慢性丙型肝炎患者中丙型肝炎病毒准种变异研究进展

丙型肝炎病毒（HCV）具有较高的变异性,通常以准种的形式分布在感染者的体内,病毒容易逃离机体的免疫监控,因而无法被有效地清除,导致机体很难控制其感染的发展,故易转变成慢性肝炎。HCV准种变异在宿主体内的持续存在对病毒感染的控制、抗病毒药物和疫苗的发展都是一个巨大的挑战。,我们重点阐述近年来关于HCV准种变异及其与慢性丙型肝炎患者的机体免疫、疾病进展、治疗效果之间关系的研究进展。     

Altered Functionality of Anti-Bacterial Antibodies in Patients with Chronic Hepatitis C Virus Infection

Background

Using comparative glycoproteomics, we have previously identified a glycoprotein that is altered in both amount and glycosylation as a function of liver cirrhosis. The altered glycoprotein is an agalactosylated (G0) immunoglobulin G molecule (IgG) that recognizes the heterophilic alpha-gal epitope. Since the alpha gal epitope is found on gut enterobacteria, it has been hypothesized that anti-gal antibodies are generated as a result of increased bacterial exposure in patients with liver disease.

Methods

The N-linked glycosylation of anti-gal IgG molecules from patients with fibrosis and cirrhosis was determined and the effector function of anti-bacterial antibodies from over 100 patients examined. In addition, markers of microbial exposure were determined.

Results

Surprisingly, the subset of agalactosylated anti-gal antibodies described here, was impaired in their ability to mediate complement mediated lysis and inhibited the complement-mediated destruction of common gut bacteria. In an analysis of serum from more than 100 patients with liver disease, we have shown that those with increased levels of this modified anti-gal antibody had increased levels of markers of bacterial exposure.

Conclusions

Anti-gal antibodies in patients with liver cirrhosis were reduced in their ability to mediate complement mediated lysis of target cells. As bacterial infection is a major complication in patients with cirrhosis and bacterial products such as LPS are thought to play a major role in the development and progression of liver fibrosis, this finding has many clinical implications in the etiology, prognosis and treatment of liver disease.     

Longitudinal Liver Stiffness Assessment in Patients with Chronic Hepatitis C Undergoing Antiviral Therapy

Background/Aims

Liver stiffness (LS) measurement by means of transient elastography (TE) is accurate to predict fibrosis stage. The effect of antiviral treatment and virologic response on LS was assessed and compared with untreated patients with chronic hepatitis C (CHC).

Methods

TE was performed at baseline, and at weeks 24, 48, and 72 in 515 patients with CHC.

Results

323 treated (62.7%) and 192 untreated patients (37.3%) were assessed. LS experienced a significant decline in treated patients and remained stable in untreated patients at the end of study (P<0.0001). The decline was significant for patients with baseline LS ≥ 7.1 kPa (P<0.0001 and P 0.03, for LS ≥9.5 and ≥7.1 kPa vs lower values, respectively). Sustained virological responders and relapsers had a significant LS improvement whereas a trend was observed in nonresponders (mean percent change −16%, −10% and −2%, for SVR, RR and NR, respectively, P 0.03 for SVR vs NR). In multivariate analysis, high baseline LS (P<0.0001) and ALT levels, antiviral therapy and non-1 genotype were independent predictors of LS improvement.

Conclusions

LS decreases during and after antiviral treatment in patients with CHC. The decrease is significant in sustained responders and relapsers (particularly in those with high baseline LS) and suggests an improvement in liver damage.     

Significant Fibrosis Is Not Rare in Chinese Chronic Hepatitis B Patients with Persistent Normal ALT

Background

Limited studies have been done on chronic hepatitis B (CHB) patients defined according to the latest Asian-Pacific Association for the Study of the Liver guideline with liver histology by a large sample size.

Methods

We retrospectively evaluated liver histological characteristics on a cohort of consecutive treatment-naive CHB patients with persistent normal alanine aminotransferase (PNALT) or elevated ALT from May 2005 to October 2011. Histological assessment was based on the Metavir scoring system, significant abnormality was defined as necroinflammation grade ≥A2 and/or fibrosis stage ≥F2.

Results

A total of 675 CHB patients were recruited, including 516 HBeAg-positive and 159 HBeAg-negative patients. In HBeAg-positive patients, significant fibrosis was found 49.4% (42/85) in PNALT, 69.8% (88/126) in ALT 1-2×upper limit normal (ULN) and 81.6% (249/305) in ALT>2×ULN group, respectively. In HBeAg-negative patients, significant fibrosis was found 30.9% (17/55) in PNALT, 73.3% (33/45) in ALT 1-2×ULN and 94.9% (56/59) in ALT>2×ULN group, respectively. HBeAg-positive patients with PNALT over 30 years old had a higher frequency of significant fibrosis than those under 30 years old (87.5% vs. 45.5%, P = 0.058). Multivariate logistic regression analysis indicated increasing age (P = 0.012), higher aspartate aminotransferase (AST) (P < 0.001) and lower HBV DNA (P < 0.001) were associated with significant necroinflammation, while higher AST (P < 0.001), lower albumin (P = 0.027) and HBV DNA (P = 0.004) were associated with significant fibrosis in HBeAg-positive patients with elevated ALT. Higher AST was associated with significant necroinflammation in HBeAg-negative patients with elevated ALT (P = 0.009).

Conclusions

Significant fibrosis is not rare in Chinese CHB patients with PNALT, especially HBeAg-positive patients over 30 years old.     

Effect of Retreatment with Interferon Alone or Interferon plus Ribavirin on Hepatitis C Virus Quasispecies Diversification in Nonresponder Patients with Chronic Hepatitis C

Alpha interferon (IFN-alpha) treatment is effective on a long-term basis in only 15 to 25% of patients with chronic hepatitis C. The results of recent trials indicate that response rates can be significantly increased when IFN-alpha is given in combination with ribavirin. However, a large number of patients do not respond even to combination therapy. Nonresponsiveness to IFN is characterized by evolution of the hepatitis C virus (HCV) quasispecies. Little is known about the changes occurring within the HCV genomes when nonresponder patients are retreated with IFN or with IFN plus ribavirin. In the present study we have examined the genetic divergence of HCV quasispecies during unsuccessful retreatment with IFN or IFN plus ribavirin. Fifteen nonresponder patients with HCV-1 (4 patients with HCV-1a and 11 patients with HCV-1b) infection were studied while being retreated for 2 months (phase 1) with IFN-alpha (6 MU given three times a week), followed by IFN plus ribavirin or IFN alone for an additional 6 months (phase 2). HCV quasispecies diversification in the E2 hypervariable region-1 (HVR1) and in the putative NS5A IFN sensitivity determining region (ISDR) were analyzed for phase 1 and phase 2 by using the heteroduplex tracking assay and clonal frequency analysis techniques. A major finding of this study was the relatively rapid evolution of the HCV quasispecies observed in both treatment groups during the early phase 1 compared to the late phase 2 of treatment. The rate of quasispecies diversification in HVR1 was significantly higher during phase 1 versus phase 2 both in patients who received IFN plus ribavirin (P = 0.017) and in patients who received IFN alone (P = 0. 05). A trend toward higher rates of quasispecies evolution in the ISDR was also observed during phase 1 in both groups, although the results did not reach statistical significance. However, the NS5A quasispecies appeared to be rather homogeneous and stable in most nonresponder patients, suggesting the presence of a single well-fit major variant, resistant to antiviral treatment, in agreement with published data which have identified an IFN sensitivity determinant region within the NS5A. During the entire 8 months of retreatment, there was no difference in the rate of fixation of mutation between patients who received combination therapy and patients who were treated with IFN alone, suggesting that ribavirin had no major effects on the evolution of the HCV quasispecies after the initial 2 months of IFN therapy.     

Glucose Tolerance and Insulin Secretion in Patients with Chronic Duodenal Ulcer

An oral glucose tolerance test has been used to compare carbohydrate metabolism in patients suffering from chronic duodenal ulceration with that in a matched group of normal subjects. A significantly greater rise in blood glucose concentration was observed in the duodenal ulcer group, and this was associated with a significantly greater output of insulin. The reason for these findings may be an abnormal secretion of one or more of the small-bowel hormones.     

Longitudinal Evaluation of the Structure of Replicating and Circulating Hepatitis C Virus Quasispecies in Nonprogressive Chronic Hepatitis C Patients

In previous cross-sectional studies, we demonstrated that, in most patients with chronic hepatitis C, the composition and complexity of the circulating hepatitis C virus (HCV) population do not coincide with those of the virus replicating in the liver. In the subgroup of patients with similar complexities in both compartments, the ratio of quasispecies complexity in the liver to that in serum (liver/serum complexity ratio) of paired samples correlated with disease stage. In the present study we investigated the dynamic behavior of viral population parameters in consecutive paired liver and serum samples, obtained 3 to 6 years apart, from four chronic hepatitis C patients with persistently normal transaminases and stable liver histology. We sequenced 359 clones of a genomic fragment encompassing the E2(p7)-NS2 junction, in two consecutive liver-serum sample pairs from the four patients and in four intermediate serum samples from one of the patients. The results show that the liver/serum complexity ratio is not stable but rather fluctuates widely over time. Hence, the liver/serum complexity ratio does not identify a particular group of patients but a particular state of the infecting quasispecies. Phylogenetic analysis and signature mutation patterns showed that virtually all circulating sequences originated from sequences present in the liver specimens. The overall behavior of the circulating viral quasispecies appears to originate from changes in the relative replication kinetics of the large mutant spectrum present in the infected liver.     

乙型肝炎病人重叠感染丙型肝炎的评价

应用ＥＬＩＳＡ和ＰＣＲ法检测５０２例乙肝病人血清,４０１例ＨＢｓＡｇ阳性血清中,有１１４例（２８．４％）抗－ＨＣＶ和ＨＣＶＲＮＡ双项阳性,２５例（６．２％）ＨＣＶＲＮＡ单项阳性;２１例（５．２％）抗－ＨＣＶ单项阳性。将ＨＢｓＡｇ乙肝病人分成ＨＢＶＤＮＡ,ＨＢｅＡｇ阳性组和ＨＢＶＤＮＡ,ＨＢｅＡｇ阴性组。前者抗－ＨＣＶ阳性率为１１．６％～２０．５％,ＨＣＶＲＮＡ阳性率为１６．２％～２０．５％。后者抗－ＨＣＶ阳性率为２０．２％～５５．６％,ＨＣＶＲＮＡ阳性率为２３％～６０．３％。结果说明长期携带ＨＢＶ者和慢性乙肝病人均可重叠ＨＣＶ感染。ＨＢＶＤＮＡ阳性组抗－ＨＣＶ和ＨＣＶＲＮＡ阳性率明显高于ＨＢＶＤＮＡ阳性组     

IP-10 Can Be Measured in Dried Plasma Spots in Patients with Chronic Hepatitis C Infection

The chemokine IP-10 (CXCL10) is a candidate marker for hepatitis C virus (HCV) fibrosis monitoring. The aim of this proof-of-concept study is to assess if IP-10 measurements from dried plasma spots (DPS) are accurate in HCV-infected patients with either minimal or significant fibrosis. We measured IP-10 levels in plasma and DPS of 21 HCV-infected patients with cirrhosis and 19 patients with no/little fibrosis (determined with FibroScan). Cirrhotic patients had significantly higher levels of IP-10 compared to patients with minimal fibrosis. DPS and plasma measurements of IP-10 are comparable and the correlation was excellent (r² = 0.97, p<0.0001). The DPS based method for IP-10 detection performs well in HCV-infected patients with either minimal or significant fibrosis.     

Markers of Collagen Remodeling Detect Clinically Significant Fibrosis in Chronic Hepatitis C Patients

Background and Aim

Detection of advanced fibrosis (Metavir F≥3) is important to identify patients with a high urgency of antiviral treatments vs. those whose treatment could be deferred (F≤2). The aim was to assess the diagnostic value of novel serological extracellular matrix protein fragments as potential biomarkers for clinically significant and advanced fibrosis.

Methods

Specific protein fragments of matrix metalloprotease degraded type I, III, IV and VI collagen (C1M, C3M, C4M, C6M) and type III and IV collagen formation (Pro-C3 and P4NP7S) were assessed in plasma from 403 chronic hepatitis C patients by specific ELISAs. Patients were stratified according to Metavir Fibrosis stage; F0 (n = 46), F1 (n = 161), F2 (n = 95), F3 (n = 44) and F4 (n = 33) based on liver biopsy.

Results

Pro-C3 was significantly elevated in patients with significant fibrosis (≥F2) compared to F0-F1 (p<0.05), while the markers C3M, C4M, C6M and P4NP7S were significantly elevated in patients with advanced fibrosis (≥F3) compared to F0-F2 (p<0.05). C1M showed no difference between fibrosis stages. Using Receiver Operating Characteristics analysis, the best marker for detecting ≥F2 and ≥F3 was Pro-C3 with AUC = 0.75 and AUC = 0.86. Combination of Pro-C3 and C4M with age, BMI and gender in a multiple ordered logistic regression model improved the diagnostic value for detecting ≥F2 and ≥F3 with AUC = 0.80 and AUC = 0.88.

Conclusion

The Pro-C3 protein fragment provided clinically relevant diagnostic accuracy as a single marker of liver fibrosis. A model combining Pro-C3 and C4M along with patient’s age, body mass index and gender increased the diagnostic power for identifying clinically significant fibrosis.     

Cost-Effectiveness Analysis of Sofosbuvir Compared to Current Standard Treatment in Swiss Patients with Chronic Hepatitis C

In clinical trials, sofosbuvir showed high antiviral activity in patients infected with hepatitis C virus (HCV) across all genotypes. We aimed to determine the cost-effectiveness of sofosbuvir-based treatment compared to current standard treatment in mono-infected patients with chronic hepatitis C (CHC) genotypes 1–4 in Switzerland. Cost-effectiveness was modelled from the perspective of the Swiss health care system using a lifetime Markov model. Incremental cost-effectiveness ratios (ICERs) used an endpoint of cost per quality-adjusted life year (QALY) gained. Treatment characteristics, quality of life, and transition probabilities were obtained from published literature. Country-specific model inputs such as patient characteristics, mortality and costs were obtained from Swiss sources. We performed extensive sensitivity analyses. Costs and effects were discounted at 3% (range: 0–5%) per year. Sofosbuvir-containing treatment in mixed cohorts of cirrhotic and non-cirrhotic patients with CHC genotypes 1–4 showed ICERs between CHF 10,337 and CHF 91,570 per QALY gained. In subgroup analyses, sofosbuvir dominated telaprevir- and boceprevir-containing treatment in treatment-naïve genotype 1 cirrhotic patients. ICERs of sofosbuvir were above CHF 100,000 per QALY in treatment-naïve, interferon eligible, non-cirrhotic patients infected with genotypes 2 or 3. In deterministic and probabilistic sensitivity analyses, results were generally robust. From a Swiss health care system perspective, treatment of mixed cohorts of cirrhotic and non-cirrhotic patients with CHC genotypes 1–4 with sofosbuvir-containing treatment versus standard treatment would be cost-effective if a threshold of CHF 100,000 per QALY was assumed.     

An Interferon Response Gene Signature Is Associated with the Therapeutic Response of Hepatitis C Patients

Infection with the hepatitis C virus (HCV) is a major cause of chronic liver diseases and hepatocellular carcinoma worldwide, and thus represents a significant public health problem. The type I interferon (IFN), IFNα, has been successful in treating HCV-infected patients, but current IFN-based treatment regimens for HCV have suboptimal efficacy, and relatively little is known about why IFN therapy eliminates the virus in some patients but not in others. Therefore, it is critical to understand the basic mechanisms that underlie the therapeutic resistance to IFN action in HCV-infected individuals, and there is an urgent need to identify those patients most likely to respond to IFN therapy for HCV. To characterize the response of HCV-infected patients to treatment with IFNα, the expression of an IFN-response gene signature comprised of IFN-stimulated genes and genes that play an important role in the innate immune response was examined in liver biopsies from HCV-infected patients enrolled in a clinical trial. In the present study we found that the expression of a subset of IFN-response genes was dysregulated in liver biopsy samples from nonresponsive hepatitis C patients as compared with virologic responders. Based on these findings, a statistical model was developed to help predict the response of patients to IFN therapy, and compared to results obtained to the IL28 mutation model, which is highly predictive of the response to IFN-based therapy in HCV-infected patients. We found that a model incorporating gene expression data can improve predictions of IFN responsiveness compared to IL28 mutation status alone.     

IL28B Polymorphism Correlates with Active Hepatitis in Patients with HBeAg-Negative Chronic Hepatitis B

Background & Aims

The clinical relevance of single nucleotide polymorphisms (SNPs) near the IL28B gene is controversial in patients with hepatitis B virus (HBV) infection. This study aimed to investigate the role of viral and host factors, including IL28B genotypes, in the natural course of chronic hepatitis B (CHB).

Methods

The study enrolled consecutive 115 treatment-naive CHB patients. HBV viral loads, genotypes, precore and basal core promotor mutations, serum hepatitis B surface antigen (HBsAg) and interferon-gamma inducible protein 10 (IP-10) levels as well as four SNPs of IL28B were determined. Serial alanine transaminase (ALT) levels in the previous one year before enrollment at an interval of three months were recorded. Factors associated with active hepatitis, defined as persistent ALT >2× upper limit of normal (ULN) or a peak ALT level >5× ULN, were evaluated.

Results

The prevalence of rs8105790 TT, rs12979860 CC, rs8099917 TT, and rs10853728 CC genotypes were 88.3%, 87.4%, 88.4% and 70.9%, respectively. In HBeAg-positive patients (n = 48), HBV viral load correlated with active hepatitis, while in HBeAg-negative patients (n = 67), rs10853728 CC genotype (p = 0.032) and a trend of higher IP-10 levels (p = 0.092) were associated with active hepatitis. In multivariate analysis, high viral load (HBV DNA >10⁸ IU/mL, p = 0.042, odds ratio = 3.946) was significantly associated with HBeAg-positive hepatitis, whereas rs10853728 CC genotype (p = 0.019, odds ratio = 3.927) was the only independent factor associated with active hepatitis in HBeAg-negative population.

Conclusions

HBV viral load and IL28B rs10853728 CC genotype correlated with hepatitis activity in HBeAg-positive and HBeAg-negative CHB, respectively. Both viral and host factors play roles in disease activity during different phases of CHB.     

Growth Arrest Specific 2 Is Up-Regulated in Chronic Myeloid Leukemia Cells and Required for Their Growth

Although the generation of BCR-ABL is the molecular hallmark of chronic myeloid leukemia (CML), the comprehensive molecular mechanisms of the disease remain unclear yet. Growth arrest specific 2 (GAS2) regulates multiple cellular functions including cell cycle, apoptosis and calpain activities. In the present study, we found GAS2 was up-regulated in CML cells including CD34⁺ progenitor cells compared to their normal counterparts. We utilized RNAi and the expression of dominant negative form of GAS2 (GAS2DN) to target GAS2, which resulted in calpain activity enhancement and growth inhibition of both K562 and MEG-01 cells. Targeting GAS2 also sensitized K562 cells to Imatinib mesylate (IM). GAS2DN suppressed the tumorigenic ability of MEG-01 cells and impaired the tumour growth as well. Moreover, the CD34⁺ cells from CML patients and healthy donors were transduced with control and GAS2DN lentiviral vectors, and the CD34⁺ transduced (YFP⁺) progeny cells (CD34⁺YFP⁺) were plated for colony-forming cell (CFC) assay. The results showed that GAS2DN inhibited the CFC production of CML cells by 57±3% (n = 3), while affected those of normal hematopoietic cells by 31±1% (n = 2). Next, we found the inhibition of CML cells by GAS2DN was dependent on calpain activity but not the degradation of beta-catenin. Lastly, we generated microarray data to identify the differentially expressed genes upon GAS2DN and validated that the expression of HNRPDL, PTK7 and UCHL5 was suppressed by GAS2DN. These 3 genes were up-regulated in CML cells compared to normal control cells and the growth of K562 cells was inhibited upon HNRPDL silence. Taken together, we have demonstrated that GAS2 is up-regulated in CML cells and the inhibition of GAS2 impairs the growth of CML cells, which indicates GAS2 is a novel regulator of CML cells and a potential therapeutic target of this disease.     

肝内HDAg阳性慢性肝炎病理特点探讨

用直接酶标和直接免疫荧光法在120例HBsAg阳性肝组织中检测HDAg,发现5例阳性(4.2％),其中3例活动性肝硬化,2例慢性中型活动性肝炎。另设立相对应的单纯乙型肝炎作为对照,通过光镜、电镜、免疫组化的方法,重点探讨比较丁型肝炎与单纯乙型肝炎二者肝组织学的形态变化。丁型肝炎组肝细胞灶状坏死,桥接坏死,融合性坏死的程度比单纯乙型肝炎组重,丁型肝炎组较单纯乙型肝炎组常见的组织学改变是肝细胞呈灶状微小空泡的脂肪变性及灶状嗜酸性变性。HDAg阳性肝细胞呈疏松化、核固缩,其周围未见淋巴细胞浸润。     

A Study of the Immunoloregulation of Double Filtration Plasmapheresis in Maintenance Hemodialysis Patients with Chronic Hepatitis C

Although a large number of drugs have been used to treat chronic hepatitis C (CHC), there still remains a great challenge to treat maintenance hemodialysis (MHD) patients with chronic hepatitis C. To clarify the immunnoloregulation of double filtration plasmapheresis (DFPP) in MHD patients with CHC, DFPP was performed in 20 MHD patients with CHC (HCV-antibody positive, serum HCV RNA >500 IU/ml more than 6 months and HCV genotype 1b). The clinical data was collected and peripheral blood mononuclear cells (PBMCs) were analyzed by flow cytometry at the time of hour 0, hour 3, day 3, day 7 and day 28 after the DFPP, respectively. Serum HCV particles could be removed partially by the DFPP. The titer of serum HCV RNA could remain in a lower level even 28 days after the treatment. Compared to MHD patients without HCV infection, the frequencies of innate immune cells were similar in MHD patients with CHC, while Th1/Th2 was elevated and the frequencies of regulatory T (Treg) cells were higher in those MHD patients with CHC. The frequencies of monocytes and natural killer (NK) cells remained after the DFPP in MHD patients with CHC. There were no significant changes of Th1, Th2 and Th1/Th2 in PBMC after DFPP. DFPP could reduce the frequencies of Th17 cells and Treg cells in PBMC from 7 days after DFPP in MHD patients with CHC. DFPP could partially remove the serum HCV particles mechanically. The titer of HCV RNA could remain in a lower level at least for 28 days probably due to the redistribution of the immunocytes in circulation.     

Immune Activation Response in Chronic HIV-Infected Patients: Influence of Hepatitis C Virus Coinfection

Objectives

We have analyzed the parameters (bacterial translocation, immune activation and regulation, presence of HCV coinfection) which could be implicated in an inappropriate immune response from individuals with chronic HIV infection. The influence of them on the evolution of CD4+ T cell count has been investigated.

Patients and methods

Seventy HIV-infected patients [monoinfected by HIV (n = 20), HCV-coinfected (with (n = 25) and without (n = 25) liver cirrhosis)] and 25 healthy controls were included. Median duration of HIV infection was 20 years. HIV- and HCV-related parameters, as well as markers relative to bacterial translocation, monocyte and lymphocyte activation and regulation were considered as independent variables. Dependent variables were the increase of CD4+ T cell count during the follow-up (12 months).

Results

Increased values of bacterial translocation, measured by lipopolysaccharide-binding protein, monocyte and lymphocyte activation markers and T regulatory lymphocytes were detected in HIV-monoinfected and HIV/HCV coinfected patients. Serum sCD14 and IL-6 were increased in HIV/HCV-coinfected patients with liver cirrhosis in comparison with those with chronic hepatitis or HIV-monoinfected individuals. Time with undetectable HIV load was not related with these parameters. The presence of cirrhosis was negatively associated with a CD4+ T cell count increase.

Conclusion

In patients with a chronic HIV infection, a persistent increase of lipopolysaccharide-binding protein and monocyte and lymphocyte modifications are present. HCV-related cirrhosis is associated with more elevated serum concentrations of monocyte-derived markers. Cirrhosis influences the continued immune reconstitution of these patients.     

PEG-IFN Alpha but Not Ribavirin Alters NK Cell Phenotype and Function in Patients with Chronic Hepatitis C

Background

Ribavirin (RBV) remains part of several interferon-free treatment strategies even though its mechanisms of action are still not fully understood. One hypothesis is that RBV increases responsiveness to type I interferons. Pegylated Interferon alpha (PEG-IFNa) has recently been shown to alter natural killer (NK) cell function possibly contributing to control of hepatitis C virus (HCV) infection. However, the effects of ribavirin alone or in combination with IFNa on NK cells are unknown.

Methods

Extensive ex vivo phenotyping and functional analysis of NK cells from hepatitis C patients was performed during antiviral therapy. Patients were treated for 6 weeks with RBV monotherapy (n = 11), placebo (n = 13) or PEG-IFNa-2a alone (n = 6) followed by PEG-IFNa/RBV combination therapy. The effects of RBV and PEG-IFNa-2a on NK cells were also studied in vitro after co-culture with K562 or Huh7.5 cells.

Results

Ribavirin monotherapy had no obvious effects on NK cell phenotype or function, neither ex vivo in patients nor in vitro. In contrast, PEG-IFNa-2a therapy was associated with an increase of CD56^bright cells and distinct changes in expression profiles leading to an activated NK cell phenotype, increased functionality and decline of terminally differentiated NK cells. Ribavirin combination therapy reduced some of the IFN effects. An activated NK cell phenotype during therapy was inversely correlated with HCV viral load.

Conclusions

PEG-IFNa activates NK cells possibly contributing to virological responses independently of RBV. The role of NK cells during future IFN-free combination therapies including RBV remains to be determined.     

Increased Baseline Proinflammatory Cytokine Production in Chronic Hepatitis C Patients with Rapid Virological Response to Peginterferon Plus Ribavirin

Background

Chronic hepatitis C (CHC) patients achieving rapid virological response (RVR) on PEG-IFN/ribavirin (P/R) therapy have high chance of sustained virological response (SVR). To analyze host immunological factors associated with RVR, viral kinetics, phenotype distribution and Th1/Th2 cytokine production by peripheral blood mononuclear cells (PBMC) were studied prior to and during P/R therapy.

Methods

TNF-α, IFN-γ, IL-2, IL-6, IL-4 and IL-10 production by PBMC were measured after Toll-like receptor 4 (TLR-4) or phorbol myristate acetate/Ionomycin stimulation in 20 healthy controls and in 50 CHC patients before receiving and during P/R therapy. RVR was achieved by 14, complete early virological response (cEVR) by 19 patients and 17 patients were null-responders (NR).

Results

Patients with RVR showed an increased baseline TNF-α and IL-6 production by TLR-4 activated monocytes and increased IFN-γ, decreased IL-4 and IL-10 production by lymphocytes compared to non-RVR patients. SVR was also associated with increased baseline TNF-α production and decreased IL-10 levels compared to patients who did not achieve SVR. Baseline IL-2 production was higher in cEVR compared to NR patients. Antiviral treatment increased TNF-α, IL-6 production by monocytes and IFN-γ secretion by lymphocytes and decreased IL-4 and IL-10 production by lymphocytes in cEVR compared to NR patients.

Conclusion

RVR was associated with increased baseline proinflammatory cytokine production by TLR-4 stimulated monocytes and by activated lymphocytes. In null-responders and in patients who did not achieve SVR both TLR-4 sensing function and proinflammatory cytokine production were impaired, suggesting that modulation of TLR activity and controlled induction of inflammatory cytokine production may provide further therapeutic strategy for CHC patients non-responding to P/R treatment.