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The androgen receptor expression and its activity have different relationships with prognosis in hepatocellular carcinoma. Sci Rep 2020; 10:22046. [PMID: 33328560 PMCID: PMC7744520 DOI: 10.1038/s41598-020-79177-2] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2020] [Accepted: 12/02/2020] [Indexed: 01/01/2023] Open
Abstract
The role of the Androgen Receptor (AR) expression and its activity in the prognosis of hepatocellular carcinoma (HCC) remains inconclusive. The aim of this study is to analyze the role of the AR expression and its activity as prognostic biomarkers in HCC. Three-hundred and thirty-seven patients from The Cancer Genome Atlas (TCGA) (107 females; 59.42 years [SD = 13.0]) were included. To infer AR activity, the expression-profile of previously validated androgen responsive genes (ARGs) was included. AR activity was shown by the AR-Score-21 (21 ARGs) and AR-Score-13 (13 ARGs) that were computed based on the expression of the selected ARGs. Those ARGs whose expression was significantly different between histological grades were used for computing two new AR-Scores. HCC patients with higher AR expression showed a higher median overall survival (OS). AR-Score 21 and AR-Score-13 did not show any association with prognosis. Six of the 21 ARGs of the AR-Score-21 and 7 of the 13 ARGs of the AR-Score-13 showed a significant different expression profile among histological grades. Based on these differences, another two AR-Scores were computed (AR-Score-6 and AR-Score-7). They showed the relative increase of upregulated to downregulated ARGs in high-grade HCC. Higher AR activity inferred by these AR-Scores was associated with worse outcomes. The expression of AR is associated with a better prognosis in HCC. However, the activity of the AR seems to be qualitatively different among histological grades. The AR activity inferred by the shifted ARGs is associated with a worse prognosis in HCC patients.
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Haga Y, Kanda T, Nakamoto S, Nakamura M, Sasaki R, Wu S, Yokosuka O. Interferon induces interleukin 8 and bone marrow stromal cell antigen 2 expression, inhibiting the production of hepatitis B virus surface antigen from human hepatocytes. Biochem Biophys Res Commun 2017; 486:858-863. [PMID: 28363866 DOI: 10.1016/j.bbrc.2017.03.150] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2017] [Accepted: 03/27/2017] [Indexed: 02/07/2023]
Abstract
Hepatitis B virus (HBV) surface antigen (HBsAg) loss is one of the treatment goals of chronic HBV infection. Bone marrow stromal cell antigen 2 (BST2) is one of the interferon (IFN)-stimulated genes (ISGs) and inhibits the release of various enveloped viruses. Here we examined the effects of antiviral treatment on HBsAg levels and its intracellular mechanism in HBsAg-producing hepatocytes. In PLC/PRF/5 and Huh1, IFNα-2a treatment decreased HBsAg levels in their conditioned media. Upregulation of interleukin 8 (IL8), toll-like receptor 2 (TLR2) and interferon gamma-induced protein 10 (IP10) mRNAs was associated with the reduction of HBsAg in both PLC/PRF/5 and Huh1. The HBsAg level was upregulated by knockdown of IL8, TLR2 or IP10. Exogenous addition of IL8 enhanced BST2 promoter activity and BST2 mRNA expression. Additionally, knockdown of IL8 could lead to the downregulation of BST2 mRNA. Transfection of poly(I-C) enhanced IL8 and BST2 mRNA expression and inhibited HBsAg secretion from PLC/PRF/5 cells. In conclusion, IL8 might play an important role in the enhancement of BST2 and be involved in HBsAg eradication.
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Affiliation(s)
- Yuki Haga
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba 260-8670, Japan
| | - Tatsuo Kanda
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba 260-8670, Japan.
| | - Shingo Nakamoto
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba 260-8670, Japan; Department of Molecular Virology, Chiba University, Graduate School of Medicine, Chiba 260-8670, Japan
| | - Masato Nakamura
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba 260-8670, Japan
| | - Reina Sasaki
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba 260-8670, Japan
| | - Shuang Wu
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba 260-8670, Japan
| | - Osamu Yokosuka
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba 260-8670, Japan
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Sasaki R, Kanda T, Nakamura M, Nakamoto S, Haga Y, Wu S, Shirasawa H, Yokosuka O. Possible Involvement of Hepatitis B Virus Infection of Hepatocytes in the Attenuation of Apoptosis in Hepatic Stellate Cells. PLoS One 2016; 11:e0146314. [PMID: 26731332 PMCID: PMC4701422 DOI: 10.1371/journal.pone.0146314] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2015] [Accepted: 12/15/2015] [Indexed: 01/05/2023] Open
Abstract
Background The induction of apoptosis in hepatic stellate cells (HSCs) is a promising therapeutic strategy against hepatitis B virus (HBV)-related hepatic fibrosis. The underlying mechanisms of apoptosis in HSCs, however, are unknown under consideration of HBV infection. In this study, the effects of HBV on apoptosis and endoplasmic reticulum (ER) stress signaling in HSCs were examined. Methods The effects of conditioned media (CM) from HepG2.2.15 on apoptosis induced by the proteasome inhibitor MG132 in LX-2 and HHSteC were studied in regard to c-Jun. In combination with c-Fos, c-Jun forms the AP-1 early response transcription factor, leading to AP-1 activation, signal transduction, endoplasmic reticulum (ER) stress and apoptosis. Results In LX-2 cells, MG132 treatment was associated with the phosphorylation of c-Jun, activation of AP-1 and apoptosis. However, in the presence of CM from HepG2.2.15, these phenomena were attenuated. In HHSteC cells, similar results were observed. HBV genomic DNA is not involved in the process of HSC apoptosis. It is possible that HBeAg has an inhibitory effect on MG132-induced apoptosis in LX-2. We also observed the upregulation of several ER stress-associated genes, such as cAMP responsive element binding protein 3-like 3, inhibin-beta A and solute carrier family 17-member 2, in the presence of CM from HepG2.2.15, or CM from PXB cells infected with HBV. Conclusions HBV inhibits the activation of c-Jun/AP-1 in HSCs, contributing to the attenuation of apoptosis and resulting in hepatic fibrosis. HBV also up-regulated several ER stress genes associated with cell growth and fibrosis. These mechanistic insights might shed new light on a treatment strategy for HBV-associated hepatic fibrosis.
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Affiliation(s)
- Reina Sasaki
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, 260–8670, Japan
| | - Tatsuo Kanda
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, 260–8670, Japan
- * E-mail:
| | - Masato Nakamura
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, 260–8670, Japan
| | - Shingo Nakamoto
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, 260–8670, Japan
- Department of Molecular Virology, Chiba University, Graduate School of Medicine, Chiba, 260–8670, Japan
| | - Yuki Haga
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, 260–8670, Japan
| | - Shuang Wu
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, 260–8670, Japan
| | - Hiroshi Shirasawa
- Department of Molecular Virology, Chiba University, Graduate School of Medicine, Chiba, 260–8670, Japan
| | - Osamu Yokosuka
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, 260–8670, Japan
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Sarkar N, Panigrahi R, Pal A, Biswas A, Singh SP, Kar SK, Bandopadhyay M, Das D, Saha D, Kanda T, Sugiyama M, Chakrabarti S, Banerjee A, Chakravarty R. Expression of microRNA-155 correlates positively with the expression of Toll-like receptor 7 and modulates hepatitis B virus via C/EBP-β in hepatocytes. J Viral Hepat 2015; 22:817-27. [PMID: 25720442 DOI: 10.1111/jvh.12390] [Citation(s) in RCA: 36] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/26/2014] [Accepted: 11/15/2014] [Indexed: 12/11/2022]
Abstract
Effective recognition of viral infection and successive activation of antiviral innate immune responses are vital for host antiviral defence, which largely depends on multiple regulators, including Toll-like receptors (TLRs) and microRNAs. Several early reports suggest that specific TLR-mediated immune responses can control hepatitis B virus (HBV) replication and express differentially with disease outcome. Considering the versatile function of miR-155 in the TLR-mediated innate immune response, we aimed to study the association between miR-155 and TLRs and their subsequent impact on HBV replication using both a HBV-replicating stable cell line (HepG2.2.15) and HBV-infected liver biopsy and serum samples. Our results showed that miR-155 was suppressed during HBV infection and a subsequent positive correlation of miR-155 with TLR7 activation was noted. Further, ectopic expression of miR-155 in vitro reduced HBV load as evidenced from reduced viral DNA, mRNA and subsequently reduced level of secreted viral antigens (HBsAg and HBeAg). Our results further suggested that CCAAT/enhancer-binding protein-β (C/EBP-β), a positive regulator of HBV transcription, was inhibited by miR-155. Taken together, our study established a correlation between miR-155 and TLR7 during HBV infection and also demonstrated in vitro that increased miR-155 level could help to reduce HBV viral load by targeting C/EBP-β.
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Affiliation(s)
- N Sarkar
- ICMR Virus Unit, Kolkata, ID & BG Hospital Campus, Kolkata, India
| | - R Panigrahi
- ICMR Virus Unit, Kolkata, ID & BG Hospital Campus, Kolkata, India
| | - A Pal
- ICMR Virus Unit, Kolkata, ID & BG Hospital Campus, Kolkata, India
| | - A Biswas
- ICMR Virus Unit, Kolkata, ID & BG Hospital Campus, Kolkata, India
| | - S P Singh
- Department of Gastroenterology, SCB Medical College, Cuttack, India.,Kalinga Gastroenterology Foundation, Beam Diagnostics Premises, Cuttack, India
| | - S K Kar
- Department of Gastroenterology, SCB Medical College, Cuttack, India
| | - M Bandopadhyay
- ICMR Virus Unit, Kolkata, ID & BG Hospital Campus, Kolkata, India
| | - D Das
- ICMR Virus Unit, Kolkata, ID & BG Hospital Campus, Kolkata, India
| | - D Saha
- ICMR Virus Unit, Kolkata, ID & BG Hospital Campus, Kolkata, India
| | - T Kanda
- Department of Medicine and Clinical Oncology, Chiba University, Graduate School of Medicine, Chiba, Japan
| | - M Sugiyama
- National Center for Global Health and Medicine (NCGM), Ichikawa, Japan
| | - S Chakrabarti
- ICMR Virus Unit, Kolkata, ID & BG Hospital Campus, Kolkata, India.,National Institute of Cholera and Enteric Diseases, Kolkata, India
| | - A Banerjee
- ICMR Virus Unit, Kolkata, ID & BG Hospital Campus, Kolkata, India
| | - R Chakravarty
- ICMR Virus Unit, Kolkata, ID & BG Hospital Campus, Kolkata, India
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Kanda T, Yokosuka O. The androgen receptor as an emerging target in hepatocellular carcinoma. J Hepatocell Carcinoma 2015; 2:91-9. [PMID: 27508198 PMCID: PMC4918288 DOI: 10.2147/jhc.s48956] [Citation(s) in RCA: 41] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Hepatocellular carcinoma (HCC) is one of the male-dominant liver diseases with poor prognosis, although treatments for HCC have been progressing in the past decades. Androgen receptor (AR) is a member of the nuclear receptor superfamily. Previous studies reported that AR was expressed in human HCC and non-HCC tissues. AR is activated both ligand-dependently and ligand-independently. The latter is associated with a mitogen-activated protein kinase–, v-akt murine thymoma viral oncogene homolog 1–, or signal-transducer and activator of transcription–signaling pathway, which has been implicated in the development of HCC. It has been reported that more than 200 RNA expression levels are altered by androgen treatment. In the liver, androgen-responsive genes are cytochrome P450s, transforming growth factor β, vascular endothelial growth factor, and glucose-regulated protein 78 kDa, which are also associated with human hepatocarcinogenesis. Recent studies also revealed that AR plays a role in cell migration and metastasis. It is possible that cross-talk among AR-signaling, endoplasmic reticulum stress, and innate immune response is important for human hepatocarcinogenesis and HCC development. This review shows that AR could play a potential role in human HCC and represent one of the important target molecules for the treatment of HCC.
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Affiliation(s)
- Tatsuo Kanda
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, Japan
| | - Osamu Yokosuka
- Department of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, Japan
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Kanda T, Jiang X, Yokosuka O. Androgen receptor signaling in hepatocellular carcinoma and pancreatic cancers. World J Gastroenterol 2014; 20:9229-9236. [PMID: 25071315 PMCID: PMC4110552 DOI: 10.3748/wjg.v20.i28.9229] [Citation(s) in RCA: 45] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/17/2014] [Revised: 03/07/2014] [Accepted: 04/30/2014] [Indexed: 02/06/2023] Open
Abstract
Hepatocellular carcinoma (HCC) and pancreatic cancer remain difficult to treat, and despite the ongoing development of new treatments, the overall survival rate has only modestly improved over the past decade. Liver and pancreatic progenitors commonly develop from endoderm cells in the embryonic foregut. A previous study showed that HCC and pancreatic cancer cell lines variably express androgen receptor (AR), and these cancers and the surrounding tissues also express AR. AR is a ligand-dependent transcription factor that belongs to the nuclear receptor superfamily. Androgen response element is present in regulatory elements on the AR-responsive target genes, such as transforming growth factor beta-1 (TGF beta-1) and vascular endothelial growth factor (VEGF). It is well known that the activation of AR is associated with human carcinogenesis in prostate cancer as well as HCC and pancreatic cancer and that GRP78, TGF beta, and VEGF all play important roles in carcinogenesis and cancer development in these cancers. HCC is a male-dominant cancer irrespective of its etiology. Previous work has reported that vertebrae forkhead box A 1/2 are involved in estrogen receptors and/or AR signaling pathways, which may contribute to the gender differences observed with HCC. Our recent work also showed that AR has a critical role in pancreatic cancer development, despite pancreatic cancer not being a male dominant cancer. Aryl hydrocarbon (or dioxin) receptor is also involved in both HCC and pancreatic cancer through the formation of complex with AR. It is possible that AR might be involved in their carcinogenesis through major histocompatibility complex class I chain-related gene A/B. This review article describes AR and its role in HCC and pancreatic cancer and suggests that more specific AR signaling-inhibitors may be useful in the treatment of these "difficult to treat" cancers.
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Jiang X, Kanda T, Wu S, Nakamura M, Miyamura T, Nakamoto S, Banerjee A, Yokosuka O. Regulation of microRNA by hepatitis B virus infection and their possible association with control of innate immunity. World J Gastroenterol 2014; 20:7197-7206. [PMID: 24966589 PMCID: PMC4064064 DOI: 10.3748/wjg.v20.i23.7197] [Citation(s) in RCA: 32] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/17/2013] [Revised: 12/11/2013] [Accepted: 01/03/2014] [Indexed: 02/06/2023] Open
Abstract
Hepatitis B virus (HBV) chronically infects more than 350 million people worldwide. HBV causes acute and chronic hepatitis, and is one of the major causes of cirrhosis and hepatocellular carcinoma. There exist complex interactions between HBV and the immune system including adaptive and innate immunity. Toll-like receptors (TLRs) and TLR-signaling pathways are important parts of the innate immune response in HBV infections. It is well known that TLR-ligands could suppress HBV replication and that TLRs play important roles in anti-viral defense. Previous immunological studies demonstrated that HBV e antigen (HBeAg) is more efficient at eliciting T-cell tolerance, including production of specific cytokines IL-2 and interferon gamma, than HBV core antigen. HBeAg downregulates cytokine production in hepatocytes by the inhibition of MAPK or NF-κB activation through the interaction with receptor-interacting serine/threonine protein kinase. MicroRNAs (miRNAs) are also able to regulate various biological processes such as the innate immune response. When the expressions of approximately 1000 miRNAs were compared between human hepatoma cells HepG2 and HepG2.2.15, which could produce HBV virion that infects chimpanzees, using real-time RT-PCR, we observed several different expression levels in miRNAs related to TLRs. Although we and others have shown that HBV modulates the host immune response, several of the miRNAs seem to be involved in the TLR signaling pathways. The possibility that alteration of these miRNAs during HBV infection might play a critical role in innate immunity against HBV infection should be considered. This article is intended to comprehensively review the association between HBV and innate immunity, and to discuss the role of miRNAs in the innate immune response to HBV infection.
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Curtil C, Enache LS, Radreau P, Dron AG, Scholtès C, Deloire A, Roche D, Lotteau V, André P, Ramière C. The metabolic sensors FXRα, PGC-1α, and SIRT1 cooperatively regulate hepatitis B virus transcription. FASEB J 2013; 28:1454-63. [PMID: 24297698 DOI: 10.1096/fj.13-236372] [Citation(s) in RCA: 39] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
Hepatitis B virus (HBV) genome transcription is highly dependent on liver-enriched, metabolic nuclear receptors (NRs). Among others, NR farnesoid X receptor α (FXRα) enhances HBV core promoter activity and pregenomic RNA synthesis. Interestingly, two food-withdrawal-induced FXRα modulators, peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α) and deacetylase SIRT1, have been found to be associated with HBV genomes ex vivo. Whereas PGC-1α induction was shown to increase HBV replication, the effect of SIRT1 on HBV transcription remains unknown. Here, we showed that, in hepatocarcinoma-derived Huh-7 cells, combined activation of FXRα by GW4064 and SIRT1 by activator 3 increased HBV core promoter-controlled luciferase expression by 25-fold, compared with a 10-fold increase with GW4064 alone. Using cell lines differentially expressing FXRα in overexpression and silencing experiments, we demonstrated that SIRT1 activated the core promoter in an FXRα- and PGC-1α-dependent manner. Maximal activation (>150-fold) was observed in FXRα- and PGC-1α-overexpressing Huh-7 cells treated with FXRα and SIRT1 activators. Similarly, in cells transfected with full-length HBV genomes, maximal induction (3.5-fold) of core promoter-controlled synthesis of 3.5-kb RNA was observed in the same conditions of transfection and treatments. Thus, we identified a subnetwork of metabolic factors regulating HBV replication, strengthening the hypothesis that transcription of HBV and metabolic genes is similarly controlled.
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Affiliation(s)
- Claire Curtil
- 2Centre International de Recherche en Infectiologie, INSERM U1111, 21 Ave. Tony Garnier, 69365 Lyon Cedex 07, France.
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Kanda T, Jiang X, Nakamoto S, Nakamura M, Miyamura T, Wu S, Yokosuka O. Different effects of three interferons L on Toll-like receptor-related gene expression in HepG2 cells. Cytokine 2013; 64:577-583. [PMID: 24041672 DOI: 10.1016/j.cyto.2013.08.010] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2013] [Revised: 08/07/2013] [Accepted: 08/16/2013] [Indexed: 02/07/2023]
Abstract
IFNL1 (IL29), IFNL2 (IL28A) and IFNL3 (IL28B) might play important roles in anti-viral defense. IFNL3 genotypes have been shown to be associated with hepatitis C spontaneous and treatment-induced viral clearance. The effects of IFNL1, IFNL2 and IFNL3 on innate immunity including Toll-like receptor (TLR)-related pathway in human hepatocytes were examined. After G418 screening, we established the human hepatoma stable cell lines HepG2-IL28A, HepG2-IL28B, and HepG2-IL29, expressing IFNL2, IFNL3, and IFNL1 in conditioned medium, respectively, and a control cell line, HepG2-pcDNA3.1. We performed real-time RT-PCR to investigate 84 Toll-like receptor-related gene expressions in triplicate and, using ddCt methods, compared these gene expressions in each cell line. IFNL2, IFNL3 and IFNL1 were respectively detected by ELISA in HepG2-IL28A, HepG2-IL28B and HepG2-IL29. Compared to HepG2-pcDNA3.1 cells, 17 (20.2%), 11 (13.0%) and 16 genes (19.0%) were up-regulated 1.5-fold or more (p<0.05); 10 (11.9%), 2 (2.3%) and 10 genes (11.9%) were 1.5-fold or more down-regulated (p<0.05) in HepG2-IL28A, HepG2-IL28B and HepG2-IL29, respectively. EIF2AK2 and SARM1 were up-regulated among all cells. Of interest, TLR3, TLR4 and related molecules CXCL10 (IP10), IL6, EIF2K2, IFNB1, and IRF1, important genes in the progression of HCV-related pathogenesis and antiviral activities against HCV, in HepG2-IL28B, presented different profiles from those of HepG2-IL28A and HepG2-IL29. IFNL3 induces interferon-stimulated genes (ISGs) that are reportedly associated with the progression of HCV-related pathogenesis and antiviral activities against HCV. IFNL is a powerful modulator of innate immune response and it is supposed that the 3 IFNLs may play different roles in the antiviral activity against HBV and HCV.
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Affiliation(s)
- Tatsuo Kanda
- Department of Gastroenterology and Nephrology, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8677, Japan.
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