Gastroenterology

Gastroenterology

Volume 117, Issue 6, December 1999, Pages 1386-1396
Gastroenterology

Liver, Pancreas, and Biliary Tract
Direct ex vivo analysis of hepatitis B virus-specific CD8+ T cells associated with the control of infection,☆☆,

https://doi.org/10.1016/S0016-5085(99)70289-1Get rights and content

Abstract

Background & Aims: Cytotoxic T cells have been suggested to be responsible for lysis of hepatitis B virus (HBV)-infected hepatocytes and control of virus infection. The frequency, kinetics, phenotype, and capacity for clonal expansion of circulating HBV-specific CD8 cells were analyzed directly in patients with acute HBV infection to clarify their pathogenetic role. Methods: Three HLA-A2 peptide tetramers able to visualize HBV core, envelope, and polymerase epitope–specific cytotoxic T lymphocytes were synthesized and used for flow cytometric analysis of antigen-specific populations. Results: Tetramer-positive cells specific for the core 18–27 epitope were found at a higher frequency than those specific for polymerase 575–583 and envelope 335–343 epitopes in most patients with acute HBV. The number of HBV-specific CD8 cells was highest during the clinically acute stage of infection and decreased after recovery. These cells expressed an activated phenotype and had an impaired capacity to expand in vitro and to display cytolytic activity in response to peptide stimulation. Recovery of these functions was observed when the frequency of specific CD8 cells decreased, coincident with a progressive decrease in their expression of activation markers. Conclusions: This study provides the first ex vivo evidence that the highest frequency of circulating HBV-specific CD8 cells coincides with the clinically acute phase of hepatitis B. These cells exhibit an activated phenotype with limited further proliferative capacity that is restored during recovery.

GASTROENTEROLOGY 1999;117:1386-1396

Section snippets

Patients and controls

Twenty-three adult subjects (18 male and 5 female; age range, 24–54 years) with self-limited acute HBV infection were studied. Diagnosis of acute hepatitis was based on increased serum alanine aminotransferase (ALT) activity (at least 10 times the upper limit of normal) and detection of hepatitis B surface (HBsAg) antigen and immunoglobulin (Ig) M anti–hepatitis B core antigen (anti-HBc) antibodies in the serum along with the recent onset of jaundice and associated symptoms. All patients were

Synthesis and characterization of HLA-A2/HBV tetramers

Tetramers of HLA-A2 complexed with peptides from core (amino acids 18–27), polymerase (amino acids 575–583), and envelope (amino acids 335–343) proteins were generated as described previously8 (hereafter indicated as T c18–27, T p575–583, and T e335–343, respectively) to study the HBV-specific CTL response to these dominant epitopes4, 5, 6 during acute HBV infection. The specificity of tetramers was determined using CTL clones2 and CTL lines generated by stimulation of PBMCs from

Discussion

The direct analysis of 14 HLA-A2–positive patients with acute HBV infection using the tetramer technology described in this study provides a quantitative characterization of the kinetics and hierarchy of the HBV-specific CD8 response during a self-limited infection with a noncytopathic virus. The same technology has been applied to the study of other acute viral infections, revealing the potential extent of virus-specific T-cell expansion. Fifty to 70% of CD8+ T cells are virus specific in the

Acknowledgements

The authors thank Ludovica Bruno for comments on the manuscript and help with fluorescence-activated cell sorting; and Peter Beverley, Persephone Borrow, and Jethro Herberg for critical reading of the manuscript.

References (34)

  • R Nayersina et al.

    HLA-A2 restricted cytotoxic T lymphocyte responses to multiple hepatitis B surface antigen epitopes during hepatitis B virus infection

    J Immunol

    (1993)
  • B Rehermann et al.

    The cytotoxic T lymphocyte response to multiple hepatitis B virus polymerase epitopes during and after acute viral hepatitis

    J Exp Med

    (1995)
  • MA Alexander-Miller et al.

    Role of antigen, CD8, and CTL avidity in high dose antigen induction of apoptosis of effector CTL

    J Exp Med

    (1996)
  • JD Altman et al.

    Phenotypic analysis of antigen-specific T lymphocytes

    Science

    (1996)
  • MF Callan et al.

    Direct visualization of antigen-specific CD8(+) T cells during the primary immune response to Epstein–Barr virus in vivo

    J Exp Med

    (1998)
  • A Gallimore et al.

    Induction and exhaustion of lymphocytic choriomeningitis virus–specific cytotoxic T lymphocytes visualized using soluble tetrameric major histocompatibility complex class I–peptide complexes

    J Exp Med

    (1998)
  • GS Ogg et al.

    Quantitation of HIV-1–specific cytotoxic T lymphocytes and plasma load of viral RNA

    Science

    (1998)
  • Cited by (319)

    View all citing articles on Scopus

    Address requests for reprints to: Antonio Bertoletti, M.D., Institute of Hepatology, University College London, 69-75 Chenies Mews, London, WC1E 6HX England. e-mail: [email protected]; fax: (44) 171-380-0405.

    ☆☆

    Supported by a Collaborative Research Grant from the Edward Jenner Institute for Vaccine Research (to M.K.M.); by the Fondo de Investigaciones Sanitarias (B.E.F.I. 98/9155) from the Ministerio de Sanidad y Consumo of Spain (J.R.L.); and partially supported by a University College London Clinical Research and Development Committee grant.

    M. K. Maini and C. Boni contributed equally to this work.

    View full text