negative for type Ⅰ interferon action. J Biol Chem 1997; 272: 11002-11005.
8. Radaeva S, Jaruga B, Hong F, KimWH, Fan S, Cai H, Strom S, et al. Interferon-alpha activates multiple STAT signals and down-regulates c-Met in primary human hepatocytes. Gastroenterology 2002; 122: 1020-1034.
9. Ji X, Cheung R, Cooper S, Li Q, GreenbergHB, He XS. Interferon alfa regulated gene expression in patients initiating interferon treatment for chronic hepatitis C. HEPATOLOGY 2003; 37: 610-621.
10. Brassard DL, Grace MJ, Bordens RW. Interferon-alpha as an immunotherapeutic protein. J Leukoc Biol 2002; 71: 565-581.
11. Krebs DL, Hilton DJ. SOCS: physiological suppressors of cytokine signaling. J Cell Sci 2000; 113:2813-2819.
12. Hong F, Jaruga B, Kim WH, Radaeva S, El-Assal ON, Tian Z, Nguyen VA, et al. Opposing roles of STAT1 and STAT3 in T cell-mediated hepatitis: regulation by SOCS. J Clin Invest 2002; 110:1503-1513.
13. Campbell JS, Prichard L, Schaper F, Schmitz J, Stephenson-Famy A, Rosenfeld ME, Argast GM, et al. Expression of suppressors of cytokine signaling during liver regeneration. J Clin Invest 2001:107:1285-1292.
14. Song MM, Shuai K. The suppressor of cytokine signaling (SOCS) 1 and S0CS3 but not S0CS2 proteins inhibit interferon-mediated antiviral and antiproliferative activities. J Biol Chem 1998; 273:35056 - 35062.
15. David M, Chen HE, Goelz S, Larner AC, Neel BG. Differential regulation of the alpha/beta interferon-stimulated Jak/Stat pathway by the SH2 domain-containing tyrosine phosphatase SHPTPl. Mol Cell Biol 1995; 15:7050 -7058.
16. Morita K, Tanaka K, Saito S, Kitamura T, Kiba T, Fujii T, Numata K, et al. Expression of interferon receptor genes in the liver as a predictor of interferon response in patients with chronic hepatitis C. J Med Virol 1999; 58:359 -365.
17. Mizukoshi E, Kaneko S, Kaji K, Terasaki S, Matsushita E, Muraguchi M, Ohmoto Y, et al. Serum levels of soluble interferon Alfa/Beta receptor as an inhibitory factor of interferon in the patients with chronic hepatitis C. HEPATOLOGY 1999:30:1325-1331.
18. Larrea E, Garcia N, QianC, CiveiraMP, Prieto J. Tumor necrosis factor alpha gene expression and the response to interferon in chronic hepatitis C. HEPATOLOGY 1996:23:210 -217.
19. Neuman MG, Benhamou JP, Malkiewicz IM, Akremi R, Shear NH, Asselah T, Ibrahim A, et al. Cytokines as predictors for sustained response and as markers for immunomodulation in patients with chronic hepatitis C. Clin Biochem 2001:34:173-182.
20. Nelson DR, Tu Z, Soldevila-Pico C, Abdelmalek M, Zhu H, Xu YL, Cabrera R, et al. Long-term interleukin 10 therapy in chronic hepatitis C patients has a proviral and anti-inflammatory effect. HEPATOLOGY 2003; 38:859-868.
21. Edwards-Smith CJ, Jonsson JR, Purdie DM, Bansal A, Shorthouse C, Powell EE. Interleukin-10 promoter polymorphism predicts initial response of chronic hepatitis C to interferon alfa. HEPATOLOGY 1999;30: 526 - 530.
22. Yee LJ, Tang J, Gibson AW, Kimberly R, Van Leeuwen DJ, Kaslow RA. Interleukin 10 polymorphisms as predictors of sustained response in antiviral therapy for chronic hepatitis C infection. HEPATOLOGY 2001:33:708-712
23. Kishihara Y, Hayashi J, Yoshimura E, Yamaji K, Nakashima K, Kashiwagi S. IL—1 beta and TNF-alpha produced by peripheral blood mononuclear cells before and during interferon therapy in patients with chronic hepatitis C. Dig Dis Sci 1996:41:315-321.
24. Tian Z, Shen X, Feng H, Gao B. IL-1 beta attenuates IFN-alpha beta induced antiviral activity and STAT1 activation in the liver: involvement of proteasome-dependent pathway. J Immunol 2000; 165:3959 - 3965.
25. Thimme R, Bukh J, Spangenberg HC, Wieland S, Pemberton J, Steiger C, Govindarajan S, et al. Viral and immunological determinants of hepatitis C virus clearance, persistence, and disease. Proc Natl Acad Sci U S A 2002; 99:15661 -15668.
26. Saez-Royuela F, Porres JC, Moreno A, Castillo I, Martinez G, Galiana F, Carreno V. High doses of recombinant alpha-interferon or gamma-interferon for chronic hepatitis C: a randomized, controlled trial. HEPATOLOGY 1991; 13:327-331.
27. Katayama K, Kasahara A, Sasaki Y, Kashiwagi T, Naito M, Masuzawa M, Katoh M, et al. Immunological response to interferon-gamma priming prior to interferon- alpha treatment in refractory chronic hepatitis C in relation to viral clearance. J Viral Hepat 2001; 8:180 -185.
28. Dumoulin FL, Wennrich U, Nischalke HD, Leifeld L, Fischer HP, Sauerbruch T, Spengler U. Intrahepatic mRNA levels of interferon gamma and tumor necrosis factor alpha and response to antiviral treatment of chronic hepatitis C. J Hum Virol 2001;4:195 - 199.
29. Polyak SJ, Khabar KS, Rezeiq M, Gretch DR. Elevated levels of interleukin- 8 in serum are associated with hepatitis C virus infection and resistance to interferon therapy. J Virol 2001; 75:6209-6211.
30. 69. Khabar KS, Al-Zoghaibi F, Al-Ahdal MN, Murayama T, Dhalla M, Mukaida N, Taha M, et al. The alpha chemokine, interleukin 8, inhibits the antiviral action of interferon alpha. J Exp Med 1997:186:1077-1085.
31. Promrat K, McDermott DH, Gonzalez CM, Kleiner DE, Koziol DE, Lessie M, Merrell M, et al. Associations of chemokine system polymorphisms with clinical outcomes and treatment responses of chronic hepatitis C. Gastroenterology 2003:124:352-360.
32. Hayashi J, Kishihara Y, Ueno K, Yamaji K, Kawakami Y, Furusyo N, Sawayama Y, et al. Age-related response to interferon alfa treatment in women vs men with chronic hepatitis C virus infection. Arch Intern Med 1998:158:177-181.
33. Layden-Almer JE, Ribeiro RM, Wiley T, Perelson AS, Layden TJ. Viral dynamics and response differences in HCV-infected African American and white patients treated with IFN and ribavirin. HEPATOLOGY 2003; 37:1343-1350.
34. Sim H, Wojcik J, Margulies M, Wade JA, Heathcote J. Response to interferon therapy: influence of human leucocyte antigen alleles in patients with chronic hepatitis C. J Viral Hepat 1998; 5:249 -253.
35. Wawrzynowicz-Syczewska M, Underhill JA, Clare MA, Boron-Kaczmarska A, McFarlane IG, Donaldson PT. HLA class II genotypes associated with chronic hepatitis C virus infection and response to alpha-interferon treatment in Poland. Liver 2000; 20:234 -239.
36. Romero-Gomez M, Gonzalez-Escribano MF, Torres B, Barroso N, Montes- Cano MA, Sanchez-Munoz D, Nunez-Roldan A, et al. HLA class I B44 is associated with sustained response to interferon_ribavirin therapy in patients with chronic hepatitis C.Am J Gastroenterol 2003:98:1621-1626.
37. McCullough AJ. Obesity and its nurturing effect on hepatitis C. HEPATOLOGY 2003; 38:557-559.
38. Bressler BL, Guindi M, Tomlinson G, Heathcote J. High body mass index is an independent risk factor for nonresponse to antiviral treatment in chronic hepatitis C. HEPATOLOGY 2003:38:639-644.
39. Banner BF, Barton AL, Cable EE, Smith L, Bonkovsky HL. A detailed analysis of the Knodell score and other histologic parameters as predictors of response to interferon therapy in chronic hepatitis C. Mod Pathol 1995; 8:232-238.
40. Myers RP, Patel K, Pianko S, Poynard T, McHutchison JG. The rate of fibrosis progression is an independent predictor of the response to antiviral therapy in chronic hepatitis C. J Viral Hepat 2003; 10:16 -22.
41. Farci, P. Hepatitis C virus. The importance of viral heterogeneity. Clin Liver Dis 2001; 5, 895-916
42. Farci, P. et al. Early changes in hepatitis C viral quasispecies during interferon therapy predict the therapeutic outcome. Proc Natl Acad Sci USA 2002; 99, 3081 -3086
43. Farci, P. et al. The outcome of acute hepatitis C predicted by the evolution of the viral quasispecies. Science 2000-, 288, 339 - 344.
44. Enomoto, N. et al. Mutations in the nonstructural protein 5A gene and response to interferon in patients with chronic hepatitis C virus 1b infection. N Engl J Med 1996; 334, 77-81
45. Schinkel, J., Spoon, W. J. & Kroes, A. C. Meta-analysis of mutations in the NS5A gene and hepatitis C virus resistance to interferon therapy: uniting discordant conclusions. Antivir Ther 2004; 9, 275 - 286.
46. Geiss, G. K. et al. Gene expression profiling of the cellular transcriptional network regulated by α/β interferon and its partial attenuation by the hepatitis C virus nonstructural 5A protein. J Virol 2003; 77, 6367-6375.
47. Lin W, Choe WH, Hiasa Y, Kamegaya Y, Blackard JT, Schmidt EV, Chung RT. Hepatitis C virus expression suppresses interferon signaling by degrading STAT1.Gastroenterology. 2005; 128(4):1034-41.
48. Bode JG, Ludwig S, Ehrhardt C, Albrecht U, Erhardt A, Schaper F, Heinrich PC, Haussinger D. IFN-alpha antagonistic activity of HCV core protein involves induction of suppressor of cytokine signaling-3. FASEB J. 2003; 17(3):488-90.
49. Melen K, Fagerlund R, Nyqvist M, Keskinen P, Julkunen I Expression of hepatitis C virus core protein inhibits interferon-induced nuclear import of STATs. J Med Virol. 2004; 73(4):536-47.
50. Duong FH, Filipowicz M, Tripodi M, La Monica N, Heim MH. Hepatitis C virus inhibits interferon signaling through up-regulation of protein phosphatase 2A. Gastroenterology. 2004;126(1):263-77.
51. Gale, M. Jr et al. Control of PKR protein kinase by hepatitis C virus nonstructural 5A protein: molecular mechanisms of kinase regulation. Mol Cell Biol 1998; 18, 5208-5218
52. Taylor, D. R., Shi, S. T., Romano, P. R., Barber, G. N. & Lai, M. M. C. Inhibition of the interferon inducible protein kinase PKR by HCV E2 protein. Science 1999; 285, 107-110.
53. Au, W. C., Yeow, W. S. & Pitha, P. M. Analysis of functional domains of interferon regulatory factor 7 and its association with IRF-3. Virology 2001; 280, 273-282.
54. Breiman, A. et al. Inhibition of RIG-I-dependent signaling to the interferon pathway during hepatitis C virus expression and restoration of signaling by IKK_. J. Virol. 2005; 79, 3969-3978.
55. Han, J. Q., Wroblewski, G., Xu, Z., Silverman, R. H. & Barton, D. J. Sensitivity of hepatitis C virus RNA to the antiviral enzyme ribonuclease L is determined by a subset of efficient cleavage sites. J. Interferon Cytokine Res. 2004; 24, 664 - 676.