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Hepatocyte expression of hepatitis B surface and core antigens across phases of chronic hepatitis B infection

      Linked Article

      To the Editor:
      In a recent issue of the Journal, Montanari et al.
      • Montanari N.R.
      • Ramírez R.
      • Aggarwal A.
      • van Buuren N.
      • Doukas M.
      • Moon C.
      • et al.
      Multi-parametric analysis of human livers reveals variation in intrahepatic inflammation across phases of chronic hepatitis B infection.
      comprehensively analyzed intrahepatic inflammation across phases of chronic hepatitis B infection. We have 2 comments on this study:
      Classification of immune tolerance (IT) phase: In this study, alanine aminotransferase (ALT) of 30 IU/ml was considered normal and ALT of 45 IU/ml (1.5 x upper limit of normal [ULN]) was used as the cut-off threshold to differentiate patients with inactive infection from those with chronic hepatitis. However, the HBeAg-positive chronic infection or IT phase is better diagnosed by persistent ALT within the normal range. Even with a cut-off threshold of 1.5x ULN for ALT, 4 of the 15 IT patients in this study did have ALT levels >50 IU/ml. In addition, 1 IT patient had advanced fibrosis (F3 or F4 of Metavir fibrosis score). These patients have likely been misclassified as IT patients.
      Intrahepatic viral antigen expression: Using formalin-fixed paraffin-embedded sections of liver, the present study showed that the percentage of HBsAg-positive hepatocytes ranged from 0 to 100% (0–100% on the membrane and 0–80% in the cytoplasm) during the IT or immune active (IA) phase, whereas during the immune control (IC) phase (HBeAg-negative chronic HBV infection), most patients had low levels of HBsAg expression (<25% of hepatocytes), with membranous expression exceeding cytoplasmic expression. These results contradict previous observations. Studies using frozen sections of liver revealed that HBsAg was diffusely distributed on the plasma membrane as well as focally (usually <25% of hepatocytes) in the cytoplasm during the IT or IA phase, whereas HBsAg was extensively (usually >50% of hepatocytes) distributed in the cytoplasm only during the IC phase.
      • Chu C.M.
      • Liaw Y.F.
      Intrahepatic distribution of hepatitis B surface and core antigens in chronic hepatitis B virus infection. Hepatocyte with cytoplasmic/membranous hepatitis B core antigen as a possible target for immune hepatocytolysis.
      Membranous HBsAg expression correlates with HBcAg expression in the liver
      • Ray M.B.
      • Desmet V.J.
      • Bradburne A.F.
      • Desmyter J.
      • Fevery J.
      • De Groote J.
      Differential distribution of hepatitis B surface antigen and hepatitis B core antigen in the liver of hepatitis B patients.
      and also with high levels of HBV DNA in serum.
      • Chu C.M.
      • Liaw Y.F.
      Membrane staining for hepatitis B surface antigen on hepatocytes: a sensitive and specific marker of active viral replication in hepatitis B.
      Extensive cytoplasmic HBsAg expression in the IC phase correlates with high numbers of classical ground glass hepatocytes seen in IC patients.
      • Ray M.B.
      • Desmet V.J.
      • Bradburne A.F.
      • Desmyter J.
      • Fevery J.
      • De Groote J.
      Differential distribution of hepatitis B surface antigen and hepatitis B core antigen in the liver of hepatitis B patients.
      Of note, immunohistochemical staining on paraffin sections of liver is much less sensitive and specific for detecting HBsAg on hepatocyte membrane than on frozen sections of liver,
      • Gowans E.J.
      • Burrell C.J.
      Widespread presence of cytoplasmic HBcAg in hepatitis B infected liver detected by improved immunochemical methods.
      which may partly explain the discrepant results of membranous HBsAg expression. However, the reason for low numbers of hepatocytes expressing cytoplasmic HBsAg in IC patients in this study remains unclear. Regarding hepatocyte HBcAg expression, the present study revealed that, during both IT and IA phases, HBcAg was mainly localized in the cytoplasm and rarely in the nucleus, and the degrees of liver inflammation did not correlate with intrahepatic HBcAg levels. However, it has long been recognized that HBcAg is distributed in the nucleus mainly during the IT phase, whereas during the IA phase, nuclear HBcAg expression decreases markedly alongside a concomitant increase in cytoplasmic HBcAg expression.
      • Chu C.M.
      • Liaw Y.F.
      Intrahepatic distribution of hepatitis B surface and core antigens in chronic hepatitis B virus infection. Hepatocyte with cytoplasmic/membranous hepatitis B core antigen as a possible target for immune hepatocytolysis.
      ,
      • Hsu H.C.
      • Su I.J.
      • Lai M.Y.
      • Chen D.S.
      • Chang M.H.
      • Chuang S.M.
      • et al.
      Biologic and prognostic significance of hepatocyte hepatitis B core antigen expressions in the natural course of chronic hepatitis B virus infection.
      ,
      • Mason W.S.
      • Gill U.S.
      • Litwin S.
      • Zhou Y.
      • Peri S.
      • Pop O.
      • et al.
      HBV DNA integration and clonal hepatocyte expansion in chronic hepatitis B patients considered immune tolerant.
      Patients with predominantly cytoplasmic HBcAg expression had significantly higher degrees of liver inflammation and liver cell regeneration, compared to patients with predominantly nuclear HBcAg expression.
      • Chu C.M.
      • Yeh C.T.
      • Sheen I.S.
      • Liaw Y.F.
      Subcellular localization of hepatitis B core antigen in relation to hepatocyte regeneration in chronic hepatitis B.
      It is not appropriate to correlate liver inflammation with intrahepatic HBcAg levels without considering topographic distribution. It also seems that some of the IT patients in this study are indeed previously unrecognized IA patients with remission of hepatitis at the time of enrollment.
      In conclusion, some of the IT patients in this study had high ALT, advanced fibrosis and predominantly cytoplasmic HBcAg expression, which was inconsistent with the diagnosis of IT. However, since the proportion of such patients is low, its impact on the results of the overall case analysis should be limited.

      Financial support

      The authors received no financial support to produce this manuscript.

      Authors’ contributions

      CM Chu: Conception and design of the letter; Analysis and interpretation of data; Drafting of the manuscript; Approval of the final version of the manuscript. YF Liaw: Design of the letter; Interpretation of data; Critical revision of manuscript; Approval of the final version of the manuscript.

      Conflict of interest

      The authors have no financial and personal relationships with other people or organizations that could inappropriately influence (bias) their work.
      Please refer to the accompanying ICMJE disclosure forms for further details.

      Supplementary data

      The following are the supplementary data to this article:

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        Multi-parametric analysis of human livers reveals variation in intrahepatic inflammation across phases of chronic hepatitis B infection.
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