Th1 and Th2 cytokine profiles induced by hepatitis C virus F protein in peripheral blood mononuclear cells from chronic hepatitis C patients

doi:10.1016/j.imlet.2013.05.002

Immunology Letters

Volume 152, Issue 2, May 2013, Pages 89-95

https://doi.org/10.1016/j.imlet.2013.05.002 Get rights and content

Highlights

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Th1/Th2 cytokine profiles induced by F protein in HCV patients were studied.
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Levels of IFN-γ, IL-2, IL-4, IL-5 in PBMC post F protein were compared.
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Different Th1/Th2 cytokine profiles exhibited in patients with and without anti-F.
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F protein may contribute to the Th1/Th2 response imbalance in CHC patients.
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F protein may involve in the progress of HCC pathogenesis.

Abstract

Th1 and Th2 cytokine response has been confirmed to be correlated with the pathogenesis of HCV infection. The aim of the study is to investigate the Th1 and Th2 cytokine profiles induced by HCV alternate reading frame protein (F protein) in chronic hepatitis C patients. We assessed the immune responses specific to HCV F protein in 55 chronic HCV patients. IFN-γ, IL-2, IL-4 and IL-5 secretion by peripheral blood mononuclear cells (PBMC) post F protein stimulation were compared among HCV patients and healthy donors. Finally, the associations between HCV F protein and HLA class II alleles were explored. We found that the seroprevalence of anti-F antibodies in HCV-related hepatocellular carcinoma (HCC) patients was significantly higher than that of patients without HCC, but such a significant difference in humoral immune responses to F protein was not observed in HCV 1b-infected- and non-HCV 1b-infected-patients. Additionally, the PBMC proliferation of HCC patients was significantly lower than that of patients without HCC. Furthermore, F protein stimulation of PBMCs from F-seropositive patients resulted in Th2 biased cytokine responses (significantly decreased IFN-γ and/or IL-2 and significantly increased IL-4 and/or IL-5 levels) that reportedly may contribute to HCC progression and pathogenesis. However, no significant difference in the association between HCV F protein and HLA-DRB1*0201, 0301, 0405, 1001 and HLA-DQB1*0201, 0401, 0502, 0602 was observed in this study. These findings suggest that F protein may contribute to the HCV-associated bias in Th1/Th2 responses of chronic hepatitis C patients including the progress of HCC pathogenesis.

Introduction

Hepatitis C virus (HCV) is responsible for more than 170 million individuals infected with the virus worldwide. It is one of the major causes of chronic liver diseases, with the risk of severe liver damage, such as cirrhosis and hepatocellular carcinoma (HCC) [1]. An important clinical feature of hepatitis C is the high rate of chronic hepatitis, with more than 50% HCV-infected patients developing chronic infection [2], among whom some will evolve to cirrhosis and HCC [1]. Unfortunately, no effective vaccine is available. The mechanisms underlying hepatocarcinogenesis in chronic HCV infection are not well understood, but it has been accepted that the viral genetics and host immune status are the major factors determining the outcome of HCV infection [3]. By means of signal interference, effector modulation and continual viral genetic variation, HCV evades host immune surveillance to establish and maintain persistent infection in vivo [4], [5].

The HCV genome (∼9.6 kb) includes a long open reading frame, encoding a polyprotein of approximately 3000 amino acids, which is cleaved by viral and cellular proteases to generate at least ten gene products: four structural proteins (core, E1, E2, p7) and six nonstructural proteins (NS2, NS3, NS4A, NS4B, NS5A, and NS5B) [6]. A novel HCV protein, termed alternate reading frame protein (F protein/ARFP/core +1 protein), has been previously reported [7], [8], [9]. F protein is encoded from an alternative +1 reading frame overlapping the core genomic region, starting at codon 11 and ending at codon 161 in HCV 1a genotype while shifting from codon 42 and ending at codon 141 in HCV 1b genotype [7], [8], [9]. However, the functions of F protein are still not fully understood. Evidence shows that F protein is produced during HCV infection [7], [8], [9], [10] and is characterized by a high degree of sequence conservation within the same genotypes [11] and immunogenicity to induce specific humoral and cellular immune response in vivo [7], [8], [9], [10], [12]. By using enzyme-linked immunosorbent assay (ELISA), the F protein was recognized by specific antibodies in the sera of HCV-infected patients [7], [9]. Furthermore, data also demonstrate that the proliferation and cytokine secretion of T helper (Th) cells can be induced specifically by the F protein in HCV patients [10]. Studies reveal that the F protein plays a role in the interaction of HCV with the host immune system. However, the Th1 and Th2 cytokine profiles specific to the F protein and the significance of it in the progress of HCV infection are not well understood. As we all know, human leukocyte antigen (HLA) class II are associated with HCV infection [13]. Additionally, eight HLA class II alleles were reported to influence on the serum ALT level, which related with the risk of liver cirrhosis or HCC in Chinese HCV patients, including HLA-DRB1*0201, 0301, 0405, 1001 and HLA-DQB1*0201, 0401, 0502, 0602 [14]. However, no one has yet assessed whether these HLA class II alleles can influence the immune response to F protein.

To explore the mechanisms behind F protein pathogenesis, we investigated Th1 and Th2 cytokine (including IFN-γ, IL-2, IL-4 and IL-5) secretion by F protein stimulated PBMCs from chronic hepatitis C (CHC) patients with and without HCC. Moreover, we analyzed the genotypes of HLA-DRB1*0201, 0301, 0405, 1001 and DQB1*0201, 0401, 0502, 0602 alleles to estimate the possible associations between the distribution of these HLA class II alleles and host humoral immune response to F protein.

Section snippets

Subjects

Fifty-five Chinese Han patients (32 men and 23 women, mean age 51.3 ± 10.2 years) were investigated here. The patients were subdivided as follows: (1) CHC group consisting of 31 CHC patients without HCC confirmed to be positive for anti-HCV in sera by second-generation ELISA kits (Abbott Laboratories, North Chicago, IL, USA) and HCV RNA tests (Amplicor, Roche Diagnostics, Manheim, Germany), with persistently elevated ALT levels for more than 2 years. Liver biopsy and ultrasound showed no

Detection of anti-F and anti-core antibodies in sera of HCV patients

The anti-F and anti-core antibodies in the sera of chronic HCV-infected patients with HCC (HCC patients) and without HCC (CHC patients) were examined using ELISA (Fig. 1). Anti-F antibodies were found in 18 (75%) of 24 HCC patients and 14 (45.2%) of 31 CHC patients. The prevalence of anti-F antibodies in HCC patients was significantly higher than that of the CHC patients (p = 0.025). However, no difference of anti-F antibodies prevalence was detected in HCV genotype 1b-infected patients compared

Discussion

In earlier studies, the existence of T- and B-cell mediated immune response specific to F protein during HCV infection has been demonstrated [7], [8], [9], [10], [20], [23]. A study [10] reported that IFN-γ, TNF-α, IL-2, IL-4, IL-5 and IL-10 were produced by T cell in response to F protein in HCV-infected or recovered patients. Additionally, we had also found a positive correlation between the sera IL-6 levels and anti-F antibody prevalence in some HCV patients [20]. These studies indicated

Conclusions

Evidence show that HCV F protein makes a difference in Th1/Th2 cytokine secretions, but the Th1/Th2 cytokine profiles induced by F protein are unlike those induced by the core protein in PBMC derived from CHC patients with or without HCC in vitro. It suggests that F protein may contribute to the HCV-associated bias in Th1/Th2 responses of chronic hepatitis C patients including the progress of HCC pathogenesis.

To our knowledge, this is the first report on Th1/Th2 cytokine profiles induced by the

Acknowledgements

This study was supported by grants from the National Natural Science Foundation of China (Nos. 81172724 and 30972628), National Major Scientific and Technological Projects (No. 2011ZX10004-902), Natural Science Foundation of Jiangsu Province of China (No. BK2011840) and from China Postdoctoral Science Foundation (No. 20090451569).

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Th1 and Th2 cytokine profiles induced by hepatitis C virus F protein in peripheral blood mononuclear cells from chronic hepatitis C patients

Highlights

Abstract

Introduction

Section snippets

Subjects

Detection of anti-F and anti-core antibodies in sera of HCV patients

Discussion

Conclusions

Acknowledgements

Perspect Med Virol

Clin Immunol

Clin Immunol

J Biol Chem

Hepatology

Lancet

Hum Immunol

J Hepatol

J Immunol Methods

Hepatology

J Biol Chem

Gene

J Clin Virol

Biochem Biophys Res Commun

J Hepatol

J Hepatol

Course and outcome of hepatitis C

Hepatology

Unscrambling hepatitis C virus–host interactions

Nature

Replication of hepatitis C virus

Nat Rev Microbiol

Evidence for a new hepatitis C virus antigen encoded in an overlapping reading frame

RNA (New York, NY)

Synthesis of a novel hepatitis C virus protein by ribosomal frameshift

EMBO J