丙型肝炎病毒与其合并人类免疫缺陷病毒感染者的病毒基因型差异

HCV与HIV具有相似的传播途径,都可以通过使用污染的血制品、注射用具、性接触及母婴传播,因此,HIV与HCV混合感染较常见.HIV感染改变HCV的自然病程,加速肝纤维化、肝硬化及肝细胞癌的进程已达成共识;HCV基因型对肝病的进程和预后亦起到重要作用.那么HCV基因型的分布在HⅣ/HCV混合感染与HCV单纯感染之间是否存在差异?各自的感染途径有何不同?值得我们作进一步的探讨.我们对昆明市第三人民医院收治的昆明地区85例HCV感染住院患者采用PCR等分子生物学方法研究了HIV/HCV感染与HCV单纯感染患者HCV基因型分布的差异.     

HIV/HCV混合感染者与HCV感染者感染HCV基因型分析*

目的 使用二代测序技术检测广州地区人类免疫缺陷病毒（HIV）/丙型肝炎病毒（HCV）混合感染者和HCV感染者HCV基因型,了解本地区HCV基因型的流行特点。方法 在233例HCV感染者中,包括95例HIV/HCV混合感染者和138例HCV感染者,使用Illumina Miseq平台PE300模式对HCV Core和NS5B片段进行测序,采用生物信息学分析确定HCV基因型,对测序结果提示混合基因型感染的样本,使用NS5A基因型特异引物进行验证。结果 HIV/HCV混合感染者静脉吸毒感染构成比显著高于HCV感染者（77.7%对19.6% P<0.001）,HCV感染者输血感染的构成比显著高于HIV/HCV混合感染者（48.6%对3.2% P<0.001）;HIV/HCV混合感染者HCV Core基因片段平均数据量为（15456±6689） reads,HCV感染者为（14323±5321） reads,两组无显著差异（P>0.05）;HIV/HCV混合感染者HCV NS5B基因片段平均数据量为（16432±3467） reads,HCV感染者为（17611±5632） reads,也无显著差异（P>0.05）;本组228例（97.9%）HCV感染者为单一HCV基因型感染,HIV/HCV混合感染者HCV 6a型和3b型感染率显著高于HCV感染者（分别为47.4%对26.8%,P=0.001和13.7%对3.6%,P=0.005）,HCV 1b型感染率显著低于HCV感染者（20.0%对59.4%,P<0.001）;在233例入组的HCV感染者中,发现5例（2.1%）为混合基因型感染,其中4例为HIV/HCV混合感染者,1例为HCV感染者。结论 广州地区HIV/HCV混合感染者与HCV感染者感染HCV基因型构成比存在差异,其临床意义还需要探讨。     

HBV and HCV therapy

One year of interferon therapy inhibits HBV replication in one third of the patients whereas long-term administration of oral nucleos(t)ide analogues is efficient in most of them, as long as early treatment adaptation in patients with partial virological response and resistance is provided. Following the demonstration of a more potent antiviral effect in terms of sustained virological response (SVR) rates, Pegylated-IFN coupled with Ribavirin has become the standard treatment for chronic hepatitis C, with nearly 65% of all treated patients achieving a SVR. Long-term suppression of HBV and eradication of HCV would halt the progression of chronic hepatitis to cirrhosis, hepatocellular carcinoma and liver decompensation.     

HCV and HIV Coinfection

Chronic hepatitis C (CHC) is estimated to affect about 20% of the 34 million individuals currently living with HIV worldwide, with greater rates (~ 75%) in intravenous drug users or persons exposed to blood products. Individuals who are coinfected with HIV and hepatitis C virus (HCV) show an accelerated course of liver disease, with faster progression to cirrhosis and its clinical complications. The combination of pegylated interferon plus ribavirin given for 6–18 months leads to sustained HCV clearance in no more than half of HIV-HCV coinfected patients. Thus, new direct anti-HCV drugs are eagerly awaited for this population. Appropriate diagnosis and monitoring of CHC, including the use of noninvasive tools for assessing liver fibrosis (eg, elastometry) as well as provision of therapy guided by early viral kinetics and IL28B genotyping, are improving the management of CHC in HIV-infected persons.     

HCV and extrahepatic diseases

Besides being a hepatotropic virus and a common cause of chronic hepatitis, hepatitis C virus (HCV) has been linked to a variety of extrahepatic immunological manifestations. The high prevalence of HCV infections in some of these conditions suggests an important pathogenetic role of the virus. The recent observation that HCV infects peripheral blood mononuclear cells, such as CD8+ T lymphocytes, CD19+ B lymphocytes and monocytes/microphages, has given an insight into the possible mechanisms of HCV associated autoimmunity. In the clinical practice it is recommended not only to search for symptoms and signs of autoimmune disorders in patients with chronic hepatitis C, but also to test for hepatitis C virus infection patients with extrahepatic conditions known to be related to HCV. Even if the occurrence of autoimmune disorders or the exacerbations of autoimmune diseases has been reported during interferon therapy, antiviral therapy is effective in treating some of the extrahepatic disorders associated with HCV, namely mixed cryoglobulinemia and membranoproliferative glomerulonephritis. The extrahepatic manifestations associated with hepatitis C virus infection are reviewed according to the available data and the option of interferon therapy is discussed.     

HCV不同功能区抗体与HCVRNA的关系探讨

探讨HCV不同功能区抗体(抗-C、抗-NS3、抗-NS4及抗-NS5)与HCVRNA的关系。HCV不同功能区抗体测定采用ELISA法,用RT-PCR进行HCV RNA检测。结果显示55例抗HCV阳性血清中有7种抗体阳性组合。抗-NS3、抗-C、抗-NS5及抗-NS4在7种阳性组合中的检出率分别为96．4%、89．1%、58．2%、56．4%。HCVRNA阳性血清中抗-NS3、抗-C、抗-NS5及抗NS4的检出率分别为96．2%,88．5%,57．7%,57．7%。HCV不同功能区抗体ELISA法有很高的敏感性和特异性,与RT-PCR法基本一致。抗-NS3、抗-C在HCV的诊断中有重要的意义,抗-NS5和抗-NS4有诊断的互补作用。     

HCV dsRNA-Activated Macrophages Inhibit HCV Replication in Hepatocytes

Background:

Macrophages play critical roles in innate immune response in the liver. Whether macrophages participate in liver innate immunity against HCV replication is poorly understood

Objectives:

The aim of this study was to investigate the role of macrophages in liver innate immunity against HCV replication.

Materials and Methods:

Freshly isolated monocytes were purified from peripheral blood of healthy adult donors. Macrophages refer to 7-day-cultured monocytes in vitro. A hepatoma cell line (Huh7) was infected with HCV JFH-1 to generate in vitro HCV infectious system. RT-PCR was used to determine HCV RNA and mRNA levels of genes expression. ELISA was used to measure the protein level of interferon-α (IFN-α) and western blot was used to determine protein expression level of Toll-like receptor 3 (TLR3).

Results:

HCV dsRNA induced the expression of type I IFN (IFN-α/β) in monocyte-derived macrophages. HCV dsRNA also induced the expression of TLR3 and IFN regulatory factor-7 (IRF-7), the key regulators of the IFN signaling pathway. When HCV JFH-1-infected Huh7 cells were co-cultured with macrophages activated with HCV dsRNA or incubated in media conditioned with supernatant (SN) from HCV dsRNA-activated macrophages, HCV replication was significantly suppressed. This macrophage SN action on HCV inhibition was mediated through type I IFN, which was evidenced by the observation that antibody to type I IFN receptor could neutralize the macrophages-mediated anti-HCV effect. The role of type I IFN in macrophages-mediated anti-HCV activity is further supported by the observation that HCV dsRNA-activated macrophages SN treatment induced the expression of several IFN-stimulated genes (ISGs), ISG15, ISG56, OAS-1, OAS-2, MxA and Viperin in HCV-infected Huh7 cells.

Conclusions:

Macrophages may play an important role in liver innate immunity against HCV replication through a type I IFN-dependent mechanism.     

FQ-PCR和AMPLICOR HCV MONITOR 2.0方法测定HCV病毒载量的比较

目的 探讨荧光定量(FQ-PCR)和AMPLICOR HCV MONITOR 2.0试剂盒检测相同样品中丙型肝炎病毒(HCV)病毒载量值之间的关系.方法 收集84份样品,经ELISA检测抗-HCV,再分别使用两种方法测定HCV病毒载量,利用统计分析软件SPSS 15.0对病毒载量Log对数值进行统计分析.结果 当HCV病毒载量值大于1 000IU/ml时,两种方法检测结果的相关性为0.818(P<0.0001).结论 FQ-PCR和AMPLICOR HCV MONI-TOR 2.0两种方法测定的HCV病毒载量值有较高的相关性.     

HCV抗原、HCV抗体及HCV-RNA联合检测与ALT的相关性及其临床意义

[目的]探讨HCV-RNA阳性丙肝患者核心抗原（HCV-cAg）、抗体（HCV-Ab）以及HCV-RNA含量3种检测指标与丙氨酸氨基转移酶（ALT）水平的相关性及其临床意义.[方法]对100例HCV-RNA阳性丙肝患者应用酶速率法检测ALT含量、ELISA法检测HCV-cAg、CLIA法检测HCV-Ab、FQ-PCR法检测HCV-RNA,并对所得数据进行统计分析.[结果[100例患者血清中HCV-Ab阳性率为97.0％,HCV-cAg阳性率为83.0％;HCV-RNA载体含量越高,HCV-Ab阳性率越高;ALT含量的变化与HCV-RNA载体含量并无明显相关性（P＞0.05）;ALT异常率随着HCV-RNA含量的增高而升高,呈正相关（P＜0.05）.[结论]HCV-Ab检测可反映机体对HCV感染的免疫状态,能间接证实HCV感染;HCV-RNA敏感性和特异性较高,HCV-RNA与HCV-cAg阳性检出率呈现一致性,且ALT异常率随着HCV-RNA含量的增高而升高,可以反映临床病情;3种方法同时检测能准确诊断丙肝病毒的感染,以及预测是否具有传染性,联合ALT可有效预测和评价肝脏损伤和药物疗效.     

不同HCV感染者各区段抗HCV抗体的反应性及HCV-RNA检测结果分析

利用丙型肝炎病毒（ＨＣＶ）不同区段抗原的单片段酶免疫（ＥＩＡ）试剂及反转录聚合酶链反应（ＲＴ－ＰＣＲ）技术,对不同ＨＣＶ感染者血清不同区段抗ＨＣＶ抗体及丙型肝炎病毒核糖核酸（ＨＣＶ－ＲＮＡ）进行检测。结果显示,慢性肝炎患者各抗体检出率及Ｓ／Ｃｏ（Ｓ为样品光密度值,Ｃｏ为临界值）值均高,肝癌患者各抗体检出率及ＨＣＶ－ＲＮＡ阳性率均较低;ＡＬＴ正常而抗ＨＣＶ阳性者各区段抗体阳性率低于急、慢性肝炎患者,而其ＨＣＶ－ＲＮＡ的阳性率高于其他各组;抗－ＮＳ３和抗－Ｃ抗体在各组的检出率和反应性均强。表明抗－Ｃ和抗－ＮＳ３抗体的检测对ＨＣＶ感染最有诊断价值,同时用ＲＴ－ＰＣＲ技术检测血清ＨＣＶ－ＲＮＡ有助于ＨＣＶ感染的早期诊断     

HCV和HIV-1合并感染标本中HCV基因分型

目的了解丙型肝炎病毒（ＨＣＶ）和人类免疫缺陷病毒Ｉ型（ＨＩＶ－１）混合感染标本中ＨＣＶ基因型情况。方法采用了根据ＨＣＶ５′端非转译区基因组所设计的反相探针杂交测定方法（ＩＮＮＯＬｉＰＡ，ＨＣＶⅡ），对来自六个地区的１７份抗－ＨＣＶ和抗－ＨＩＶ－１双阳性标本进行ＨＣＶ基因型分型研究。结果在１７份标本中，７份（４１．１８％）为Ｉ（１ｂ）型病毒，５份（２９．４１％）为Ⅱ（２ａ／２ｃ）型病毒，５份（２９．４１％）为Ｉ型（１ｂ）和Ⅱ（２ａ／２ｃ）混合型病毒。结论发现ＨＣＶ与ＨＩＶ－１合并感染标本中存在Ｉ（１ｂ）型、Ⅱ（２ａ／２ｃ）型以及Ｉ（１ｂ）型与Ⅱ（２ａ／２ｃ）型混合型ＨＣＶ。值得注意的是对α－干扰素治疗不敏感并且较易趋于导致严重的慢性肝炎、肝硬化和肝细胞癌的Ⅱ型ＨＣＶ感染的比率占７０．５８％（１２／１７）。     

Factors influencing liver fibrosis and necroinflammation in HIV/HCV coinfection and HCV monoinfection

Objectives

To define differences in liver histology between HIV/HCV coinfection and HCV monoinfection, and to investigate possible causative factors.

Methods

Liver biopsies (LBs) from 440 consecutive HIV/HCV-coinfected patients (Group HIV/HCV) and 374 consecutive HCV-monoinfected patients (Group HCV) were evaluated for necroinflammation and fibrosis (Ishak) by a pathologist unaware of the clinical and laboratory data. All patients were HBsAg-negative, with no history of alcohol abuse and naïve to anti-HCV treatment. At LB, 78.4 % of patients in Group HIV/HCV were on an antiretroviral regimen.

Results

HIV/HCV-coinfected patients compared to the HCV-monoinfected patients were younger (p < 0.0001), more frequently males (p < 0.0001), and had HCV genotype 3 (p < 0.0001); they showed a good immunological condition (CD4+ cell count: 518 ± 166 cells/mm³). Patients in Group HIV/HCV more frequently showed a fibrosis score ≥4 (27.5 vs. 20.6 %, p < 0.05) and a necroinflammation score ≥9 (25.9 vs. 13.4 %; p < 0.0001). The prevalence of patients with fibrosis score ≥4 was significantly higher in older age classes in both Group HIV/HCV (p < 0.005) and Group HCV (p < 0.05). A necroinflammation score ≥9 was significantly higher in older age classes only in Group HIV/HCV (p < 0.05). A multivariate analysis for Group HIV/HCV revealed that the patient age and nadir of CD4+ cell count were independently associated to higher degrees of fibrosis, the patient age and antiretroviral treatment were associated to higher degrees of necroinflammation, and HCV genotype 3 was associated to higher degrees of steatosis.

Conclusion

The data suggest a need for early anti-HCV treatment in both HCV-monoinfected and HIV/HCV-coinfected patients.