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Aasarey R, Yadav K, Kashyap BK, Prabha S, Kumar P, Kumar A, Ruokolainen J, Kesari KK. Role of Immunological Cells in Hepatocellular Carcinoma Disease and Associated Pathways. ACS Pharmacol Transl Sci 2023; 6:1801-1816. [PMID: 38093838 PMCID: PMC10714437 DOI: 10.1021/acsptsci.3c00216] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2023] [Revised: 10/12/2023] [Accepted: 10/13/2023] [Indexed: 03/28/2024]
Abstract
Hepatocellular carcinoma (HCC) remains one of the predominant causes of cancer-related mortality across the globe. It is attributed to obesity, excessive alcohol consumption, smoking, and infection by the hepatitis virus. Early diagnosis of HCC is essential, and local treatments such as surgical excision and percutaneous ablation are effective. Palliative systemic therapy, primarily with the tyrosine kinase inhibitor Sorafenib, is used in advanced cases. However, the prognosis for advanced HCC remains poor. This Review additionally describes the pathophysiological mechanisms of HCC, which include aberrant molecular signaling, genomic instability, persistent inflammation, and the paradoxical position of the immune system in promoting and suppressing HCC. The paper concludes by discussing the growing body of research on the relationship between mitochondria and HCC, suggesting that mitochondrial dysfunction may contribute to the progression of HCC. This Review focuses on immunological interactions between different mechanisms of HCC progression, including obesity, viral infection, and alcohol consumption.
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Affiliation(s)
- Ram Aasarey
- Department
of Laboratory Medicine, All India Institute
of Medical Science, New Delhi-11029, India
| | - Kajal Yadav
- Department
of Biotechnology, All India Institute of
Medical Science, New Delhi-11029, India
| | - Brijendra Kumar Kashyap
- Department
of Biotechnology Engineering, Institute of Engineering and Technology, Bundelkhand University, Jhansi-284128, Uttar Pradesh, India
| | - Sarit Prabha
- Department
of Biological Science and Engineering, Maulana
Azad National Institute of Technology, Bhopal-462003, Madhya Pradesh,India
| | - Pramod Kumar
- Indian
Council of Medical Research, National Institute
of Cancer Prevention and Research (NICPR), l-7, Sector-39, Noida-201301, National Capital Region, India
| | - Anil Kumar
- Department
of Life Sciences, School of Natural Sciences, Central University of Jharkhand, Cheri-Manatu, Karmre, Kanke-835222, Ranchi, India
| | - Janne Ruokolainen
- Department
of Applied Physics, School of Science, Aalto
University, FI-00076 Espoo, Finland
| | - Kavindra Kumar Kesari
- Department
of Applied Physics, School of Science, Aalto
University, FI-00076 Espoo, Finland
- Research
and Development Cell, Lovely Professional
University, Phagwara-144411, Punjab, India
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Annexins in Translational Research: Hidden Treasures to Be Found. Int J Mol Sci 2018; 19:ijms19061781. [PMID: 29914106 PMCID: PMC6032224 DOI: 10.3390/ijms19061781] [Citation(s) in RCA: 48] [Impact Index Per Article: 6.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2018] [Revised: 06/06/2018] [Accepted: 06/12/2018] [Indexed: 12/12/2022] Open
Abstract
The vertebrate annexin superfamily (AnxA) consists of 12 members of a calcium (Ca2+) and phospholipid binding protein family which share a high structural homology. In keeping with this hallmark feature, annexins have been implicated in the Ca2+-controlled regulation of a broad range of membrane events. In this review, we identify and discuss several themes of annexin actions that hold a potential therapeutic value, namely, the regulation of the immune response and the control of tissue homeostasis, and that repeatedly surface in the annexin activity profile. Our aim is to identify and discuss those annexin properties which might be exploited from a translational science and specifically, a clinical point of view.
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Kuehnl A, Musiol A, Raabe CA, Rescher U. Emerging functions as host cell factors - an encyclopedia of annexin-pathogen interactions. Biol Chem 2017; 397:949-59. [PMID: 27366904 DOI: 10.1515/hsz-2016-0183] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/19/2016] [Accepted: 06/28/2016] [Indexed: 12/14/2022]
Abstract
Emerging infectious diseases and drug-resistant infectious agents call for the development of innovative antimicrobial strategies. With pathogenicity now considered to arise from the complex and bi-directional interplay between a microbe and the host, host cell factor targeting has emerged as a promising approach that might overcome the limitations of classical antimicrobial drug development and could open up novel and efficient therapeutic strategies. Interaction with and modulation of host cell membranes is a recurrent theme in the host-microbe relationship. In this review, we provide an overview of what is currently known about the role of the Ca2+ dependent, membrane-binding annexin protein family in pathogen-host interactions, and discuss their emerging functions as host cell derived auxiliary proteins in microbe-host interactions and host cell targets.
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Petrareanu C, Macovei A, Sokolowska I, Woods AG, Lazar C, Radu GL, Darie CC, Branza-Nichita N. Comparative proteomics reveals novel components at the plasma membrane of differentiated HepaRG cells and different distribution in hepatocyte- and biliary-like cells. PLoS One 2013; 8:e71859. [PMID: 23977166 PMCID: PMC3748114 DOI: 10.1371/journal.pone.0071859] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/25/2013] [Accepted: 07/04/2013] [Indexed: 12/11/2022] Open
Abstract
Hepatitis B virus (HBV) is a human pathogen causing severe liver disease and eventually death. Despite important progress in deciphering HBV internalization, the early virus-cell interactions leading to infection are not known. HepaRG is a human bipotent liver cell line bearing the unique ability to differentiate towards a mixture of hepatocyte- and biliary-like cells. In addition to expressing metabolic functions normally found in liver, differentiated HepaRG cells support HBV infection in vitro, thus resembling cultured primary hepatocytes more than other hepatoma cells. Therefore, extensive characterization of the plasma membrane proteome from HepaRG cells would allow the identification of new cellular factors potentially involved in infection. Here we analyzed the plasma membranes of non-differentiated and differentiated HepaRG cells using nanoliquid chromatography-tandem mass spectrometry to identify the differences between the proteomes and the changes that lead to differentiation of these cells. We followed up on differentially-regulated proteins in hepatocytes- and biliary-like cells, focusing on Cathepsins D and K, Cyclophilin A, Annexin 1/A1, PDI and PDI A4/ERp72. Major differences between the two proteomes were found, including differentially regulated proteins, protein-protein interactions and intracellular localizations following differentiation. The results advance our current understanding of HepaRG differentiation and the unique properties of these cells.
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Affiliation(s)
- Catalina Petrareanu
- Department of Viral Glycoproteins, Institute of Biochemistry of the Romanian Academy, Bucharest, Romania
- Department of Analytical Chemistry and Enviromental Engineering, Faculty of Applied Chemistry and Materials Science, Politehnica University of Bucharest, Bucharest, Romania
| | - Alina Macovei
- Department of Viral Glycoproteins, Institute of Biochemistry of the Romanian Academy, Bucharest, Romania
| | - Izabela Sokolowska
- Biochemistry and Proteomics Group, Department of Chemistry and Biomolecular Science, Clarkson University, Potsdam, New York, United States of America
| | - Alisa G. Woods
- Biochemistry and Proteomics Group, Department of Chemistry and Biomolecular Science, Clarkson University, Potsdam, New York, United States of America
| | - Catalin Lazar
- Department of Viral Glycoproteins, Institute of Biochemistry of the Romanian Academy, Bucharest, Romania
| | - Gabriel L. Radu
- Department of Analytical Chemistry and Enviromental Engineering, Faculty of Applied Chemistry and Materials Science, Politehnica University of Bucharest, Bucharest, Romania
| | - Costel C. Darie
- Biochemistry and Proteomics Group, Department of Chemistry and Biomolecular Science, Clarkson University, Potsdam, New York, United States of America
| | - Norica Branza-Nichita
- Department of Viral Glycoproteins, Institute of Biochemistry of the Romanian Academy, Bucharest, Romania
- * E-mail:
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5
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Sun P, Zheng J, She G, Wei X, Zhang X, Shi H, Zhou X. Expression pattern of asialoglycoprotein receptor in human testis. Cell Tissue Res 2013; 352:761-8. [PMID: 23604802 DOI: 10.1007/s00441-013-1616-8] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/26/2012] [Accepted: 03/11/2013] [Indexed: 02/05/2023]
Abstract
During acute or chronic hepatitis B virus (HBV) infection, the virus can invade the male reproductive system, pass through the blood-testis barrier and integrate into the germ line, resulting in abnormal spermatozoa. However, the pathway remains unclear. The asialoglycoprotein receptor (ASGR), a potential receptor for HBV, is mainly distributed in hepatocytes. We have examined the distribution of ASGR in human testis and found it in the seminiferous tubules and interstitial region but its enrichment in human testis is much lower than that in liver. By multiple immunoenzyme histochemistry staining, ASGR was precisely co-localized with vimentin (Sertoli cell marker) but not proliferating cell nuclear antigen (spermatogonial cell marker) in testis tissue. ASGR was expressed in human Leydig cells, stromal cells in the seminiferous tubules and Sertoli cells but seldom in spermatogonial cells. Therefore, ASGR could provide HBV with access to the luminal compartment of human testis. The mechanism by which HBV invades germ cells remains unknown.
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Affiliation(s)
- Pingnan Sun
- Department of Pathology, Shantou University Medical College, Shantou, People's Republic of China
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6
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Entry of hepatitis B virus into immortalized human primary hepatocytes by clathrin-dependent endocytosis. J Virol 2012; 86:9443-53. [PMID: 22740403 DOI: 10.1128/jvi.00873-12] [Citation(s) in RCA: 123] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023] Open
Abstract
The lack of a suitable in vitro hepatitis B virus (HBV) infectivity model has limited examination of the early stages of the virus-cell interaction. In this study, we used an immortalized cell line derived from human primary hepatocytes, HuS-E/2, to study the mechanism of HBV infection. HBV infection efficiency was markedly increased after dimethyl sulfoxide (DMSO)-induced differentiation of the cells. Transmission electron microscopy demonstrated the presence of intact HBV particles in DMSO-treated HBV-infected HuS-E/2 cells, which could be infected with HBV for up to at least 50 passages. The pre-S1 domain of the large HBsAg (LHBsAg) protein specifically interacted with clathrin heavy chain (CHC) and clathrin adaptor protein AP-2. Short hairpin RNA knockdown of CHC or AP-2 in HuS-E/2 cells significantly reduced their susceptibility to HBV, indicating that both are necessary for HBV infection. Furthermore, HBV entry was inhibited by chlorpromazine, an inhibitor of clathrin-mediated endocytosis. LHBsAg also interfered with the clathrin-mediated endocytosis of transferrin by human hepatocytes. This infection system using an immortalized human primary hepatocyte cell line will facilitate investigations into HBV entry and in devising therapeutic strategies for manipulating HBV-associated liver disorders.
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7
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Hofmann A, Osman A, Leow CY, Driguez P, McManus DP, Jones MK. Parasite annexins--new molecules with potential for drug and vaccine development. Bioessays 2011; 32:967-76. [PMID: 21105292 DOI: 10.1002/bies.200900195] [Citation(s) in RCA: 40] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Abstract
In the last few years, annexins have been discovered in several nematodes and other parasites, and distinct differences between the parasite annexins and those of the hosts make them potentially attractive targets for anti-parasite therapeutics. Annexins are ubiquitous proteins found in almost all organisms across all kingdoms.Here, we present an overview of novel annexins from parasitic organisms, and summarize their phylogenetic and biochemical properties, with a view to using them as drug or vaccine targets. Building on structural and biological information that has been accumulated for mammalian and plant annexins, we describe a predicted additional secondary structure element found in many parasite annexins that may confer unique functional properties, and present a specific antigenic epitope for use as a vaccine.
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Affiliation(s)
- Andreas Hofmann
- Structural Chemistry Program, Eskitis Institute for Cell and Molecular Therapies, Griffith University, Nathan, Queensland, Australia.
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8
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Roelandt P, Pauwelyn KA, Sancho-Bru P, Subramanian K, Bose B, Ordovas L, Vanuytsel K, Geraerts M, Firpo M, De Vos R, Fevery J, Nevens F, Hu WS, Verfaillie CM. Human embryonic and rat adult stem cells with primitive endoderm-like phenotype can be fated to definitive endoderm, and finally hepatocyte-like cells. PLoS One 2010; 5:e12101. [PMID: 20711405 PMCID: PMC2920330 DOI: 10.1371/journal.pone.0012101] [Citation(s) in RCA: 63] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2010] [Accepted: 07/13/2010] [Indexed: 01/29/2023] Open
Abstract
Stem cell-derived hepatocytes may be an alternative cell source to treat liver diseases or to be used for pharmacological purposes. We developed a protocol that mimics mammalian liver development, to differentiate cells with pluripotent characteristics to hepatocyte-like cells. The protocol supports the stepwise differentiation of human embryonic stem cells (ESC) to cells with characteristics of primitive streak (PS)/mesendoderm (ME)/definitive endoderm (DE), hepatoblasts, and finally cells with phenotypic and functional characteristics of hepatocytes. Remarkably, the same protocol can also differentiate rat multipotent adult progenitor cells (rMAPCs) to hepatocyte-like cells, even though rMAPC are isolated clonally from cultured rat bone marrow (BM) and have characteristics of primitive endoderm cells. A fraction of rMAPCs can be fated to cells expressing genes consistent with a PS/ME/DE phenotype, preceding the acquisition of phenotypic and functional characteristics of hepatocytes. Although the hepatocyte-like progeny derived from both cell types is mixed, between 10-20% of cells are developmentally consistent with late fetal hepatocytes that have attained synthetic, storage and detoxifying functions near those of adult hepatocytes. This differentiation protocol will be useful for generating hepatocyte-like cells from rodent and human stem cells, and to gain insight into the early stages of liver development.
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Affiliation(s)
- Philip Roelandt
- Interdepartmental Stem Cell Institute Leuven, Catholic University Leuven, Belgium
- Hepatology Department, University Hospitals Leuven, Belgium
| | - Karen Ann Pauwelyn
- Interdepartmental Stem Cell Institute Leuven, Catholic University Leuven, Belgium
- Hepatology Department, University Hospitals Leuven, Belgium
| | - Pau Sancho-Bru
- Interdepartmental Stem Cell Institute Leuven, Catholic University Leuven, Belgium
| | - Kartik Subramanian
- Stem Cell Institute Minnesota, University of Minnesota, Minneapolis, Minnesota, United States of America
- Department of Chemical Engineering and Materials Science, University of Minnesota, Minneapolis, Minnesota, United States of America
| | - Bipasha Bose
- Interdepartmental Stem Cell Institute Leuven, Catholic University Leuven, Belgium
| | - Laura Ordovas
- Interdepartmental Stem Cell Institute Leuven, Catholic University Leuven, Belgium
| | - Kim Vanuytsel
- Interdepartmental Stem Cell Institute Leuven, Catholic University Leuven, Belgium
| | - Martine Geraerts
- Interdepartmental Stem Cell Institute Leuven, Catholic University Leuven, Belgium
| | - Meri Firpo
- Stem Cell Institute Minnesota, University of Minnesota, Minneapolis, Minnesota, United States of America
| | - Rita De Vos
- Pathology Department, University Hospitals Leuven, Leuven, Belgium
| | - Johan Fevery
- Hepatology Department, University Hospitals Leuven, Belgium
| | | | - Wei-Shou Hu
- Stem Cell Institute Minnesota, University of Minnesota, Minneapolis, Minnesota, United States of America
- Department of Chemical Engineering and Materials Science, University of Minnesota, Minneapolis, Minnesota, United States of America
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9
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Xie Y, Zhai J, Deng Q, Tiollais P, Wang Y, Zhao M. Entry of hepatitis B virus: mechanism and new therapeutic target. ACTA ACUST UNITED AC 2010; 58:301-7. [PMID: 20570056 DOI: 10.1016/j.patbio.2010.04.001] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2010] [Accepted: 04/12/2010] [Indexed: 12/21/2022]
Abstract
Entry of hepatitis B virus (HBV) into human hepatocytes constitutes the initial step in viral infection. The study of HBV entry had long been hampered by the lack of efficient cell culture systems and small animal models. The situation was greatly improved in the last decade with the development of HBV-infectible HepaRG cell line and primary Tupaia hepatocyte culture. Armed with these new tools, marked progresses have been achieved in the elucidation of the mechanism of HBV entry. Plenty of evidences indicate that the viral large surface protein (LHBs) is essential for HBV entry. Several regions in the PreS1 domain of LHBs have been verified to contribute directly to the viral attachment. In addition, a myristate moiety linked to the N-terminal glycine of PreS1 appears critical for HBV infectivity. Recently, the cysteine-rich antigenic loop of the S domain was identified as another crucial determinant for HBV infectivity. On the other hand, several cellular proteins were implicated in HBV attachment to hepatic cells, though definitive proofs are required in support to their functional involvement in HBV infection. Aiming to blocking viral entry, a couple of approaches based on acylated PreS1-derived peptides and short PreS1-binding peptides are currently under investigation, which have the potential to become novel antiviral therapeutics.
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Affiliation(s)
- Y Xie
- Key Laboratory of Medical Molecular Virology, Shanghai Medical College, Fudan University, Shanghai, China.
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10
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Fibronectin and asialoglyprotein receptor mediate hepatitis B surface antigen binding to the cell surface. Arch Virol 2010; 155:881-8. [PMID: 20364278 DOI: 10.1007/s00705-010-0657-5] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2009] [Accepted: 03/03/2010] [Indexed: 12/15/2022]
Abstract
Both fibronectin and the asialoglycoprotein receptor (ASGPR) have been identified by some investigators as partners for hepatitis B virus (HBV) envelope proteins. Because fibronectin is a natural ligand for ASGPR, we speculated that HBV might attach to ASGPR expressed on the hepatocyte surface via fibronectin. To test this hypothesis, we first confirmed by co-immunoprecipitation that ASGPR, fibronectin and HBsAg bind to each other in HepG2.2.15 cells, and possible binding domains were identified by GST pull-down. In addition, by measuring binding of HBsAg to cells, we found that ASGPR and fibronectin enhanced the binding capability of HBsAg to HepG2 cells, and even to 293T and CHO cells, which normally do not bind HBV. In conclusion, our findings suggest that both fibronectin and ASGPR mediate HBsAg binding to the cell surface, which provides further evidence for the potential roles of these two proteins in mediating HBV binding to liver cells.
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Guidotti LG, Chisari FV. Immunobiology and pathogenesis of viral hepatitis. ANNUAL REVIEW OF PATHOLOGY-MECHANISMS OF DISEASE 2007; 1:23-61. [PMID: 18039107 DOI: 10.1146/annurev.pathol.1.110304.100230] [Citation(s) in RCA: 589] [Impact Index Per Article: 32.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Among the many viruses that are known to infect the human liver, hepatitis B virus (HBV) and hepatitis C virus (HCV) are unique because of their prodigious capacity to cause persistent infection, cirrhosis, and liver cancer. HBV and HCV are noncytopathic viruses and, thus, immunologically mediated events play an important role in the pathogenesis and outcome of these infections. The adaptive immune response mediates virtually all of the liver disease associated with viral hepatitis. However, it is becoming increasingly clear that antigen-nonspecific inflammatory cells exacerbate cytotoxic T lymphocyte (CTL)-induced immunopathology and that platelets enhance the accumulation of CTLs in the liver. Chronic hepatitis is characterized by an inefficient T cell response unable to completely clear HBV or HCV from the liver, which consequently sustains continuous cycles of low-level cell destruction. Over long periods of time, recurrent immune-mediated liver damage contributes to the development of cirrhosis and hepatocellular carcinoma.
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Affiliation(s)
- Luca G Guidotti
- Department of Molecular and Experimental Medicine, Scripps Research Institute, La Jolla, California 92037, USA.
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12
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Abstract
Hepadnaviridae is a family of hepatotropic DNA viruses that is divided into the genera orthohepadnavirus of mammals and avihepadnavirus of birds. All members of this family can cause acute and chronic hepatic infection, which in the case of human hepatitis B virus (HBV) constitutes a major global health problem. Although our knowledge about the molecular biology of these highly liver-specific viruses has profoundly increased in the last two decades, the mechanisms of attachment and productive entrance into the differentiated host hepatocytes are still enigmatic. The difficulties in studying hepadnaviral entry were primarily caused by the lack of easily accessible in vitro infection systems. Thus, for more than twenty years, differentiated primary hepatocytes from the respective species were the only in vitro models for both orthohepadnaviruses (e.g. HBV) and avihepadnaviruses (e.g. duck hepatitis B virus [DHBV]). Two important discoveries have been made recently regarding HBV: (1) primary hepatocytes from tree-shrews; i.e., Tupaia belangeri, can be substituted for primary human hepatocytes, and (2) a human hepatoma cell line (HepaRG) was established that gains susceptibility for HBV infection upon induction of differentiation in vitro. A number of potential HBV receptor candidates have been described in the past, but none of them have been confirmed to function as a receptor. For DHBV and probably all other avian hepadnaviruses, carboxypeptidase D (CPD) has been shown to be indispensable for infection, although the exact role of this molecule is still under debate. While still restricted to the use of primary duck hepatocytes (PDH), investigations performed with DHBV provided important general concepts on the first steps of hepadnaviral infection. However, with emerging data obtained from the new HBV infection systems, the hope that DHBV utilizes the same mechanism as HBV only partially held true. Nevertheless, both HBV and DHBV in vitro infection systems will help to: (1) functionally dissect the hepadnaviral entry pathways, (2) perform reverse genetics (e.g. test the fitness of escape mutants), (3) titrate and map neutralizing antibodies, (4) improve current vaccines to combat acute and chronic infections of hepatitis B, and (5) develop entry inhibitors for future clinical applications.
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Affiliation(s)
- Dieter Glebe
- Institute of Medical Virology, Justus-Liebig University of Giessen, Frankfurter Strasse 107, D-35392 Giessen, Germany.
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13
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Deng Q, Zhai JW, Michel ML, Zhang J, Qin J, Kong YY, Zhang XX, Budkowska A, Tiollais P, Wang Y, Xie YH. Identification and characterization of peptides that interact with hepatitis B virus via the putative receptor binding site. J Virol 2006; 81:4244-54. [PMID: 17192308 PMCID: PMC1866126 DOI: 10.1128/jvi.01270-06] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
Abstract
A direct involvement of the PreS domain of the hepatitis B virus (HBV) large envelope protein, and in particular amino acid residues 21 to 47, in virus attachment to hepatocytes has been suggested by many previous studies. Several PreS-interacting proteins have been identified. However, they share few common sequence motifs, and a bona fide cellular receptor for HBV remains elusive. In this study, we aimed to identify PreS-interacting motifs and to search for novel HBV-interacting proteins and the long-sought receptor. PreS fusion proteins were used as baits to screen a phage display library of random peptides. A group of PreS-binding peptides were obtained. These peptides could bind to amino acids 21 to 47 of PreS1 and shared a linear motif (W1T2X3W4W5) sufficient for binding specifically to PreS and viral particles. Several human proteins with such a motif were identified through BLAST search. Analysis of their biochemical and structural properties suggested that lipoprotein lipase (LPL), a key enzyme in lipoprotein metabolism, might interact with PreS and HBV particles. The interaction of HBV with LPL was demonstrated by in vitro binding, virus capture, and cell attachment assays. These findings suggest that LPL may play a role in the initiation of HBV infection. Identification of peptides and protein ligands corresponding to LPL that bind to the HBV envelope will offer new therapeutic strategies against HBV infection.
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Affiliation(s)
- Qiang Deng
- State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes of Biological Sciences, Chinese Academy of Sciences, and Ruijin Hospital, Department of Infectious Diseases, Shanghai, China
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14
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Iwasaki Y, Ueda M, Yamada T, Kondo A, Seno M, Tanizawa K, Kuroda S, Sakamoto M, Kitajima M. Gene therapy of liver tumors with human liver-specific nanoparticles. Cancer Gene Ther 2006; 14:74-81. [PMID: 16990844 DOI: 10.1038/sj.cgt.7700990] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/11/2023]
Abstract
The development of safe and efficient liver-specific gene delivery approaches offers new perspectives for the treatment of liver disease, in particular, liver cancer. We evaluated the therapeutic potential of hepatotropic nanoparticles for gene therapy of liver tumor. These nanoparticles do not contain a viral genome and display the hepatitis B virus L antigen, which is essential to confer hepatic specificity. It has not been shown whether a therapeutic effect could be obtained using L nanoparticles in a human liver tumor xenograft model. Rats bearing human hepatic (NuE) and non-hepatic tumors were injected with L nanoparticles containing a green fluorescent protein (GFP) expression plasmid. GFP expression was observed only in NuE-derived tumors but not in the non-hepatic tumor. The potential for treatment of liver tumors was analyzed using L nanoparticles containing the herpes simplex virus thymidine kinase gene, in conjunction with ganciclovir pro-drug administration. The growth of NuE-derived tumors in L particle-injected rats was significantly suppressed, but not of the non-hepatic tumor control. In summary, this is the first demonstration that nanoparticles could be used for delivery of therapeutic genes with anti-tumor activity into human liver tumors. This intravenous delivery system may be one of the major advantages as compared to many other viral vector systems.
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Affiliation(s)
- Y Iwasaki
- Department of Surgery, School of Medicine, Keio University, Tokyo, Japan
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15
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Severi T, Ying C, Vermeesch JR, Cassiman D, Cnops L, Verslype C, Fevery J, Arckens L, Neyts J, van Pelt JF. Hepatitis B virus replication causes oxidative stress in HepAD38 liver cells. Mol Cell Biochem 2006; 290:79-85. [PMID: 16960659 DOI: 10.1007/s11010-006-9167-x] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2005] [Accepted: 02/22/2006] [Indexed: 02/08/2023]
Abstract
UNLABELLED We used human hepatoma HepAD38 cells, in which HBV production is under the control of a tetracycline-regulated promotor, to investigate changes induced in the host cell by HBV replication that could contribute to malignant transformation. Parameters of oxidative stress (malondialdehyde, glutathione) and cell proliferation were determined at different times after induction (0-96 h). In HBV-producing cells, the redox status peaked at 72 h. cDNA micro array analysis at 72 h post induction revealed 3 groups of genes that were up-regulated by HBV: (i) heat shock proteins, (ii) oxidative and metabolic stress and (iii) growth and apoptosis related genes. Continuous HBV production did not accelerate karyotypic changes in cells cultured for 4 months (18 passages). IN CONCLUSION HBV replication modulates host gene expression and induces oxidative stress. In this HepAD38 model early events (0-4 days) in the host cell after induction of HBV replication can be studied under strictly defined conditions.
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Affiliation(s)
- Tamara Severi
- Department of Hepatology, University Hospital Gasthuisberg, Leuven, Belgium
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16
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Owada T, Matsubayashi K, Sakata H, Ihara H, Sato S, Ikebuchi K, Kato T, Azuma H, Ikeda H. Interaction between desialylated hepatitis B virus and asialoglycoprotein receptor on hepatocytes may be indispensable for viral binding and entry. J Viral Hepat 2006; 13:11-8. [PMID: 16364077 DOI: 10.1111/j.1365-2893.2005.00648.x] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
The cellular receptor for hepatitis B virus (HBV) infection has not yet been identified. The purpose of this study was to address the possibility of participation by desialylated HBV and the asialoglycoprotein receptor (ASGP-R) exclusively expressed on liver parenchymal cells, in infection. Assays for viral binding and entry were performed by culturing a hepatoblastoma cell line, HepG2, and HBV particles derived from the HBV carrier in the presence or absence of neuraminidase (NA). Viral binding and entry were clearly enhanced in the presence of NA, and the enhancement of the binding could be blocked by asialo-fetuin and ethylenediamine-tetraacetic acid (EDTA). In addition, covalently closed circular (CCC)-DNA, as a marker of infectivity, was detected in the presence of NA, but not in its absence. The optimal concentration of NA raised infectivity more than 1000 times. We concluded that this method makes it feasible to evaluate the infectivity of HBV in vitro and that ASGP-R may be a specific HBV receptor once viral particles are desialylated.
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Affiliation(s)
- T Owada
- Hokkaido Red Cross Blood Center, Nishi-ku, Sapporo-shi, Hokkaido, Japan.
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17
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Deng Q, Zhuang M, Kong YY, Xie YH, Wang Y. Screening for PreS specific binding ligands with a phage displayed peptides library. World J Gastroenterol 2005; 11:4018-23. [PMID: 15996026 PMCID: PMC4502097 DOI: 10.3748/wjg.v11.i26.4018] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To construct a random peptide phage display library and search for peptides that specifically bind to the PreS region of hepatitis B virus (HBV).
METHODS: A phage display vector, pFuse8, based on the gene 8 product (pVIII) of M13 phage was made and used to construct a random peptide library. E.coli derived thioredoxin-PreS was purified with Thio-bond beads, and exploited as the bait protein for library screening. Five rounds of bio-panning were performed. The PreS-binding specificities of enriched phages were characterized with phage ELISA assay.
RESULTS: A phage display vector was successfully constructed as demonstrated to present a pVIII fused HBV PreS1 epitope on the phage surface with a high efficiency. A cysteine confined random peptide library was constructed containing independent clones exceeding 5±108 clone forming unit (CFU). A pool of phages showing a PreS-binding specificity was obtained after the screening against thio-PreS with an enrichment of approximately 400 times. Five phages with high PreS-binding specificities were selected and characterized. Sequences of the peptides displayed on these phages were determined.
CONCLUSION: A phage library has been constructed, with random peptides displaying as pVIII-fusion proteins. Specific PreS-binding peptides have been obtained, which may be useful for developing antivirals against HBV infection.
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Affiliation(s)
- Qiang Deng
- Institute of Biochemistry and Cell Biology, 320 Yue-Yang Road, Shanghai 200031, China
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18
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Yeh CT, Lai HY, Chu SP, Tseng IC. Anti-sense expression of a metallopeptidase gene enhances nuclear entry of HBV-DNA. Biochem Biophys Res Commun 2004; 323:32-7. [PMID: 15351696 DOI: 10.1016/j.bbrc.2004.08.079] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2004] [Indexed: 01/14/2023]
Abstract
Although several putative hepatitis B virus (HBV) receptors have been identified, none of them is capable of initiating HBV replication in a non-permissive human cell line. Using an Epstein-Barr virus-based extrachromosomal replication system, we have screened through a human liver cDNA library and successfully identified a clone capable of facilitating nuclear transport of HBV-DNA during the early phase of HBV infection. This clone contained a cDNA encoding a metallopeptidase-like protein in anti-sense orientation. Pretreatment of naïve HepG2 cells with 1,10-phenanthroline, an inhibitor for liver metallopeptidases, led to nuclear entry of HBV-DNA after HBV infection. However, cccDNA was still undetectable in the nuclei, indicating other cellular factors required to complete the replication cycle were still missing. Our present data suggest that in the initial stage of HBV infection, liver metallopeptidase constitutes a barrier for effective nuclear entry of HBV genomic DNA. Attenuation of metallopeptidase activity may facilitate HBV infection.
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Affiliation(s)
- Chau-Ting Yeh
- Liver Research Unit, Chang Gung Memorial Hospital and Chang Gung University, College of Medicine, Taipei, Taiwan, ROC.
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19
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Ying C, Van Pelt JF, Van Lommel A, Van Ranst M, Leyssen P, De Clercq E, Neyts J. Sulphated and sulphonated polymers inhibit the initial interaction of hepatitis B virus with hepatocytes. Antivir Chem Chemother 2002; 13:157-64. [PMID: 12448688 DOI: 10.1177/095632020201300302] [Citation(s) in RCA: 21] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/15/2023] Open
Abstract
The initial step during hepatitis B virus (HBV) infection is the specific attachment of the virus to the hepatocyte. Here we studied whether the binding of HBV to hepatocytes can, as is the case with most other enveloped viruses, be blocked by polyanionic compounds. Viral particles produced by HepAD38 cells were used as inoculum and HBV-negative HepG2 cells, as well as primary human hepatocytes, as target cells. Three sulphated polymers, that is, PAVAS (a co-polymer of acrylic acid with vinyl alcohol sulphate), heparin and dextran sulphate (DS) (MW 5000), and the sulphonated polymer PAMPS [poly(2-acryl-amido-2-methyl-1-propanesulfonic acid] (MW approximately 7000-12000), proved strong inhibitors of the binding of HBV to HepG2 cells and primary hepatocytes. The 50% effective concentration (EC50) for inhibition of HBV binding to HepG2 cells by PAVAS, heparin, DS and PAMPS was 1.3 microg/ml, 1.6 microg/ml, 1.8 microg/ml and 3.3 microg/ml, respectively, and to primary hepatocytes 1.6 microg/ml (PAVAS), 1.6 microg/ml (heparin), 2.6 microg/ml (DS) and 4.1 microg/ml (PAMPS). These values are in the same range as the concentrations required for these compounds to prevent such viruses as herpesviruses and HIV from binding to cells. These findings may be helpful in elucidating the mechanism of the initial interaction of HBV with hepatocytes.
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Affiliation(s)
- C Ying
- Rega Institute for Medical Research, Laboratory of Virology, KU Leuven, Belgium
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20
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De Falco S, Ruvoletto MG, Verdoliva A, Ruvo M, Raucci A, Marino M, Senatore S, Cassani G, Alberti A, Pontisso P, Fassina G. Cloning and expression of a novel hepatitis B virus-binding protein from HepG2 cells. J Biol Chem 2001; 276:36613-23. [PMID: 11389143 DOI: 10.1074/jbc.m102377200] [Citation(s) in RCA: 62] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
Abstract
A direct involvement of the hepatitis B virus (HBV) preS1-(21-47) sequence in virus attachment to cell membrane receptor(s) and the presence on the plasma membranes of HepG2 cells of protein(s) with receptor activity for HBV have been suggested by many previous experiments. In this study, by using a tetravalent derivative of the preS1-(21-47) sequence, we have isolated by affinity chromatography from detergent-solubilized HepG2 plasma membranes a 44-kDa protein (HBV-binding protein; HBV-BP), which was found to closely correspond to the human squamous cell carcinoma antigen 1 (SCCA1), a member of the ovalbumin family of serine protease inhibitors. Comparison of SCCA1 sequence with the sequence of the corresponding HBV-BP cDNA, cloned by polymerase chain reaction starting from RNA poly(A)(+) fractions extracted from HepG2 cells, indicated the presence of only four nucleotide substitutions in the coding region, leading to three amino acid changes. Intact recombinant HBV-BP lacked inhibitory activity for serine proteases such as alpha-chymotrypsin and trypsin but inhibited with high potency cysteine proteases such as papain and cathepsin L. Direct binding experiments confirmed the interaction of recombinant HBV-BP with the HBV preS1 domain. HepG2 cells overexpressing HBV-BP after transfection of corresponding cDNA showed a virus binding capacity increased by 2 orders of magnitude compared with untransfected cells, while Chinese hamster ovary cells, which normally do not bind to HBV, acquired susceptibility to HBV binding after transfection. Native HBV particle entry was enhanced in transfected cells. Both recombinant HBV-BP and antibodies to recombinant HBV-BP blocked virus binding and internalization in transfected cells as well as in primary human hepatocytes in a dose-dependent manner. Our findings suggest that this protein plays a major role in HBV infection.
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MESH Headings
- Amino Acid Sequence
- Animals
- Antigens, Neoplasm/chemistry
- Base Sequence
- Binding, Competitive
- CHO Cells
- Cathepsin L
- Cathepsins/antagonists & inhibitors
- Cell Line
- Cell Membrane/chemistry
- Cells, Cultured
- Chromatography
- Chymotrypsin/metabolism
- Cloning, Molecular
- Cricetinae
- Cysteine Endopeptidases
- DNA, Complementary/metabolism
- Dose-Response Relationship, Drug
- Hepatitis B virus/metabolism
- Hepatocytes/metabolism
- Humans
- Kinetics
- Molecular Sequence Data
- Papain/antagonists & inhibitors
- Poly A/metabolism
- Protease Inhibitors
- Protein Binding
- Protein Structure, Tertiary
- Rats
- Receptors, Virus/biosynthesis
- Receptors, Virus/chemistry
- Receptors, Virus/metabolism
- Recombinant Proteins/metabolism
- Sequence Homology, Amino Acid
- Serpins
- Time Factors
- Transfection
- Trypsin/metabolism
- Tumor Cells, Cultured
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Affiliation(s)
- S De Falco
- TECNOGEN S.C.p.A., Parco Scientifico, 81015 Piana di Monte Verna (CE), Caserta 81015, Italy
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21
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Heinz D, Peters M, Prange R, Gerken G, Rose-John S. Possible role of human interleukin-6 and soluble interleukin-6 receptor in hepatitis B virus infection. J Viral Hepat 2001; 8:186-93. [PMID: 11380796 DOI: 10.1046/j.1365-2893.2001.00281.x] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/28/2022]
Abstract
Human interleukin-6 has been shown to promote hepatitis B virus (HBV) infection. However, it is not clear whether this influence is the result of a direct interaction between interleukin-6 (IL-6) and the HBV envelope proteins or of a rather indirect mechanism. A direct interaction of IL-6 and the preS region of the large envelope protein (L-protein) of HBV has been reported. In this study we assessed the binding of IL-6 and of the IL-6 receptor subunits to the preS region of the L-protein of HBV. Binding of IL-6 and IL-6 receptor subunits sIL-6R and gp130 to preS was assessed by immunoprecipitation with recombinant preS proteins. In patient sera IL-6 and sIL-6R concentrations were analysed with respect to the course of hepatitis B infection during and after interferon-alpha (IFN-alpha) therapy. The IL-6 and IL-6 receptor subunits could not be precipitated with recombinant preS proteins. In sera of patients who responded to IFN-alpha therapy by virus elimination, a significant increase in sIL-6R concentration was measured. No increase in sIL-6R levels was seen in patients who did not respond to IFN-alpha. Hence, IL-6 and IL-6 receptor subunits do not bind to preS directly. A possible role for sIL-6R in the elimination of HBV infection is discussed.
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Affiliation(s)
- D Heinz
- I. Medizinische Klinik, Abteilung Pathophysiologie, Johannes Gutenberg Universität Mainz, Mainz, Germany
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22
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Smela ME, Currier SS, Bailey EA, Essigmann JM. The chemistry and biology of aflatoxin B(1): from mutational spectrometry to carcinogenesis. Carcinogenesis 2001; 22:535-45. [PMID: 11285186 DOI: 10.1093/carcin/22.4.535] [Citation(s) in RCA: 186] [Impact Index Per Article: 7.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023] Open
Abstract
Dietary exposure to aflatoxin B(1) (AFB(1)) is associated with an increased incidence of hepatocellular carcinoma (HCC), especially in populations in which exposure to hepatitis B virus (HBV) is a common occurrence. Most HCC samples from people living where HBV is prevalent have one striking mutational hotspot: a GC-->TA transversion at the third position of codon 249 of the p53 gene. In this review, the chemical reaction of an electrophilic derivative of aflatoxin with specific DNA sequences is examined, along with the types of mutations caused by AFB(1) and the sequence context dependence of those mutations. An attempt is made to assign the source of these mutations to specific chemical forms of AFB(1)-DNA damage. In addition, epidemiological and experimental data are examined regarding the synergistic effects of AFB(1) and HBV on HCC formation and the predominance of one hotspot GC-->TA transversion in the p53 gene of affected individuals.
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Affiliation(s)
- M E Smela
- Department of Chemistry and Division of Bioengineering and Environmental Health Massachusetts Institute of Technology, Cambridge, MA 02139, USA
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23
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Abstract
Annexins are ubiquitous multifunctional Ca2+ and phospholipid-binding proteins whose mechanism of function remains largely unknown. The accumulated in vitro experimental evidence indicates that ATP and GTP are functional ligands for nucleotide-sensitive annexin isoforms. Such nucleotide binding could modulate Ca2+ homeostasis, vesicular transport and/or signal transduction pathways and link them to cellular energy metabolism. Alternatively, since annexins are able to interact with other nucleotide-utilizing proteins, such as various kinases, GTPases and structural proteins, these proteins could influence the guanine nucleotide exchange metabolism and/or control the activity of various G proteins. The nucleotide-binding properties of annexins may affect the development or maintenance of some pathologies and diseases in which changes in physiological concentrations of purine nucleotides or disruption of Ca2+ homeostasis are crucial targets.
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Affiliation(s)
- J Bandorowicz-Pikula
- Department of Cellular Biochemistry, Nencki Institute of Experimental Biology, Warsaw, Poland.
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24
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Ohta A, Sekimoto M, Sato M, Koda T, Nishimura S, Iwakura Y, Sekikawa K, Nishimura T. Indispensable role for TNF-alpha and IFN-gamma at the effector phase of liver injury mediated by Th1 cells specific to hepatitis B virus surface antigen. JOURNAL OF IMMUNOLOGY (BALTIMORE, MD. : 1950) 2000; 165:956-61. [PMID: 10878371 DOI: 10.4049/jimmunol.165.2.956] [Citation(s) in RCA: 57] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
We report the development and characterization of a novel model of severe hepatitis induced against hepatitis B virus surface Ag (HBsAg). HBsAg was successfully targeted into the liver in soluble form. Using this unique property of HBsAg, we established a liver injury model induced by HBsAg-specific Th1 cells. Severe liver injury was induced in C57BL/6 mice by injection of HBsAg together with HBsAg-specific Th1 cells. Histochemical examination demonstrated extensive necroinflammatory hepatic lesions in these animals. Application of this liver injury model to mutant or gene knockout mice enabled us to define the effector mechanisms of Th1 cells in fulminant hepatitis. When Fas-deficient lpr mice were used as recipients, a similar degree of liver injury was induced as in wild-type mice. Moreover, HBsAg-specific Th1 cells obtained from perforin-/- mice could induce severe liver injury in both wild-type and lpr mice. These results indicated that neither Fas ligand nor perforin are essential for Th1-mediated liver injury in this model. Pretreatment with anti-TNF-alpha mAb prevented liver injury, whereas severe liver injury was induced in TNF-alpha-/- mice. Moreover, IFN-gamma receptor-deficient mice were resistant to Th1-mediated liver injury. Therefore, TNF-alpha and IFN-gamma, which were produced by HBsAg-specific Th1 cells during the effector phase, appeared to be indispensable in the pathogenesis of fulminant hepatitis.
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Affiliation(s)
- A Ohta
- Division of Immunoregulation, Section of Disease Control, Institute for Genetic Medicine, and Division of Biological Sciences, Graduate School of Science, Hokkaido University, Sapporo, Japan
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25
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Zoulim F. Therapy of chronic hepatitis B virus infection: inhibition of the viral polymerase and other antiviral strategies. Antiviral Res 1999; 44:1-30. [PMID: 10588330 DOI: 10.1016/s0166-3542(99)00056-x] [Citation(s) in RCA: 35] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Chronic hepatitis B infection remains a major public health problem worldwide. The hepatitis B virus belongs to the family of hepadnaviruses that replicate their DNA genome via a reverse transcription pathway. The chronicity of infection in infected hepatocytes is maintained by the persistence of the viral covalently closed circular DNA. The main strategies to combat chronic HBV infection rely on the stimulation of the specific antiviral immune response and on the inhibition of viral replication. While the prolonged administration of reverse transcriptase inhibitors is most often associated with a control of viral replication rather than eradication, it may select for resistant mutants. The search for new viral targets is therefore mandatory to design combination strategies to prevent the emergence of resistant mutants and eventually clear viral infection.
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26
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De Meyer S, Gong Z, Depla E, Maertens G, Yap SH. Involvement of phosphatidylserine and non-phospholipid components of the hepatitis B virus envelope in human Annexin V binding and in HBV infection in vitro. J Hepatol 1999; 31:783-90. [PMID: 10580574 DOI: 10.1016/s0168-8278(99)80278-5] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
BACKGROUND/AIMS We have previously demonstrated that human liver Annexin V (hAV), a Ca2+-dependent phospholipid binding protein, binds specifically to small HBsAg (SHBsAg). Because of the propensity of AV to bind phospholipids, we here examine the role of phospholipids, as component of the HBV envelope, in binding to hAV and in HBV infection. METHODS The influence of phospholipids (phosphatidylserine and phosphatidylcholine) on the binding of hAV to SHBsAg or to anti-hAV monoclonals was determined by ELISA. Their influence on HBV infection was investigated using an in vitro HBV infection assay. RESULTS Two monoclonals, specific against hAV, were able to block the binding of hAV to SHBsAg and recognized different epitopes of hAV. The binding of one of these monoclonals to hAV could be inhibited by phosphatidylserine, but not by phosphatidylcholine. Further experiments revealed that phosphatidylserine could also inhibit the binding of hAV to SHBsAg and could even prevent HBV infection in vitro. Phosphatidylcholine had no effect on the binding of hAV to SHBsAg and could not prevent HBV infection in vitro. However, since phosphatidylserine was not able to abolish the binding of the other blocking monoclonal to hAV, a non-phospholipid component of the HBV envelope must also be involved in hAV binding. CONCLUSIONS These results indicate that phosphatidylserine and a non-phospholipid component of the HBV envelope are involved in hAV binding and in HBV infection.
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Affiliation(s)
- S De Meyer
- Department of Liver and Pancreatic Diseases, University Hospital Gasthuisberg, Catholic University Leuven, Belgium
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27
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De Meyer S, Depla E, Maertens G, Soumillion A, Yap SH. Characterization of small hepatitis B surface antigen epitopes involved in binding to human annexin V. J Viral Hepat 1999; 6:277-85. [PMID: 10607242 DOI: 10.1046/j.1365-2893.1999.00167.x] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Previously, we have shown that small hepatitis B surface antigen (SHBsAg) binds specifically to human annexin V (hAV) and that hAV plays a key role in the initial steps of hepatitis B virus (HBV) infection. We have also demonstrated the spontaneous development of anti-idiotypic antibodies (antibodies to HBsAg Ab2) in rabbits immunized with hAV. As Ab2 is able to inhibit the binding of hAV to SHBsAg, Ab2 might contain epitope(s) mimicking a region of hAV for binding to SHBsAg. Identification of this epitope will therefore reveal a SHBsAg sequence involved in hAV binding. Using a panel of synthetic peptides covering the region of SHBsAg located on the outer surface of the virus, binding studies showed that the region incorporating amino acids (aa) 125-131 of SHBsAg is important for binding to Ab2 and consequently also for binding to hAV. Further experiments revealed that not only this region, but also the region incorporating aa 158-169, is involved in the binding of SHBsAg to hAV. As these regions are located in the structural vicinity according to the topological model of HBsAg proposed by Chen et al., our findings suggest that these regions are parts of a conformational epitope of SHBsAg for binding to hAV. Because of the crucial role of hAV in HBV infection, further studies on the HBsAg epitopes for hAV binding may lead to the development of a new generation of vaccines or molecules for prevention and for treatment of patients with chronic hepatitis B.
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Affiliation(s)
- S De Meyer
- Department of Liver and Pancreatic Diseases, University Hospital Gasthuisberg, Catholic University Leuven, Belgium
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