Elsevier

Journal of Hepatology

Volume 52, Issue 3, March 2010, Pages 322-329
Journal of Hepatology

Research Article
T cell immunoglobulin- and mucin-domain-containing molecule-3 (Tim-3) mediates natural killer cell suppression in chronic hepatitis B

https://doi.org/10.1016/j.jhep.2009.12.005Get rights and content

Background & Aims

T cell immunoglobulin- and mucin-domain-containing molecule-3 (Tim-3) has been shown to influence autoimmune diseases; however, its function in viral infection has not been well-defined. We therefore investigated the expression and regulatory function of Tim-3 in natural killer (NK) cells in chronic Hepatitis B (CHB) infection.

Methods

Seventy-six CHB patients, 38 healthy controls, and 18 patients with fatty liver disease (FLD) were tested for Tim-3 expression on peripheral blood mononuclear cells (PBMCs) and in the liver tissue by flow cytometry and immunohistochemical stainning. The effects of HBV infection on Tim-3 expression in NK cells and the roles of Tim-3 in regulation of NK-cell function were also studied.

Results

There was a significant increase of Tim-3 expression in PBMCs, circulating NK cells and liver infiltrating lymphocytes (LILs) from CHB patients compared to that of healthy controls and FLD patients. Increased Tim-3 expression was also detected in NK92 cells that had been transfected with a HBV expression vector and NK cells isolated from the liver of HBV transgenic mice. Importantly, blockage of Tim-3 signaling with anti-Tim-3 antibodies or Tim-3-Fc fusion proteins resulted in an increased cytotoxicity for NK92 cells compared to HepG2 and HepG2.2.15 cells, as well as an elevated interferon-gamma (IFN-γ) production. Similarly, enhanced cytotoxicity was also observed in PBMCs or NK cells from CHB patients treated with the Tim-3 blockade ex vivo.

Conclusion

HBV infection can up-regulate Tim-3 expression in NK cells, which may in turn suppress NK-cell functions in CHB patients.

Introduction

More than 350 million people worldwide suffer from chronic Hepatitis B virus (HBV) infection, and approximately 1 million of them die annually from HBV-induced liver diseases [1], [2]. The persistence of virus that resulted from the low antiviral immune response had been thought to contribute to the pathogenesis of HBV-induced diseases [1], [3]. However, the precise mechanisms underlying the HBV-mediated immune suppression in chronic infection are not completely understood.

Previous studies have shown that activated NK cells display their antiviral effects by mediating elevated cytotoxicity during HBV infection as well as augmenting adaptive immune responses as a result of cytokine production [4]. Accumulated data support a role for NK cells in the defense against HBV infection [5], [6], [7]. Furthermore, it has been shown that natural killer activity differs in different stages of HBV infection [8], [9], [10]. Patients with acute HBV infection showed enhanced NK cytotoxicity early in the course of infection. On the contrary, natural killing activity was significantly inhibited in chronic HBV infection [9], [10].

T cell immunoglobulin- and mucin-domain-containing molecule-3 (Tim-3) plays a pivotal role in immune regulation and tolerance induction [11], [12], [13], [14], [15], [16], [17], [18], [19], [20], [21], [22], [23]. Engagement of Tim-3 by galectin-9 negatively regulates IFN-γ secretion and influences the ability to induce T cell tolerance in both mice and humans [14], [16]. Recent studies have shown that Tim-3 is differentially expressed in innate versus adaptive immune cells, and that the ligation of Tim-3 can induce distinct signaling events, which may influence a range of inflammatory conditions [15], [16], [17], [18], [19], [20], [21], [22], [23], [24], [25], [26], [27], [28], [29], [30].

Recently, Tim-3 mRNA has been shown to be mainly expressed in NK and NKT cells in humans [15]. However, it remains unclear what role Tim-3 plays in NK cells during the course of CHB infection. In this study, we thus examined the expression of Tim-3 in NK cells and assessed the role of Tim-3 in chronic HBV infection.

Section snippets

Patients and controls

A total of 132 subjects, including 76 patients with CHB, 18 patients with fatty liver disease (FLD) and 38 healthy individuals, were enrolled in this study. The liver tissues from 16 CHB patients and 6 FLD patients were collected for immunohistochemical staining. The diagnosis of these CHB patients were made according to the criteria established in the National Viral Hepatitis Conference of China (2000). FLD was diagnosed on the basis of abnormal liver enzymes, a detailed history, and exclusion

Augmented expression of Tim-3 in PBMCs and hepatic infiltrating lymphocytes from chronic hepatitis B patients

To study the expression of Tim-3 in PBMCs, we performed real-time RT-PCR analysis in 36 CHB patients, 20 healthy individuals and 9 FLD patients. There was a significant increase in Tim-3 expression on PBMCs in CHB patients, compared to that of healthy controls and FLD patients (CHB patients vs healthy controls vs FLD patients, mean ± SD 1.40 ± 0.61 vs 0.89 ± 0.40 vs 0.86 ± 0.47, p <0.01) (Fig. 1A). Further flow cytometric studies in 40 CHB patients, 18 healthy individuals and 9 FLD patients revealed

Discussion

Tim-3 was originally identified as a member of the Tim family, especially expressed in Th1 cells [11], [15]. Later, it has also been found in many other cells [18], [19], [20]. The engagement of Tim-3 with its ligand has been linked to a wide variety of physiologic and pathologic functions [11], [12], [13], [14], [15], [16], [17], [18], [19], [20], [21], [22], [23], [24], [25], [26], [27], [28], [29], [30]. Much effort has been devoted to understanding its roles in regulating inflammatory

Acknowledgements

The authors who have taken part in this study declared that they do not have anything to declare regarding funding from industry or conflict of interest with respect to this manuscript. We thank Dr. Jian Chen, Prof. Youhai Chen and Prof. Jianfeng Li for discussion and help with the revision of the manuscript, Prof. Shuangyin Han for providing blood samples.

This work was supported in part by Grants from the National Science Foundation of China (No. 30670966 and No. 30972753), the National Basic

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    These authors contributed equally to this work.

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