Reply to: “Hepatocyte expression of hepatitis B surface and core antigens across phases of chronic hepatitis B infection”

Hepatocyte expression of hepatitis B surface and core antigens across phases of chronic hepatitis B infectionJournal of HepatologyVol. 77Issue 5PreviewIn a recent issue of the Journal, Montanari et al.1 comprehensively analyzed intrahepatic inflammation across phases of chronic hepatitis B infection. We have 2 comments on this study: Full-Text PDF Multi-parametric analysis of human livers reveals variation in intrahepatic inflammation across phases of chronic hepatitis B infectionJournal of HepatologyVol. 77Issue 2PreviewChronic HBV is clinically categorized into 4 phases by a combination of serum HBV DNA levels, HBeAg status and alanine aminotransferase (ALT): immunotolerant (IT), immune-active (IA), inactive carrier (IC) and HBeAg-negative hepatitis (ENEG). Immune and virological measurements in the blood have proven useful but are insufficient to explain the interrelation between the immune system and the virus since immune dynamics differ in the blood and liver. Furthermore, the inflammatory response in the liver and parenchymal cells cannot be fully captured in blood. Full-Text PDF Open Access We would like to thank Drs. Chu and Liaw[1]Chu C.-M. Liaw Y.-F. Hepatocyte expression of hepatitis B surface and core antigens across phases of chronic hepatitis B infection.J Hepatol. 2022; 77: 1455-1456Abstract Full Text Full Text PDF Scopus (1) Google Scholar for their interest in our work and we appreciate their comments on the findings described in our manuscript.[2]Montanari N.R. Ramirez 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.J Hepatol. 2022; 77: 332-343Abstract Full Text Full Text PDF Scopus (3) Google Scholar The classification of chronic HBV patients in clinical phases is based on the serum levels of HBV DNA, alanine aminotransferase (ALT) and HBeAg, and strict thresholds have been defined for their classification, which were originally meant to guide clinical decision making. For our transcriptomic and immunohistochemical study, we designated 4 patients with ALT levels ∼50 IU/ml as immunotolerant patients, since their ALT levels were slightly elevated but stable over time, along with minimal immune infiltrate as observed by histology. The intrahepatic viral antigen expression data generated using state of the art digital pathology techniques allowed us to profile the intrahepatic HBsAg and HBcAg burden across the different phases of chronic HBV infection. Seminal work performed by Drs. Chu and Liaw[3]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.Gastroenterology. 1987; 92: 220-225Abstract Full Text PDF PubMed Scopus (183) Google Scholar,[4]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.J Clin Pathol. 1995; 48: 470-473Crossref PubMed Scopus (36) Google Scholar has previously shown differential expression patterns of HBsAg and HBcAg in chronic HBV-infected livers using fresh frozen tissue. While there are some discrepancies in the data obtained between the studies, it is important to note that differences in technologies and primary antibody clone epitopes used makes comparison between the studies difficult. Fresh frozen tissue has traditionally been considered more sensitive than FFPE tissue; however, recent advances in technologies allowing amplification of signal, such as the InSituPlex technology used in our study, and the use of well characterized antibodies has greatly improved the sensitivity and specificity of FFPE assays.[5]Wharton K.A.J. Wood D. Manesse M. Maclean K.H. Leiss F. Zuraw A. Tissue multiplex analyte detection in anatomic pathology - pathways to clinical implementation.Front Mol Biosci. 2021; 8672531Crossref PubMed Scopus (9) Google Scholar,[6]Humphries M.P. Bingham V. Abdullahi Sidi F. Craig S.G. McQuaid S. James J. et al.Improving the diagnostic accuracy of the PD-L1 test with image analysis and multiplex hybridization.Cancers. 2020; 12: 1114Crossref Scopus (22) Google Scholar Finally, in addition to technological differences, intra- and inter-patient heterogeneity in viral burden makes it challenging to reliably interpret a small number of regions of interest from each biopsy.[7]Aggarwal A. Odirizzi P. Brodbeck J. van Buuren N. Moon C. Chang S. et al.Quantification of HBV hepatocyte burden using novel multiplex immunofluorescence staining and image analysis reveals substantial reduction in HBV liver burden with anti-viral treatment.J Hepatol. 2022; 77: S832Abstract Full Text PDF Google Scholar The ability to scan and analyze the entire biopsy using whole slide images increases our confidence by enabling consistent and unbiased analysis of our data. We appreciate the opportunity to clarify these important points. The work described in the original manuscript was funded by Foundation for Liver and Gastrointestinal Research (AB). All authors were involved and approved the final version of the manuscript. NRM, and AB declare no conflict of interest related to the content of this letter. At the time this study was conducted, AA, RR, LD, LL and BF were employees and stockholders of Gilead Sciences, Inc. Please refer to the accompanying ICMJE disclosure forms for further details. The following are the supplementary data to this article: Download .pdf (.38 MB) Help with pdf files Multimedia component 1