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1
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Serna R, Ramrakhiani A, Hernandez JC, Chen CL, Nakagawa C, Machida T, Ray RB, Zhan X, Tahara SM, Machida K. c-JUN inhibits mTORC2 and glucose uptake to promote self-renewal and obesity. iScience 2022; 25:104325. [PMID: 35601917 PMCID: PMC9121277 DOI: 10.1016/j.isci.2022.104325] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2020] [Revised: 12/30/2021] [Accepted: 04/26/2022] [Indexed: 02/08/2023] Open
Abstract
Metabolic syndrome is associated with obesity, insulin resistance, and the risk of cancer. We tested whether oncogenic transcription factor c-JUN metabolically reprogrammed cells to induce obesity and cancer by reduction of glucose uptake, with promotion of the stemness phenotype leading to malignant transformation. Liquid alcohol, high-cholesterol, fat diet (HCFD), and isocaloric dextrin were fed to wild-type or experimental mice for 12 months to promote hepatocellular carcinoma (HCC). We demonstrated 40% of mice developed liver tumors after chronic HCFD feeding. Disruption of liver-specific c-Jun reduced tumor incidence 4-fold and improved insulin sensitivity. Overexpression of c-JUN downregulated RICTOR transcription, leading to inhibition of the mTORC2/AKT and glycolysis pathways. c-JUN inhibited GLUT1, 2, and 3 transactivation to suppress glucose uptake. Silencing of RICTOR or c-JUN overexpression promoted self-renewal ability. Taken together, c-JUN inhibited mTORC2 via RICTOR downregulation and inhibited glucose uptake via downregulation of glucose intake, leading to self-renewal and obesity.
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Affiliation(s)
- Raphael Serna
- Department of Molecular Microbiology and Immunology, University of Southern California, Keck School of Medicine, 2011 Zonal Avenue, HMR503C, Los Angeles, CA 90033, USA
| | - Ambika Ramrakhiani
- Department of Molecular Microbiology and Immunology, University of Southern California, Keck School of Medicine, 2011 Zonal Avenue, HMR503C, Los Angeles, CA 90033, USA
| | - Juan Carlos Hernandez
- Department of Molecular Microbiology and Immunology, University of Southern California, Keck School of Medicine, 2011 Zonal Avenue, HMR503C, Los Angeles, CA 90033, USA
| | - Chia-Lin Chen
- Department of Molecular Microbiology and Immunology, University of Southern California, Keck School of Medicine, 2011 Zonal Avenue, HMR503C, Los Angeles, CA 90033, USA
| | - Chad Nakagawa
- Department of Molecular Microbiology and Immunology, University of Southern California, Keck School of Medicine, 2011 Zonal Avenue, HMR503C, Los Angeles, CA 90033, USA
| | - Tatsuya Machida
- Department of Molecular Microbiology and Immunology, University of Southern California, Keck School of Medicine, 2011 Zonal Avenue, HMR503C, Los Angeles, CA 90033, USA
| | | | - Xiaohang Zhan
- Chinese Academy of Sciences and Peking Union Medical College, Beijing 100050, P.R. China
| | - Stanley M. Tahara
- Department of Molecular Microbiology and Immunology, University of Southern California, Keck School of Medicine, 2011 Zonal Avenue, HMR503C, Los Angeles, CA 90033, USA
| | - Keigo Machida
- Department of Molecular Microbiology and Immunology, University of Southern California, Keck School of Medicine, 2011 Zonal Avenue, HMR503C, Los Angeles, CA 90033, USA
- Southern California Research Center for ALPD and Cirrhosis, Los Angeles, CA, USA
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Akutsu N, Sasaki S, Matsui T, Akashi H, Yonezawa K, Ishigami K, Tsujisaki M, Isshiki H, Yawata A, Yamaoka S, Ban T, Adachi T, Nakahara S, Takagi H, Nakachi K, Tanaka K, Hirano T, Yamamoto I, Kaneto H, Wagatsuma K, Numata Y, Nakase H. Association of the Low-density Lipoprotein Cholesterol/High-density Lipoprotein Cholesterol Ratio with Glecaprevir-pibrentasvir Treatment. Intern Med 2021; 60:3369-3376. [PMID: 34024854 PMCID: PMC8627811 DOI: 10.2169/internalmedicine.7098-21] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Objective The change in serum lipid levels by direct-acting antiviral (DAA) treatment for chronic hepatitis C varies depending on the type of DAA. How the lipid level changes induced by glecaprevir-pibrentasvir (G/P) treatment contribute to the clinical outcome remains unclear. We conducted a prospective observational study to evaluate the effectiveness of G/P treatment and the lipid level changes. Methods The primary endpoint was a sustained virologic response at 12 weeks (SVR12). The total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), and triglyceride (TG) levels and LDL-C/HDL-C (L/H) ratio were measured every two weeks. Patients This study included 101 patients. Seventeen cases of liver cirrhosis and nine cases of DAA retreatment were registered. The G/P treatment period was 8 weeks in 74 cases and 12 weeks in 27 cases. Results SVR12 was evaluated in 96 patients. The rate of achievement of SVR12 in the evaluable cases was 100%. We found significantly elevated TC and LDL-C levels over the observation period compared to baseline. The serum levels of HDL-C did not change during treatment but were significantly increased after treatment compared to baseline. The L/H ratio was significantly increased two weeks after the start of treatment but returned to the baseline after treatment. Conclusion The primary endpoint of the SVR12 achievement rate was 100%. G/P treatment changed the serum lipid levels. Specifically, the TC and LDL-C levels increased during and after treatment, and the HDL-C levels increased after treatment. G/P treatment may be associated with a reduced thrombotic risk. Therefore, validation in large trials is recommended.
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Affiliation(s)
- Noriyuki Akutsu
- Department of Gastroenterology and Hepatology, Sapporo Medical University School of Medicine, Japan
| | - Shigeru Sasaki
- Department of Gastroenterology and Hepatology, Sapporo Medical University School of Medicine, Japan
| | - Takeshi Matsui
- Department of Gastroenterology, Teine Keijinkai Hospital, Japan
| | - Hirofumi Akashi
- Department of Internal Medicine, Saiseikai Otaru Hospital, Japan
| | - Kazuhiko Yonezawa
- Department of Gastroenterology, Kushiro City General Hospital, Japan
| | - Keisuke Ishigami
- Department of Gastroenterology and Hepatology, Sapporo Medical University School of Medicine, Japan
| | | | - Hiroyuki Isshiki
- Department of Gastroenterology, Hakodate Goryoukaku Hospital, Japan
| | - Atsushi Yawata
- Department of Gastroenterology, Hakodate Goryoukaku Hospital, Japan
| | - Satoshi Yamaoka
- Department of Gastroenterology, Sapporo Satozuka Hospital, Japan
| | - Toshihiro Ban
- Department of Gastroenterology, Sapporo Shirakabadai Hospital, Japan
| | - Takeya Adachi
- Department of Gastroenterology, JR Sapporo Hospital, Japan
| | - Seiya Nakahara
- Department of Gastroenterology, Sapporo Teishinkai Hospital, Japan
| | - Hideyasu Takagi
- Department of Gastroenterology, Sapporo Teishinkai Hospital, Japan
| | - Kohei Nakachi
- Department of Medical Oncology, Tochigi Cancer Center, Japan
| | - Katsunori Tanaka
- Department of Gastroenterology, Sapporo Gekakinen Hospital, Japan
| | - Takehiro Hirano
- Department of Gastroenterology and Hepatology, Sapporo Medical University School of Medicine, Japan
| | - Itaru Yamamoto
- Department of Gastroenterology, Obihiro Kyokai Hospital, Japan
| | - Hiroyuki Kaneto
- Department of Gastroenterology, Muroran City General Hospital, Japan
| | - Kohei Wagatsuma
- Department of Gastroenterology and Hepatology, Sapporo Medical University School of Medicine, Japan
| | - Yasunao Numata
- Department of Gastroenterology and Hepatology, Sapporo Medical University School of Medicine, Japan
| | - Hiroshi Nakase
- Department of Gastroenterology and Hepatology, Sapporo Medical University School of Medicine, Japan
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3
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Dimitriadis A, Foka P, Kyratzopoulou E, Karamichali E, Petroulia S, Tsitoura P, Kakkanas A, Eliadis P, Georgopoulou U, Mamalaki A. The Hepatitis C virus NS5A and core proteins exert antagonistic effects on HAMP gene expression: the hidden interplay with the MTF-1/MRE pathway. FEBS Open Bio 2021; 11:237-250. [PMID: 33247551 PMCID: PMC7780115 DOI: 10.1002/2211-5463.13048] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/19/2020] [Revised: 11/12/2020] [Accepted: 11/15/2020] [Indexed: 12/26/2022] Open
Abstract
Hepcidin, a 25-amino acid peptide encoded by the HAMP gene and produced mainly by hepatocytes and macrophages, is a mediator of innate immunity and the central iron-regulatory hormone. Circulating hepcidin controls iron efflux by inducing degradation of the cellular iron exporter ferroportin. HCV infection is associated with hepatic iron overload and elevated serum iron, which correlate with poor antiviral responses. The HCV nonstructural NS5A protein is known to function in multiple aspects of the HCV life cycle, probably exerting its activity in concert with cellular factor(s). In this study, we attempted to delineate the effect of HCV NS5A on HAMP gene expression. We observed that transient transfection of hepatoma cell lines with HCV NS5A resulted in down-regulation of HAMP promoter activity. A similar effect was evident after transduction of Huh7 cells with a recombinant baculovirus vector expressing NS5A protein. We proceeded to construct an NS5A-expressing stable cell line, which also exhibited down-regulation of HAMP gene promoter activity and significant reduction of HAMP mRNA and hepcidin protein levels. Concurrent expression of HCV core protein, a well-characterized hepcidin inducer, revealed antagonism between those two proteins for hepcidin regulation. In attempting to identify the pathways involved in NS5A-driven reduction of hepcidin levels, we ruled out any NS5A-induced alterations in the expression of the well-known hepcidin inducers SMAD4 and STAT3. Further analysis linked the abundance of intracellular zinc ions and the deregulation of the MTF-1/MRE/hepcidin axis with the observed phenomenon. This effect could be associated with distinct phases in HCV life cycle.
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Affiliation(s)
- Alexios Dimitriadis
- Laboratory of Molecular Biology and ImmunobiotechnologyHellenic Pasteur InstituteAthensGreece
| | - Pelagia Foka
- Laboratory of Molecular VirologyHellenic Pasteur InstituteAthensGreece
| | - Eleni Kyratzopoulou
- Laboratory of Molecular Biology and ImmunobiotechnologyHellenic Pasteur InstituteAthensGreece
| | | | | | - Panagiota Tsitoura
- Laboratory of Molecular VirologyHellenic Pasteur InstituteAthensGreece
- Present address:
Laboratory of Molecular Biology and ImmunobiotechnologyHellenic Pasteur InstituteAthensGreece
| | | | - Petros Eliadis
- Laboratory of Molecular Biology and ImmunobiotechnologyHellenic Pasteur InstituteAthensGreece
| | | | - Avgi Mamalaki
- Laboratory of Molecular Biology and ImmunobiotechnologyHellenic Pasteur InstituteAthensGreece
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4
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Yang CW, Hsu HY, Chang HY, Lee YZ, Lee SJ. Natural cardenolides suppress coronaviral replication by downregulating JAK1 via a Na +/K +-ATPase independent proteolysis. Biochem Pharmacol 2020; 180:114122. [PMID: 32592721 PMCID: PMC7314687 DOI: 10.1016/j.bcp.2020.114122] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2020] [Revised: 06/18/2020] [Accepted: 06/22/2020] [Indexed: 12/21/2022]
Abstract
Natural cardenolides suppress coronaviral activity via downregulating JAK1. Natural cardenolides downregulate JAK1 in a Na+/K+-ATPase independent manner. Ouabain (cardenolides) activates Ndfip1/2 and NEDD4 to mediate JAK1 proteolysis. Ndfip1/2 and NEDD4 mediated JAK1 proteolysis attenuates coronaviral activity. An unprecedented biological function of natural cardenolides independent of their membrane target Na+/K+-ATPase is disclosed. Previously, we reported that cardenolides impart anti-transmissible gastroenteritis coronavirus (anti-TGEV) activity through the targeting of Na+/K+-ATPase and its associated PI3K_PDK1_RSK2 signaling. Swine testis cells with Na+/K+-ATPase α1 knocked down exhibited decreased susceptibility to TGEV infectivity and attenuated PI3K_PDK1_RSK2 signaling. Herein, we further explored a Na+/K+-ATPase-independent signaling axis induced by natural cardenolides that also afforded significant anti-coronaviral activity for porcine TGEV and human HCoV-OC43. Using pharmacological inhibition and gene silencing techniques, we found that this anti-TGEV or anti-HCoV-OC43 activity was caused by JAK1 proteolysis and mediated through upstream activation of Ndfip1/2 and its effector NEDD4. This study provides novel insights into the pharmacological effects of natural cardenolides, and is expected to inform their future development as antiviral agents.
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Affiliation(s)
- Cheng-Wei Yang
- Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 35053, Taiwan, ROC
| | - Hsing-Yu Hsu
- Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 35053, Taiwan, ROC
| | - Hsin-Yu Chang
- Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 35053, Taiwan, ROC
| | - Yue-Zhi Lee
- Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 35053, Taiwan, ROC
| | - Shiow-Ju Lee
- Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli 35053, Taiwan, ROC.
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5
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Convery O, Gargan S, Kickham M, Schroder M, O'Farrelly C, Stevenson NJ. The hepatitis C virus (HCV) protein, p7, suppresses inflammatory responses to tumor necrosis factor (TNF)-α via signal transducer and activator of transcription (STAT)3 and extracellular signal-regulated kinase (ERK)-mediated induction of suppressor of cytokine signaling (SOCS)3. FASEB J 2019; 33:8732-8744. [PMID: 31163989 DOI: 10.1096/fj.201800629rr] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
Viruses use a spectrum of immune evasion strategies that enable infection and replication. The acute phase of hepatitis C virus (HCV) infection is characterized by nonspecific and often mild clinical symptoms, suggesting an immunosuppressive mechanism that, unless symptomatic liver disease presents, allows the virus to remain largely undetected. We previously reported that HCV induced the regulatory protein suppressor of cytokine signaling (SOCS)3, which inhibited TNF-α-mediated inflammatory responses. However, the mechanism by which HCV up-regulates SOCS3 remains unknown. Here we show that the HCV protein, p7, enhances both SOCS3 mRNA and protein expression. A p7 inhibitor reduced SOCS3 induction, indicating that p7's ion channel activity was required for optimal up-regulation of SOCS3. Short hairpin RNA and chemical inhibition revealed that both the Janus kinase-signal transducer and activator of transcription (JAK-STAT) and MAPK pathways were required for p7-mediated induction of SOCS3. HCV-p7 expression suppressed TNF-α-mediated IκB-α degradation and subsequent NF-κB promoter activity, revealing a new and functional, anti-inflammatory effect of p7. Together, these findings identify a molecular mechanism by which HCV-p7 induces SOCS3 through STAT3 and ERK activation and demonstrate that p7 suppresses proinflammatory responses to TNF-α, possibly explaining the lack of inflammatory symptoms observed during early HCV infection.-Convery, O., Gargan, S., Kickham, M., Schroder, M., O'Farrelly, C., Stevenson, N. J. The hepatitis C virus (HCV) protein, p7, suppresses inflammatory responses to tumor necrosis factor (TNF)-α via signal transducer and activator of transcription (STAT)3 and extracellular signal-regulated kinase (ERK)-mediated induction of suppressor of cytokine signaling (SOCS)3.
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Affiliation(s)
- Orla Convery
- School of Biochemistry and Immunology, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland
| | - Siobhan Gargan
- School of Biochemistry and Immunology, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland
| | | | | | - Cliona O'Farrelly
- School of Biochemistry and Immunology, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland.,School of Medicine, Trinity College Dublin, Dublin, Ireland
| | - Nigel J Stevenson
- School of Biochemistry and Immunology, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland
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6
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Guo X, Liu WL, Yang D, Shen ZQ, Qiu ZG, Jin M, Li JW. Hepatitis C virus infection induces endoplasmic reticulum stress and apoptosis in human fetal liver stem cells. J Pathol 2019; 248:155-163. [PMID: 30680725 PMCID: PMC7167977 DOI: 10.1002/path.5240] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/06/2018] [Revised: 11/29/2018] [Accepted: 01/16/2019] [Indexed: 12/18/2022]
Abstract
The cellular mechanisms by which hepatitis C virus (HCV) replication might mediate cytopathic effects are controversial and not entirely clear. In this study, we found that blood-borne HCV (bbHCV) infection could lead to endoplasmic reticulum (ER)-stress and mitochondria-related/caspase-dependent apoptosis at the early stages of infection based on use of the highly efficient bbHCV cell culture model established previously. Sections of bbHCV-infected human fetal liver stem cells (hFLSCs) revealed convolution and nonlinear ER, cell vacuolization, swelling of mitochondria, and numerous double membrane vesicles (DMVs). The percentage of apoptotic hFLSCs infected by bbHCV reached 29.8% at 16 h postinfection, and the amount of cytochrome c increased remarkably in the cytosolic protein fraction. However, over time, apoptosis was inhibited due to the activation of NF-κB. The expression of NF-κB-p65, Bcl-xL, XIAP, and c-FLIPL in hFLSCs was increased significantly 24 h after in infection by bbHCV. The accelerated cell death cycles involving apoptosis, regeneration and repair by bbHCV infection might give rise to the development of cirrhosis, and ultimately to hepatocellular carcinogenesis. Copyright © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
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Affiliation(s)
- Xuan Guo
- Tianjin Institute of Environmental and Operational Medicine, Tianjin, PR China.,Research Institute of Chemical Defense, Beijing, PR China.,State Key Laboratory of NBC Protection for Civilian, Beijing, PR China
| | - Wei-Li Liu
- Tianjin Institute of Environmental and Operational Medicine, Tianjin, PR China
| | - Dong Yang
- Tianjin Institute of Environmental and Operational Medicine, Tianjin, PR China
| | - Zhi-Qiang Shen
- Tianjin Institute of Environmental and Operational Medicine, Tianjin, PR China
| | - Zhi-Gang Qiu
- Tianjin Institute of Environmental and Operational Medicine, Tianjin, PR China
| | - Min Jin
- Tianjin Institute of Environmental and Operational Medicine, Tianjin, PR China
| | - Jun-Wen Li
- Tianjin Institute of Environmental and Operational Medicine, Tianjin, PR China
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7
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Chang Z, Wang Y, Zhou X, Long JE. STAT3 roles in viral infection: antiviral or proviral? Future Virol 2018; 13:557-574. [PMID: 32201498 PMCID: PMC7079998 DOI: 10.2217/fvl-2018-0033] [Citation(s) in RCA: 35] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2018] [Accepted: 05/14/2018] [Indexed: 02/06/2023]
Abstract
Signal transducer and activator of transcription 3 (STAT3) is a transcription factor which can be activated by cytokines, growth factor receptors, and nonreceptor-like tyrosine kinase. An activated STAT3 translocates into the nucleus and combines with DNA to regulate the expression of target genes involved in cell proliferation, differentiation, apoptosis and metastasis. Recent studies have shown that STAT3 plays important roles in viral infection and pathogenesis. STAT3 exhibits a proviral function in several viral infections, including those of HBV, HCV, HSV-1, varicella zoster virus, human CMV and measles virus. However, in some circumstances, STAT3 has an antiviral function in other viral infections, such as enterovirus 71, severe acute respiratory syndrome coronavirus and human metapneumovirus. This review summarizes the roles of STAT3 in viral infection and pathogenesis, and briefly discusses the molecular mechanisms involved in these processes.
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Affiliation(s)
- Zhangmei Chang
- Key Laboratory of Medical Molecular Virology of Ministries of Education & Health, Shanghai Medical College of Fudan University, Shanghai 200032, PR China.,Kunshan Center For Disease Control & Prevention, 458 Tongfengxi Road, Kunshan, Jiangsu, 215301, PR China.,Key Laboratory of Medical Molecular Virology of Ministries of Education & Health, Shanghai Medical College of Fudan University, Shanghai 200032, PR China.,Kunshan Center For Disease Control & Prevention, 458 Tongfengxi Road, Kunshan, Jiangsu, 215301, PR China
| | - Yan Wang
- Department of Medical Microbiology & Parasitology, Laboratory of Medical Microbiology, Shanghai Medical College of Fudan University, 138 Yixueyuan R., Shanghai 200032, PR China.,Department of Medical Microbiology & Parasitology, Laboratory of Medical Microbiology, Shanghai Medical College of Fudan University, 138 Yixueyuan R., Shanghai 200032, PR China
| | - Xin Zhou
- Key Laboratory of Medical Molecular Virology of Ministries of Education & Health, Shanghai Medical College of Fudan University, Shanghai 200032, PR China.,Key Laboratory of Medical Molecular Virology of Ministries of Education & Health, Shanghai Medical College of Fudan University, Shanghai 200032, PR China
| | - Jian-Er Long
- Key Laboratory of Medical Molecular Virology of Ministries of Education & Health, Shanghai Medical College of Fudan University, Shanghai 200032, PR China.,Department of Medical Microbiology & Parasitology, Laboratory of Medical Microbiology, Shanghai Medical College of Fudan University, 138 Yixueyuan R., Shanghai 200032, PR China.,Key Laboratory of Medical Molecular Virology of Ministries of Education & Health, Shanghai Medical College of Fudan University, Shanghai 200032, PR China.,Department of Medical Microbiology & Parasitology, Laboratory of Medical Microbiology, Shanghai Medical College of Fudan University, 138 Yixueyuan R., Shanghai 200032, PR China
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8
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Dai CY, Tsai YS, Chou WW, Liu T, Huang CF, Wang SC, Tsai PC, Yeh ML, Hsieh MY, Huang CI, Vanson Liu SY, Huang JF, Chuang WL, Yu ML. The IL-6/STAT3 pathway upregulates microRNA-125b expression in hepatitis C virus infection. Oncotarget 2018; 9:11291-11302. [PMID: 29541414 PMCID: PMC5834265 DOI: 10.18632/oncotarget.24129] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2017] [Accepted: 12/01/2017] [Indexed: 12/31/2022] Open
Abstract
BACKGROUND/AIMS MicroRNA-125b (miR-125b) has been found to regulate inflammation and acts as an oncogene in many cancers. The mechanisms of miR-125b expression during hepatitis C virus (HCV) infection remain to be clarified. The present study aims to identify the factors that might regulate miR-125b expression in HCV infection. RESULTS High expression of miR-125b was found to correlate with HCV infection in replicon cells and in sera from HCV-infected patients, whereas the miR-125b inhibitor reduced HCV gene expression. The interleukin 6 (IL-6)/signal transducer and activator of transcription 3 (STAT3) pathway plays an inducible effect on miR-125b gene expression. STAT3 siRNA or inhibitor could reduce HCV replication. MATERIALS AND METHODS HCV replicon cells Con1 (type 1b) and Huh7/Ava5 (type 1b) were treated with 17-hydroxy-jolkinolide B (HJB) or STAT3 siRNA. Cell viability assay and Renilla Luciferase Assay were used. Fragments of the miR-125b-1 promoter were constructed for the luciferase reporter assay. PSMB8, PSMB9, miR-125b-1, and miR-125b-2 expression was determined using TaqMan® Gene Expression Assays. Western blot analysis was performed to assess protein abundance. CONCLUSIONS This study elucidates a novel pathway for miR-125b in the pathogenesis of chronic HCV infection and suggests it as a possible target for treating HCV infection.
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Affiliation(s)
- Chia-Yen Dai
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Department of Occupational and Environmental Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Health Management Center, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Faculty of Internal Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
- Graduate Institute of Clinical Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
- Center for Infectious Disease and Cancer Research, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Yi-Shan Tsai
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
| | - Wen-Wen Chou
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
| | - Tawei Liu
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
| | - Chung-Feng Huang
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Department of Occupational and Environmental Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Faculty of Internal Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Shu-Chi Wang
- Health Management Center, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
| | - Pei-Chien Tsai
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
| | - Ming-Lun Yeh
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Faculty of Internal Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
- Graduate Institute of Clinical Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Ming-Yen Hsieh
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
| | - Ching-I Huang
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Graduate Institute of Clinical Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Shang-Yin Vanson Liu
- Department of Marine Biotechnology and Resources, National Sun Yat-Sen University, Kaohsiung, Taiwan
| | - Jee-Fu Huang
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Faculty of Internal Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Wan-Long Chuang
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Faculty of Internal Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
- Center for Infectious Disease and Cancer Research, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Ming-Lung Yu
- Hepatobiliary Division, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Department of Occupational and Environmental Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Health Management Center, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- Faculty of Internal Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
- Graduate Institute of Clinical Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
- Center for Infectious Disease and Cancer Research, Kaohsiung Medical University, Kaohsiung, Taiwan
- Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung, Taiwan
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9
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Tsutsumi T, Okushin K, Enooku K, Fujinaga H, Moriya K, Yotsuyanagi H, Aizaki H, Suzuki T, Matsuura Y, Koike K. Nonstructural 5A Protein of Hepatitis C Virus Interferes with Toll-Like Receptor Signaling and Suppresses the Interferon Response in Mouse Liver. PLoS One 2017; 12:e0170461. [PMID: 28107512 PMCID: PMC5249188 DOI: 10.1371/journal.pone.0170461] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/04/2016] [Accepted: 01/05/2017] [Indexed: 02/06/2023] Open
Abstract
The hepatitis C virus nonstructural protein NS5A is involved in resistance to the host immune response, as well as the viral lifecycle such as replication and maturation. Here, we established transgenic mice expressing NS5A protein in the liver and examined innate immune responses against lipopolysaccharide (LPS) in vivo. Intrahepatic gene expression levels of cytokines such as interleukin-6, tumor necrosis factor-α, and interferon-γ were significantly suppressed after LPS injection in the transgenic mouse liver. Induction of the C-C motif chemokine ligand 2, 4, and 5 was also suppressed. Phosphorylation of the signal transducer and activator of transcription 3, which is activated by cytokines, was also reduced, and expression levels of interferon-stimulated genes, 2’-5’ oligoadenylate synthase, interferon-inducible double-stranded RNA-activated protein kinase, and myxovirus resistance 1 were similarly suppressed. Since LPS binds to toll-like receptor 4 and stimulates the downstream pathway leading to induction of these genes, we examined the extracellular signal-regulated kinase and IκB-α. The phosphorylation levels of these molecules were reduced in transgenic mouse liver, indicating that the pathway upstream of the molecules was disrupted by NS5A. Further analyses revealed that the interaction between interleukin-1 receptor-associated kinase-1 and tumor necrosis factor receptor associated factor-6 was dispersed in transgenic mice, suggesting that NS5A may interfere with this interaction via myeloid differentiation primary response gene 88, which was shown to interact with NS5A. Since the gut microbiota, a source of LPS, is known to be associated with pathological conditions in liver diseases, our results suggest the involvement of NS5A in the pathogenesis of HCV infected-liver via the suppression of innate immunity.
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Affiliation(s)
- Takeya Tsutsumi
- Department of Internal Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
- * E-mail:
| | - Kazuya Okushin
- Department of Internal Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
| | - Kenichiro Enooku
- Department of Internal Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
| | - Hidetaka Fujinaga
- Department of Internal Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
| | - Kyoji Moriya
- Department of Internal Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
| | - Hiroshi Yotsuyanagi
- Department of Internal Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
| | - Hideki Aizaki
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Tetsuro Suzuki
- Department of Infectious Diseases, Hamamatsu University School of Medicine, Hamamatsu, Japan
| | - Yoshiharu Matsuura
- Department of Molecular Virology, Research Institute for Microbial Diseases, Osaka University, Osaka, Japan
| | - Kazuhiko Koike
- Department of Internal Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
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10
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Pinkham C, An S, Lundberg L, Bansal N, Benedict A, Narayanan A, Kehn-Hall K. The role of signal transducer and activator of transcription 3 in Rift Valley fever virus infection. Virology 2016; 496:175-185. [PMID: 27318793 PMCID: PMC4969177 DOI: 10.1016/j.virol.2016.06.004] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/24/2015] [Revised: 05/13/2016] [Accepted: 06/08/2016] [Indexed: 12/27/2022]
Abstract
Rift Valley fever (RVF) is a zoonotic disease that can cause severe illness in humans and livestock, triggering spontaneous abortion in almost 100% of pregnant ruminants. In this study, we demonstrate that signal transducer and activator of transcription 3 (STAT3) is phosphorylated on its conserved tyrosine residue (Y705) following RVFV infection. This phosphorylation was dependent on a major virulence factor, the viral nonstructural protein NSs. Loss of STAT3 had little effect on viral replication, but rather resulted in cells being more susceptible to RVFV-induced cell death. Phosphorylated STAT3 translocated to the nucleus, coinciding with inhibition of fos, jun, and nr4a2 gene expression, and the presence of STAT3 and NSs at the nr4a2 promoter. NSs was found predominantly in the cytoplasm of STAT3 null cells, indicating that STAT3 influences NSs nuclear localization. Collectively, these data demonstrate that STAT3 functions in a pro-survival capacity through modulation of NSs localization.
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Affiliation(s)
- Chelsea Pinkham
- National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, 10650 Pyramid Place, Manassas, VA, United States
| | - Soyeon An
- National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, 10650 Pyramid Place, Manassas, VA, United States
| | - Lindsay Lundberg
- National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, 10650 Pyramid Place, Manassas, VA, United States
| | - Neha Bansal
- National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, 10650 Pyramid Place, Manassas, VA, United States
| | - Ashwini Benedict
- National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, 10650 Pyramid Place, Manassas, VA, United States
| | - Aarthi Narayanan
- National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, 10650 Pyramid Place, Manassas, VA, United States
| | - Kylene Kehn-Hall
- National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, 10650 Pyramid Place, Manassas, VA, United States.
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11
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Vallianou I, Dafou D, Vassilaki N, Mavromara P, Hadzopoulou-Cladaras M. Hepatitis C virus suppresses Hepatocyte Nuclear Factor 4 alpha, a key regulator of hepatocellular carcinoma. Int J Biochem Cell Biol 2016; 78:315-326. [PMID: 27477312 DOI: 10.1016/j.biocel.2016.07.027] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2016] [Revised: 07/20/2016] [Accepted: 07/26/2016] [Indexed: 12/18/2022]
Abstract
Hepatitis C Virus (HCV) infection presents with a disturbed lipid profile and can evolve to hepatic steatosis and hepatocellular carcinoma (HCC). Hepatocyte Nuclear Factor 4 alpha (HNF4α) is the most abundant transcription factor in the liver, a key regulator of hepatic lipid metabolism and a critical determinant of Epithelial to Mesenchymal Transition and hepatic development. We have previously shown that transient inhibition of HNF4α initiates transformation of immortalized hepatocytes through a feedback loop consisting of miR-24, IL6 receptor (IL6R), STAT3, miR-124 and miR-629, suggesting a central role of HNF4α in HCC. However, the role of HNF4α in Hepatitis C Virus (HCV)-related hepatocarcinoma has not been evaluated and remains controversial. In this study, we provide strong evidence suggesting that HCV downregulates HNF4α expression at both transcriptional and translational levels. The observed decrease of HNF4α expression correlated with the downregulation of its downstream targets, HNF1α and MTP. Ectopic overexpression of HCV proteins also exhibited an inhibitory effect on HNF4α levels. The inhibition of HNF4α expression by HCV appeared to be mediated at transcriptional level as HCV proteins suppressed HNF4α gene promoter activity. HCV also up-regulated IL6R, activated STAT3 protein phosphorylation and altered the expression of acute phase genes. Furthermore, as HCV triggered the loss of HNF4α a consequent change of miR-24, miR-629 or miR-124 was observed. Our findings demonstrated that HCV-related HCC could be mediated through HNF4α-microRNA deregulation implying a possible role of HNF4α in HCV hepatocarcinogenesis. HCV inhibition of HNF4α could be sustained to promote HCC.
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Affiliation(s)
- Ioanna Vallianou
- Department of Genetics, Development and Molecular Biology, School of Biology, Aristotle University of Thessaloniki, Thessaloniki, Greece
| | - Dimitra Dafou
- Department of Genetics, Development and Molecular Biology, School of Biology, Aristotle University of Thessaloniki, Thessaloniki, Greece
| | - Niki Vassilaki
- Molecular Virology Laboratory, Hellenic Pasteur Institute, Athens, Greece
| | - Penelope Mavromara
- Molecular Virology Laboratory, Hellenic Pasteur Institute, Athens, Greece
| | - Margarita Hadzopoulou-Cladaras
- Department of Genetics, Development and Molecular Biology, School of Biology, Aristotle University of Thessaloniki, Thessaloniki, Greece.
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12
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Li X, Bhaduri-McIntosh S. A Central Role for STAT3 in Gammaherpesvirus-Life Cycle and -Diseases. Front Microbiol 2016; 7:1052. [PMID: 27458446 PMCID: PMC4937026 DOI: 10.3389/fmicb.2016.01052] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/30/2016] [Accepted: 06/23/2016] [Indexed: 12/14/2022] Open
Abstract
Having co-evolved with humans, herpesviruses have adapted to exploit the host molecular machinery to ensure viral persistence. The cellular protein Signal Transducer and Activator of Transcription 3 (STAT3) is a leading example. STAT3 is a prominent transcription factor that functions in a variety of physiologic processes including embryonic development, inflammation, immunity, and wound healing. Generally activated via growth factor and cytokine signaling, STAT3 can transcriptionally drive oncoproteins, pro-survival and pro-proliferative proteins as well as angiogenic factors, thereby contributing to cancer. As in most non-viral cancers, STAT3 is constitutively active in EBV-related B and epithelial cell cancers and in animal models of KSHV-cancers. Again, similar to non-viral cancers, STAT3 contributes to gammaherpesvirus (EBV and KSHV)-mediated cancers by driving cell proliferation, invasion and angiogenesis. Being herpesviruses, EBV and KSHV establish latency in humans with episodic lytic activation. Importantly, both viruses activate STAT3 almost immediately upon infection of primary cells. In the setting of infection of primary B cells by EBV, this rapidly activated STAT3 plays a key role in suppressing the DNA damage response (DDR) to EBV-oncogene triggered replication stress, thereby facilitating B cell proliferation and ultimately establishment of latency. STAT3 also contributes to maintenance of latency by curbing lytic activation of EBV and KSHV in latent cells that express high levels of STAT3. In this way, gammaherpesviruses exploit STAT3 to overcome cellular anti-proliferative and anti-lytic barriers to promote viral persistence. These investigations into gammaherpesviruses and STAT3 have simultaneously revealed a novel function for STAT3 in suppression of the DDR, a process fundamental to physiologic cell proliferation as well as development of cancer.
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Affiliation(s)
- Xiaofan Li
- Pediatric Infectious Diseases, Department of Pediatrics, Stony Brook University School of Medicine Stony Brook, NY, USA
| | - Sumita Bhaduri-McIntosh
- Pediatric Infectious Diseases, Department of Pediatrics, Stony Brook University School of MedicineStony Brook, NY, USA; Department of Molecular Genetics and Microbiology, Stony Brook University School of MedicineStony Brook, NY, USA
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Klebl BM, Kurtenbach A, Salassidis K, Daub H, Herget T. Host Cell Targets in HCV Therapy: Novel Strategy or Proven Practice? ACTA ACUST UNITED AC 2016; 16:69-90. [PMID: 15889531 DOI: 10.1177/095632020501600201] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
The development of novel antiviral drugs against hepatitis C is a challenging and competitive area of research. Progress of this research has been hampered due to the quasispecies nature of the hepatitis C virus, the absence of cellular infection models and the lack of easily accessible and highly representative animal models. The current combination therapy consisting of interferon-α and ribavirin mainly acts by supporting host cell defence. These therapeutics are the prototypic representatives of indirect antiviral agents as they act on cellular targets. However, the therapy is not a cure, when considered from the long-term perspective, for almost half of the chronically infected patients. This draws attention to the urgent need for more efficient treatments. Novel anti-hepatitis C treatments under study are directed against a number of so-called direct antiviral targets such as polymerases and proteases, which are encoded by the virus. Although such direct antiviral approaches have proven to be successful in several viral indications, there is a risk of resistant viruses developing. In order to avoid resistance, the development of indirect antiviral compounds has to be intensified. These act on host cell targets either by boosting the immune response or by blocking the virus host cell interaction. A particularly interesting approach is the development of inhibitors that interfere with signal transduction, such as protein kinase inhibitors. The purpose of this review is to stress the importance of developing indirect antiviral agents that act on host cell targets. In doing so, a large source of potential targets and mechanisms can be exploited, thus increasing the likelihood of success. Ultimately, combination therapies consisting of drugs against direct and indirect viral targets will most probably provide the solution to fighting and eradicating hepatitis C virus in patients.
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Li M, Wang W, Jin R, Zhang T, Li N, Han Q, Wei P, Liu Z. Differential association of STAT3 and HK-II expression in hepatitis B virus- and hepatitis C virus-related hepatocellular carcinoma. J Med Virol 2016; 88:1552-9. [PMID: 26889748 DOI: 10.1002/jmv.24498] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 02/13/2016] [Indexed: 12/14/2022]
Abstract
STAT3 and hexokinase II (HK-II) are involved in viral infection and carcinogenesis of various cancers including hepatocellular carcinoma (HCC). The roles of STAT3 and HK-II in hepatitis B virus (HBV)- and hepatitis C virus (HCV)-related HCC remain largely unclear. This study examined STAT3 and HK-II expression in HBV- and HCV-related HCC, HBV-related liver fibrosis, and normal control liver by using tissue microarray and immunohistochemical method. Results showed that STAT3 expression in HBV-related HCC, HCV-related HCC, and HBV-related liver fibrosis was significantly higher than in control liver (P < 0.001, P = 0.016, and P = 0.005, respectively) and had no significant differences between these three diseased liver tissues. The HK-II expression in HBV-related HCC was significantly higher than that in HCV-related HCC, HBV-related liver fibrosis, and control liver (P = 0.007, P = 0.029, and P = 0.008, respectively) but had no significant elevation in and no significant differences between HCV-related HCC, HBV-related liver fibrosis, and control liver. The HK-II expression was significantly correlated to STAT3 expression in HBV-related HCC (P = 0.022), but no correlation was observed in HCV-related HCC, HBV-related liver fibrosis, and control liver. In conclusion, STAT3 expression is upregulated in both HBV- and HCV-related HCC, while HK-II is predominantly upregulated and correlated to STAT3 in HBV-related HCC. These differential expression and association may suggest the distinct roles of STAT3 and HK-II in hepatocarcinogenesis of HBV and HCV infection. Studies are needed to confirm the relationship of STAT3 and HK-II and to examine the underlying mechanisms. J. Med. Virol. 88:1552-1559, 2016. © 2016 Wiley Periodicals, Inc.
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Affiliation(s)
- Man Li
- Department of Infectious Diseases, First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, People's Republic of China.,Department of Internal Medicine, The Second Hospital of Xi'an, Xi'an, Shaanxi, People's Republic of China
| | - Weihua Wang
- Department of Pharmacogenomics, The Fourth Military Medical University, Xi'an, Shaanxi, People's Republic of China
| | - Rui Jin
- Department of Radiology, Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, People's Republic of China
| | - Tieying Zhang
- Department of Internal Medicine, The Second Hospital of Xi'an, Xi'an, Shaanxi, People's Republic of China
| | - Na Li
- Department of Infectious Diseases, First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, People's Republic of China
| | - Qunying Han
- Department of Infectious Diseases, First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, People's Republic of China
| | - Ping Wei
- Department of Infectious Diseases, First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, People's Republic of China
| | - Zhengwen Liu
- Department of Infectious Diseases, First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, People's Republic of China
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Shi Q, Hoffman B, Liu Q. PI3K-Akt signaling pathway upregulates hepatitis C virus RNA translation through the activation of SREBPs. Virology 2016; 490:99-108. [PMID: 26855332 DOI: 10.1016/j.virol.2016.01.012] [Citation(s) in RCA: 32] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/19/2016] [Accepted: 01/21/2016] [Indexed: 02/06/2023]
Abstract
Hepatitis C virus (HCV) activates PI3K-Akt signaling to enhance entry and replication. Here, we found that this pathway also increased HCV translation. Knocking down the three Akt isoforms significantly decreased, whereas ectopic expression increased HCV translation. HCV translation upregulation by Akt required their kinase activities because Akt kinase-dead mutants downregulated HCV translation; and was dependent on PI3K activity since it was sensitive to PI3K inhibitor wortmannin. The viral 3'UTR was not involved in translation upregulation by Akt. HCV NS5A increased Akt phosphorylation/activity and HCV translation in the absence of the viral 3'UTR. Sterol regulatory element-binding proteins (SREBPs) were the downstream effectors of the PI3K-Akt pathway in regulating HCV translation because Akt1 and Akt2 activated both SREBP-1 and SREBP-2, whereas Akt3 upregulated SREBP-1. Knocking down SREBPs significantly decreased, while ectopic expression of SREBPs increased HCV translation. Taken together, we showed that the PI3K-Akt signaling pathway positively regulates HCV translation through SREBPs.
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Affiliation(s)
| | - Brett Hoffman
- VIDO-InterVac, Vaccinology and Immunotherapeutics, Canada
| | - Qiang Liu
- VIDO-InterVac, Vaccinology and Immunotherapeutics, Veterinary Microbiology, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.
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Machida K, Feldman DE, Tsukamoto H. TLR4-dependent tumor-initiating stem cell-like cells (TICs) in alcohol-associated hepatocellular carcinogenesis. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2015; 815:131-44. [PMID: 25427905 PMCID: PMC10578031 DOI: 10.1007/978-3-319-09614-8_8] [Citation(s) in RCA: 22] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Alcohol abuse predisposes individuals to the development of hepatocellular carcinoma (HCC) and synergistically heightens the HCC risk in patients infected with hepatitis C virus (HCV). The mechanisms of this synergism have been elusive until our recent demonstration of the obligatory role of ectopically expressed TLR4 in liver tumorigenesis in alcohol-fed HCV Ns5a or Core transgenic mice. CD133+/CD49f+ tumor-initiating stem cell-like cells (TICs) isolated from these models are tumorigenic in a manner dependent on TLR4 and NANOG. TICs' tumor-initiating activity and chemoresistance are causally associated with inhibition of TGF-β tumor suppressor pathway due to NANOG-mediated expression of IGF2BP3 and YAP1. TLR4/NANOG activation causes p53 degradation via phosphorylation of the protective protein NUMB and its dissociation from p53 by the oncoprotein TBC1D15. Nutrient deprivation reduces overexpressed TBC1D15 in TICs via autophagy-mediated degradation, suggesting a possible role of this oncoprotein in linking metabolic reprogramming and self-renewal.
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Affiliation(s)
- Keigo Machida
- Southern California Research Center for ALPD and Cirrhosis, Keck School of Medicine of the University of Southern California, 1333 San Pablo Street, MMR-402, Los Angeles, CA, 90089-9141, USA,
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Georgopoulou U, Dimitriadis A, Foka P, Karamichali E, Mamalaki A. Hepcidin and the iron enigma in HCV infection. Virulence 2014; 5:465-76. [PMID: 24626108 PMCID: PMC4063809 DOI: 10.4161/viru.28508] [Citation(s) in RCA: 32] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
An estimated 30-40% of patients with chronic hepatitis C have elevated serum iron, transferrin saturation, and ferritin levels. Clinical data suggest that iron is a co-morbidity factor for disease progression following HCV infection. Iron is essential for a number of fundamental metabolic processes in cells and organisms. Mammalian iron homeostasis is tightly regulated and this is maintained through the coordinated action of sensory and regulatory networks that modulate the expression of iron-related proteins at the transcriptional and/or posttranscriptional levels. Disturbances of iron homeostasis have been implicated in infectious disease pathogenesis. Viruses, similarly to other pathogens, can escape recognition by the immune system, but they need iron from their host to grow and spread. Hepcidin is a 25-aa peptide, present in human serum and urine and represents the key peptide hormone, which modulates iron homeostasis in the body. It is synthesized predominantly by hepatocytes and its mature form is released in circulation. In this review, we discuss recent advances in the exciting crosstalk of molecular mechanisms and cell signaling pathways by which iron and hepcidin production influences HCV-induced liver disease.
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Affiliation(s)
- Urania Georgopoulou
- Laboratory of Molecular Virology; Hellenic Pasteur Institute; Athens, Greece
| | - Alexios Dimitriadis
- Laboratory of Molecular Biology and Immunobiotechnology; Hellenic Pasteur Institute; Athens, Greece
| | - Pelagia Foka
- Laboratory of Molecular Virology; Hellenic Pasteur Institute; Athens, Greece; Laboratory of Molecular Biology and Immunobiotechnology; Hellenic Pasteur Institute; Athens, Greece
| | - Eirini Karamichali
- Laboratory of Molecular Virology; Hellenic Pasteur Institute; Athens, Greece
| | - Avgi Mamalaki
- Laboratory of Molecular Biology and Immunobiotechnology; Hellenic Pasteur Institute; Athens, Greece
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Stevenson NJ, Bourke NM, Ryan EJ, Binder M, Fanning L, Johnston JA, Hegarty JE, Long A, O'Farrelly C. Hepatitis C virus targets the interferon-α JAK/STAT pathway by promoting proteasomal degradation in immune cells and hepatocytes. FEBS Lett 2013; 587:1571-8. [PMID: 23587486 DOI: 10.1016/j.febslet.2013.03.041] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/06/2013] [Revised: 03/22/2013] [Accepted: 03/28/2013] [Indexed: 12/19/2022]
Abstract
JAK/STAT signalling is essential for anti-viral immunity, making IFN-α an obvious anti-viral therapeutic. However, many HCV+ patients fail treatment, indicating that the virus blocks successful IFN-α signalling. We found that STAT1 and STAT3 proteins, key components of the IFN-α signalling pathway were reduced in immune cells and hepatocytes from HCV infected patients, and upon HCV expression in Huh7 hepatocytes. However, STAT1 and STAT3 mRNA levels were normal. Mechanistic analysis revealed that in the presence of HCV, STAT3 protein was preferentially ubiquitinated, and degradation was blocked by the proteasomal inhibitor MG132. These findings show that HCV inhibits IFN-α responses in a broad spectrum of cells via proteasomal degradation of JAK/STAT pathway components.
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Affiliation(s)
- Nigel J Stevenson
- School of Biochemistry and Immunology, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland.
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Hepatitis C virus and hepatocellular carcinoma. BIOLOGY 2013; 2:304-16. [PMID: 24832662 PMCID: PMC4009856 DOI: 10.3390/biology2010304] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 01/04/2013] [Revised: 01/18/2013] [Accepted: 01/23/2013] [Indexed: 12/28/2022]
Abstract
Hepatitis C virus (HCV), a hepatotropic virus, is a single stranded-positive RNA virus of ~9,600 nt. length belonging to the Flaviviridae family. HCV infection causes acute hepatitis, chronic hepatitis, cirrhosis and hepatocellular carcinoma (HCC). It has been reported that HCV-coding proteins interact with host-cell factors that are involved in cell cycle regulation, transcriptional regulation, cell proliferation and apoptosis. Severe inflammation and advanced liver fibrosis in the liver background are also associated with the incidence of HCV-related HCC. In this review, we discuss the mechanism of hepatocarcinogenesis in HCV-related liver diseases.
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Hepatitis C virus activates Bcl-2 and MMP-2 expression through multiple cellular signaling pathways. J Virol 2012; 86:12531-43. [PMID: 22951829 DOI: 10.1128/jvi.01136-12] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Hepatitis C virus (HCV) infection is associated with numerous liver diseases and causes serious global health problems, but the mechanisms underlying the pathogenesis of HCV infections remain largely unknown. In this study, we demonstrate that signal transducer and activator of transcription 3 (STAT3), matrix metalloproteinase-2 (MMP-2), and B-cell lymphoma 2 (Bcl-2) are significantly stimulated in HCV-infected patients. We further show that HCV activates STAT3, MMP-2, Bcl-2, extracellular regulated protein kinase (ERK), and c-Jun N-terminal kinase (JNK) in infected Huh7.5.1 cells. Functional screening of HCV proteins revealed that nonstructural protein 4B (NS4B) is responsible for the activation of MMP-2 and Bcl-2 by stimulating STAT3 through repression of the suppressor of cytokine signaling 3 (SOCS3). Our results also demonstrate that multiple signaling cascades, including several members of the protein kinase C (PKC) family, JNK, ERK, and STAT3, play critical roles in the activation of MMP-2 and Bcl-2 mediated by NS4B. Further studies revealed that the C-terminal domain (CTD) of NS4B is sufficient for the activation of STAT3, JNK, ERK, MMP-2, and Bcl-2. We also show that amino acids 227 to 250 of NS4B are essential for regulation of STAT3, JNK, ERK, MMP-2, and Bcl-2, and among them, three residues (237L, 239S, and 245L) are crucial for this regulation. Thus, we reveal a novel mechanism underlying HCV pathogenesis in which multiple intracellular signaling cascades are cooperatively involved in the activation of two important cellular factors, MMP-2 and Bcl-2, in response to HCV infection.
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The oncogenic role of NS5A of hepatitis C virus is mediated by up-regulation of survivin gene expression in the hepatocellular cell through p53 and NF-κB pathways. Cell Biol Int 2012; 35:1225-32. [PMID: 21612579 DOI: 10.1042/cbi20110102] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Approx. 4% of patients experiencing chronic infection of human HCV (hepatitis C virus) ultimately develop HCC (hepatocellular carcinoma). The NS5A (non-structural protein 5A) encoded by HCV has been reported to have an oncogenic role during HCV infection, but the precise mechanism remains largely unclear. The aim of this study is to investigate the signal transduction pathways that mediate the role of NS5A in hepatocarcinogenesis. HepG2 cells were transfected with a plasmid expressing HCV NS5A protein. Subsequently, cell proliferation was analysed by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide] assay and cell counting, apoptosis was analysed by Hoechst 33342 staining, and the gene expression profile was identified by microarray and subsequently validated by RT-PCR (reverse transcription-PCR). The protein levels of survivin, p53, NOS2A (nitric oxide synthase 2A), cyclin D1 and NF-κB (nuclear factor κB) were monitored by Western blotting. Our results showed that transfection of HCV NS5A expression plasmid significantly down-regulated the expression of nine genes and up-regulated the expression of ten genes among the 104 genes detectable by the microarray associated with signalling transduction. The increased expression of survivin mRNA and protein, down-regulated p53 protein levels and increased NOS2A, cyclin D1 and NF-κB protein levels were further identified. Our results suggested that HCV NS5A protein can enhance survivin transcription by increasing p53 degradation and stimulating NOS2A expression as well as NF-κB relocation to the nucleus. The functions of survivin in anti-apoptosis and regulation of cell division might mediate the role of NS5A in HCV-induced HCC.
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Downregulation of GRIM-19 is associated with hyperactivation of p-STAT3 in hepatocellular carcinoma. Med Oncol 2012; 29:3046-54. [DOI: 10.1007/s12032-012-0234-8] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2012] [Accepted: 03/28/2012] [Indexed: 11/25/2022]
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Paulino AD, Ubhi K, Rockenstein E, Adame A, Crews L, Letendre S, Ellis R, Everall IP, Grant I, Masliah E. Neurotoxic effects of the HCV core protein are mediated by sustained activation of ERK via TLR2 signaling. J Neurovirol 2011; 17:327-40. [PMID: 21660601 PMCID: PMC3919659 DOI: 10.1007/s13365-011-0039-0] [Citation(s) in RCA: 30] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2011] [Revised: 04/26/2011] [Accepted: 05/18/2011] [Indexed: 12/24/2022]
Abstract
Hepatitis C virus (HCV) infection is a serious problem among those co-infected with human immunodeficiency virus; however, its impact in the central nervous system (CNS) remains unclear. This study aimed to investigate the mechanisms underlying HCV core protein-mediated neurodegeneration. Analysis of human HCV seropositive cases demonstrated widespread damage to neuronal dendritic processes and sustained activation of extracellular signal-related kinase (ERK); analogous pathologies were observed in wild type injected with HCV core protein into the hippocampus. In vitro analysis in neuronal cells exposed to HCV core demonstrated retraction of the neuronal processes in an ERK/Signal Transducer and Activator of Transcription 3 (STAT3)-dependent manner dependent on toll-like receptor 2 (TLR2) signaling activation. These results indicate that HCV core protein neurotoxicity may be mediated by the sustained activation of ERK/STAT3 via TLR2-IRAK1 signaling pathway. These pathways provide novel targets for development of neuroprotective treatments for HCV involvement of the CNS.
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Affiliation(s)
- Amy D. Paulino
- Department of Neurosciences, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0624, USA
| | - Kiren Ubhi
- Department of Neurosciences, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0624, USA
| | - Edward Rockenstein
- Department of Neurosciences, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0624, USA
| | - Anthony Adame
- Department of Neurosciences, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0624, USA
| | - Leslie Crews
- Department of Pathology, University of California San Diego, La Jolla, CA, USA
| | - Scott Letendre
- Department of Medicine, University of California San Diego, La Jolla, CA, USA
| | - Ronald Ellis
- Department of Neurosciences, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0624, USA
| | - Ian P. Everall
- Department of Psychiatry and the HIV Neurobehavioral Research Center, University of California San Diego, La Jolla, CA, USA
| | - Igor Grant
- Department of Psychiatry and the HIV Neurobehavioral Research Center, University of California San Diego, La Jolla, CA, USA; Veterans Affairs Healthcare System, La Jolla, CA, USA
| | - Eliezer Masliah
- Department of Neurosciences, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0624, USA; Department of Pathology, University of California San Diego, La Jolla, CA, USA
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Ishdorj G, Johnston JB, Gibson SB. Inhibition of constitutive activation of STAT3 by curcurbitacin-I (JSI-124) sensitized human B-leukemia cells to apoptosis. Mol Cancer Ther 2011; 9:3302-14. [PMID: 21159613 DOI: 10.1158/1535-7163.mct-10-0550] [Citation(s) in RCA: 45] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Phosphorylation of STAT3 on serine 727 regulates gene expression and is found to be elevated in many B-leukemia cells including chronic lymphocytic leukemia (CLL). It is, however, unclear whether targeting STAT3 will be an effective antileukemia therapy. In this study, we assessed in vitro antileukemia activity of the STAT3 inhibitor JSI-124 (cucurbitacin I). JSI-124 potently induces apoptosis in 3 B-leukemia cell lines (BJAB, I-83, and NALM-6) and in primary CLL cells and was associated with a reduction in serine 727 phosphorylation of STAT3. Similarly, knockdown of STAT3 expression induced apoptosis in these leukemia cells. In addition, we found that JSI-124 and knockdown of STAT3 decreased antiapoptotic protein XIAP expression and overexpression of XIAP blocked JSI-124-induced apoptosis. Furthermore, we found that combined treatment of JSI-124 and TRAIL increased apoptosis associated with an increase in death receptor 4 expression. Besides apoptosis, we found that JSI-124 also induced cell-cycle arrest prior to apoptosis in B-leukemia cells. This corresponded with reduced expression of the cell-cycle regulatory gene, cdc-2. Thus, we present here for the first time that JSI-124 induced suppression of serine 727 phosphorylation of STAT3, leading to apoptosis and cell-cycle arrest through alterations in gene transcription in B-leukemia cells.
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Affiliation(s)
- Ganchimeg Ishdorj
- Departments of Biochemistry and Medical Genetics, Manitoba Institute of Cell Biology, University of Manitoba, Winnipeg, Manitoba, Canada
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25
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Banerjee A, Ray RB, Ray R. Oncogenic potential of hepatitis C virus proteins. Viruses 2010; 2:2108-2133. [PMID: 21994721 PMCID: PMC3185750 DOI: 10.3390/v2092108] [Citation(s) in RCA: 90] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/15/2010] [Revised: 09/23/2010] [Accepted: 09/24/2010] [Indexed: 02/06/2023] Open
Abstract
Chronic hepatitis C virus (HCV) infection is a major risk factor for liver disease progression, and may lead to cirrhosis and hepatocellular carcinoma (HCC). The HCV genome contains a single-stranded positive sense RNA with a cytoplasmic lifecycle. HCV proteins interact with many host-cell factors and are involved in a wide range of activities, including cell cycle regulation, transcriptional regulation, cell proliferation, apoptosis, lipid metabolism, and cell growth promotion. Increasing experimental evidences suggest that HCV contributes to HCC by modulating pathways that may promote malignant transformation of hepatocytes. At least four of the 10 HCV gene products, namely core, NS3, NS5A and NS5B play roles in several potentially oncogenic pathways. Induction of both endoplasmic reticulum (ER) stress and oxidative stress by HCV proteins may also contribute to hepatocyte growth promotion. The current review identifies important functions of the viral proteins connecting HCV infections and potential for development of HCC. However, most of the putative transforming potentials of the HCV proteins have been defined in artificial cellular systems, and need to be established relevant to infection and disease models. The new insight into the mechanisms for HCV mediated disease progression may offer novel therapeutic targets for one of the most devastating human malignancies in the world today.
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Affiliation(s)
- Arup Banerjee
- Department of Internal Medicine, Edward A. Doisy Research Center, 1100 S. Grand Blvd., 8th Floor, St. Louis, MO 63104, USA; E-Mail:
| | - Ratna B. Ray
- Department of Pathology, Edward A. Doisy Research Center, 1100 S. Grand Blvd., 2nd Floor, St. Louis, MO 63104, USA; E-Mail:
| | - Ranjit Ray
- Department of Internal Medicine, Edward A. Doisy Research Center, 1100 S. Grand Blvd., 8th Floor, St. Louis, MO 63104, USA; E-Mail:
- Molecular Microbiology & Immunology, Edward A. Doisy Research Center, 1100 S. Grand Blvd., 8th Floor, St. Louis, MO 63104, USA
- Author to whom correspondence should be addressed; E-Mail: ; Tel.: 1-314- 977-9034; Fax: 1-314-771-3816
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26
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Hiscott J, Lin R. Inhibition of the interferon antiviral response by hepatitis C virus. Expert Rev Clin Immunol 2010; 2:49-58. [PMID: 20477087 DOI: 10.1586/1744666x.2.1.49] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
Hepatitis C virus (HCV) causes acute and chronic hepatitis by targeting the liver hepatocyte for infection and destruction. The standard treatment for chronic HCV infection is pegylated interferon plus ribavirin. Unfortunately, the sustained response rate and associated toxicity with this treatment are far from ideal; more effective and less toxic treatment regimens are needed. With more than 170 million people infected worldwide, there is an unmet medical need for new effective treatments. Recent advances in the understanding of the signaling pathways leading to the host antiviral response to HCV, the mechanisms used by HCV to evade the immune response, the development of cell culture models of HCV infection and the development of small molecule inhibitors of HCV have generated optimism that novel therapeutic approaches to control HCV will soon be available.
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Affiliation(s)
- John Hiscott
- McGill University, Lady Davis Institute for Medical Research - Jewish General Hospital, Department of Microbiology & Immunology, McGill University, Montreal, H3T 1E2, Canada.
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27
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Shimizu S, Takehara T, Hikita H, Kodama T, Miyagi T, Hosui A, Tatsumi T, Ishida H, Noda T, Nagano H, Doki Y, Mori M, Hayashi N. The let-7 family of microRNAs inhibits Bcl-xL expression and potentiates sorafenib-induced apoptosis in human hepatocellular carcinoma. J Hepatol 2010; 52:698-704. [PMID: 20347499 DOI: 10.1016/j.jhep.2009.12.024] [Citation(s) in RCA: 269] [Impact Index Per Article: 17.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/02/2009] [Revised: 11/17/2009] [Accepted: 12/02/2009] [Indexed: 02/06/2023]
Abstract
BACKGROUND & AIMS Bcl-xL, an anti-apoptotic member of the Bcl-2 family, is over-expressed in human hepatocellular carcinoma, conferring a survival advantage to tumour cells. The mechanisms underlying its dysregulation have not been clarified. In the present study, we explored the involvement of microRNAs that act as endogenous sequence-specific suppressors of gene expression. METHODS The expression profiles of microRNAs in Huh7 hepatoma cells and primary human hepatocytes were compared by microarray analysis. The effect of let-7 on Bcl-xL expression was examined by Western blot and a reporter assay. The involvement of let-7 microRNAs in human tissues was analysed by western blot and reverse transcription-PCR. RESULTS Microarray analysis, followed by in silico target prediction, identified let-7 microRNAs as being downregulated in Huh7 hepatoma cells in comparison with primary human hepatocytes, as well as possessing a putative target site in the bcl-xl mRNA. Over-expression of let-7c or let-7g led to a clear decrease of Bcl-xL expression in Huh7 and HepG2 cell lines. Reporter assays revealed direct post-transcriptional regulation involving let-7c or let-7g and the 3'-untranslated region of bcl-xl mRNA. Human hepatocellular carcinoma tissues with low expression of let-7c displayed higher expression of Bcl-xL protein than those with high expression of let-7c, suggesting that low let-7 microRNA expression contributes to Bcl-xL over-expression. Finally, expression of let-7c enhanced apoptosis of hepatoma cells upon exposure to sorafenib, which downregulates expression of another anti-apoptotic Bcl-2 protein, Mcl-1. CONCLUSIONS let-7 microRNAs negatively regulate Bcl-xL expression in human hepatocellular carcinomas and induce apoptosis in cooperation with an anti-cancer drug targeting Mcl-1.
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Affiliation(s)
- Satoshi Shimizu
- Department of Gastroenterology and Hepatology, Osaka University Graduate School of Medicine, 2-2 Yamada-oka, Suita, Osaka 565-0871, Japan
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Inhibition of intrahepatic gamma interferon production by hepatitis C virus nonstructural protein 5A in transgenic mice. J Virol 2009; 83:8463-9. [PMID: 19553305 DOI: 10.1128/jvi.00751-09] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Hepatitis C virus (HCV) utilizes strategies to suppress or evade the host immune response for establishment of persistent infection. We have shown previously that HCV nonstructural protein 5A (NS5A) impairs tumor necrosis factor alpha (TNF-alpha)-mediated apoptosis. In this study, we have examined the immunomodulatory role of HCV NS5A protein in transgenic mouse (NS5A-Tg) liver when mice were challenged with an unrelated hepatotropic adenovirus as a nonspecific stimulus. Hepatotropic adenovirus was introduced intravenously into NS5A-Tg mice and control mice, and virus clearance from liver was compared over a time course of 3 weeks. The differential mRNA expression levels of 84 cytokine-related genes, signal pathway molecules, transcription factors, and cell surface molecules were determined using real-time reverse transcription-PCR array. NS5A-Tg mice failed to clear adenovirus from liver up to 3 weeks postinfection while control mice cleared virus within 1 to 2 weeks. Subsequent study revealed that gamma interferon (IFN-gamma) expression is inhibited at both the mRNA and protein levels in NS5A-Tg mice, and an inverse expression of transcription factors Gata-3 and Tbx21 is observed. However, TNF-alpha mRNA and protein expression were elevated in both NS5A-Tg and control mice. Together, our results suggested that HCV NS5A acts as an immunomodulator by inhibiting IFN-gamma production and may play an important role toward establishment of chronic HCV infection.
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29
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Dionisio N, Garcia-Mediavilla MV, Sanchez-Campos S, Majano PL, Benedicto I, Rosado JA, Salido GM, Gonzalez-Gallego J. Hepatitis C virus NS5A and core proteins induce oxidative stress-mediated calcium signalling alterations in hepatocytes. J Hepatol 2009; 50:872-82. [PMID: 19303156 DOI: 10.1016/j.jhep.2008.12.026] [Citation(s) in RCA: 89] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/25/2008] [Revised: 11/18/2008] [Accepted: 12/09/2008] [Indexed: 02/07/2023]
Abstract
BACKGROUND/AIMS The hepatitis C virus (HCV) structural core and non-structural NS5A proteins induce in liver cells a series of intracellular events, including elevation of reactive oxygen and nitrogen species (ROS/RNS). Since oxidative stress is associated to altered intracellular Ca(2+) homeostasis, we aimed to investigate the effect of these proteins on Ca(2+) mobilization in human hepatocyte-derived transfected cells, and the protective effect of quercetin treatment. METHODS Ca(2+) mobilization and actin reorganization were determined by spectrofluorimetry. Production of ROS/RNS was determined by flow cytometry. RESULTS Cells transfected with NS5A and core proteins showed enhanced ROS/RNS production and resting cytosolic Ca(2+) concentration, and reduced Ca(2+) concentration into the stores. Phenylephrine-evoked Ca(2+) release, Ca(2+) entry and extrusion by the plasma membrane Ca(2+)-ATPase were significantly reduced in transfected cells. Similar effects were observed in cytokine-activated cells. Phenylephrine-evoked actin reorganization was reduced in the presence of core and NS5A proteins. These effects were significantly prevented by quercetin. Altered Ca(2+) mobilization and increased calpain activation were observed in replicon-containing cells. CONCLUSIONS NS5A and core proteins induce oxidative stress-mediated Ca(2+) homeostasis alterations in human hepatocyte-derived cells, which might underlie the effects of both proteins in the pathogenesis of liver disorders associated to HCV infection.
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Affiliation(s)
- Natalia Dionisio
- Department of Physiology (Cell Physiology Research Group), University of Extremadura, Cáceres, Spain
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30
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Arsenic trioxide inhibits hepatitis C virus RNA replication through modulation of the glutathione redox system and oxidative stress. J Virol 2008; 83:2338-48. [PMID: 19109388 DOI: 10.1128/jvi.01840-08] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
Arsenic trioxide (ATO), a therapeutic reagent used for the treatment of acute promyelocytic leukemia, has recently been reported to increase human immunodeficiency virus type 1 infectivity. However, in this study, we have demonstrated that replication of genome-length hepatitis C virus (HCV) RNA (O strain of genotype 1b) was notably inhibited by ATO at submicromolar concentrations without cell toxicity. RNA replication of HCV-JFH1 (genotype 2a) and the release of core protein into the culture supernatants were also inhibited by ATO after the HCV infection. To clarify the mechanism of the anti-HCV activity of ATO, we examined whether or not PML is associated with this anti-HCV activity, since PML is known to be a target of ATO. Interestingly, we observed the cytoplasmic translocation of PML after treatment with ATO. However, ATO still inhibited the HCV RNA replication even in the PML knockdown cells, suggesting that PML is dispensable for the anti-HCV activity of ATO. In contrast, we found that N-acetyl-cysteine, an antioxidant and glutathione precursor, completely and partially eliminated the anti-HCV activity of ATO after 24 h and 72 h of treatment, respectively. In this context, it is worth noting that we found an elevation of intracellular superoxide anion radical, but not hydrogen peroxide, and the depletion of intracellular glutathione in the ATO-treated cells. Taken together, these findings suggest that ATO inhibits the HCV RNA replication through modulation of the glutathione redox system and oxidative stress.
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Abstract
Hepatitis C virus (HCV) infection is frequently associated with the development of hepatocellular carcinoma (HCC), which is one of the male-dominant diseases. Androgen signaling in liver may be related to carcinogenesis. In this study, we investigated whether HCV proteins cross talk with the androgen receptor (AR) signaling pathway for promotion of carcinogenesis. We have demonstrated that HCV core protein alone or in context with other HCV proteins enhances AR-mediated transcriptional activity and further augments in the presence of androgen. Subsequent study suggested that HCV core protein activates STAT3, which in turn enhances AR-mediated transcription. This activity was blocked by a pharmacological inhibitor of the Jak/Stat signaling pathway, AG490. Vascular endothelial growth factor (VEGF) is a target gene of AR in liver and plays an important role in angiogenesis. Therefore, we examined whether HCV infection modulates VEGF expression in hepatocytes. Our results demonstrated that HCV enhances VEGF expression and facilitates tube formation in human coronary microvascular endothelial cells in the presence of AR. Together, our results suggest that HCV core protein acts as a positive regulator in AR signaling, providing further insight into oncogenic potential in the development of HCC in HCV-infected individuals.
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32
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Isolation and gene analysis of interferon alpha-resistant cell clones of the hepatitis C virus subgenome. Virology 2008; 375:424-32. [PMID: 18353417 DOI: 10.1016/j.virol.2008.02.009] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2007] [Revised: 11/27/2007] [Accepted: 02/10/2008] [Indexed: 12/20/2022]
Abstract
Hepatitis C virus (HCV) proteins appear to play an important role in IFN-resistance, but the molecular mechanism remains unclear. To clarify the mechanism in HCV replicon RNA harboring Huh-7 cells (Huh-9-13), we isolated cellular clones with impaired IFNalpha-sensitivity. Huh-9-13 was cultured for approximately 2 months in the presence of IFNalpha, and 4 IFNalpha-resistant cell clones showing significant resistances were obtained. When total RNA from clones was introduced into Huh-7 cells, the transfected cells also exhibited IFNalpha-resistance. Although no common mutations were present, mutations in NS3 and NS5A regions were accumulated. Transactivation of IFNalpha and IFNalpha-stimulated Stat-1 phosphorylation were reduced, and the elimination of HCV replicon RNA from the clones restored the IFNalpha signaling. These results suggest that the mutations in the HCV replicon RNA, at least in part, cause an inhibition of IFN signaling and are important for acquisition of IFNalpha resistance in Huh-9-13.
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Abstract
In recent years, the effects of hepatitis C virus (HCV) proteins on hepatocarcinogenesis have undergone intense investigations. The potentially oncogenic proteins include at least three HCV proteins: core (C) protein, NS3, and NS5A. Several authors indicated relationships between subcellular localization, concentration, a specific molecular form of the proteins (full length, truncated, phosphorylated), the presence of specific domains (the nuclear localization signal homologous to e.g. Bcl-2) and their effects on the mechanisms linked to oncogenesis. The involvement of all the proteins has been described as being in control of the cell cycle, through interactions with key proteins of the process (p53, p21, cyclins, proliferating cell nuclear antigen), transcription factors, proto-oncogenes, growth factors/cytokines and their receptors, and proteins linked to the apoptotic process. Untilnow, the involvement of the core protein of HCV in liver carcinogenesis is the most recognized. One of the most common proteins affected by HCV proteins is the p53 tumor-suppressor protein. The p21/WAF1 gene is a major target of p53, and the effect of HCV proteins on the gene is frequently considered in parallel. The results of studies on the effects of HCV proteins on the apoptotic process are controversial. This work summarizes the information collected thus far in the field of HCV molecular virology and principal intracellular signaling pathways in which HCV oncogenic proteins are involved.
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Affiliation(s)
- Aldona Kasprzak
- Department of Histology and Embryology, Medical University, Poznań, Poland
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Supekova L, Supek F, Lee J, Chen S, Gray N, Pezacki JP, Schlapbach A, Schultz PG. Identification of human kinases involved in hepatitis C virus replication by small interference RNA library screening. J Biol Chem 2007; 283:29-36. [PMID: 17951261 DOI: 10.1074/jbc.m703988200] [Citation(s) in RCA: 87] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022] Open
Abstract
The propagation of the hepatitis C virus (HCV) is a complex process that requires both host and viral proteins. To facilitate identification of host cell factors that are required for HCV replication, we screened a panel of small interference RNAs that preferentially target human protein kinases using an HCV replicon expressing the firefly luciferase gene as a genetic reporter. Small interference RNAs specific for three human kinases, Csk, Jak1, and Vrk1, were identified that reproducibly reduce viral RNA and viral protein levels in HCV replicon-bearing cells. Treatment of replicon cells with a small molecule inhibitor of Csk also resulted in a significant reduction in HCV RNA and proteins, further supporting a role for Csk in HCV replication. The effects of siRNAs targeting eight kinases known to be negatively regulated by Csk were then examined; knock down of one of these kinases, Fyn, resulted in up-regulation of the HCV replicon, suggesting that Csk mediates its effect on HCV replication through Fyn. This conclusion was further corroborated by demonstration that replicon cells treated with Csk inhibitor contained lower levels of the phosphorylated form of Fyn than control cells.
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Affiliation(s)
- Lubica Supekova
- Department of Chemistry, The Scripps Research Institute, La Jolla, California 92037
| | - Frantisek Supek
- Department of Drug Discovery, Genomics Institute of the Novartis Research Foundation, San Diego, California 92121
| | - Jongkook Lee
- Department of Chemistry, The Scripps Research Institute, La Jolla, California 92037
| | - Shawn Chen
- Department of Chemistry, The Scripps Research Institute, La Jolla, California 92037
| | - Nathanael Gray
- Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Dana Farber Cancer Institute, Boston, Massachusetts 02115
| | - John P Pezacki
- Steacie Institute for Molecular Sciences Ottawa, Ontario K1A 0R6, Canada, and
| | - Achim Schlapbach
- Novartis Institute for Biomedical Research, Basel CH-4002, Switzerland
| | - Peter G Schultz
- Department of Chemistry, The Scripps Research Institute, La Jolla, California 92037; Department of Drug Discovery, Genomics Institute of the Novartis Research Foundation, San Diego, California 92121.
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Abstract
Apoptosis is central for the control and elimination of viral infections. In chronic hepatitis C virus (HCV) infection, enhanced hepatocyte apoptosis and upregulation of the death inducing ligands CD95/Fas, TRAIL and TNFα occur. Nevertheless, HCV infection persists in the majority of patients. The impact of apoptosis in chronic HCV infection is not well understood. It may be harmful by triggering liver fibrosis, or essential in interferon (IFN) induced HCV elimination. For virtually all HCV proteins, pro- and anti-apoptotic effects have been described, especially for the core and NS5A protein. To date, it is not known which HCV protein affects apoptosis in vivo and whether the infectious virions act pro- or anti-apoptotic. With the availability of an infectious tissue culture system, we now can address pathophysiologically relevant issues. This review focuses on the effect of HCV infection and different HCV proteins on apoptosis and of the corresponding signaling cascades.
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Affiliation(s)
- Richard Fischer
- Department of Internal Medicine II, University of Freiburg, Hugstetter Strasse 55, D-79106 Freiburg, Germany.
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36
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Abstract
Hepatitis C virus (HCV) is a major cause of chronic liver disease, cirrhosis and hepatocellular carcinoma (HCC). In man, the pathobiological changes associated with HCV infection have been attributed to both the immune system and direct viral cytopathic effects. Until now, the lack of simple culture systems to infect and propagate the virus has hampered progress in understanding the viral life cycle and pathogenesis of HCV infection, including the molecular mechanisms implicated in HCV-induced HCC. This clearly demonstrates the need to develop small animal models for the study of HCV-associated pathogenesis. This review describes and discusses the development of new HCV animal models to study viral infection and investigate the direct effects of viral protein expression on liver disease.
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Affiliation(s)
- Dina Kremsdorf
- INSERM U812, Universite Paris Descartes, CHU Necker, 156, rue de Vaugirard, Paris 75015, France.
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37
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O'Sullivan LA, Liongue C, Lewis RS, Stephenson SEM, Ward AC. Cytokine receptor signaling through the Jak–Stat–Socs pathway in disease. Mol Immunol 2007; 44:2497-506. [PMID: 17208301 DOI: 10.1016/j.molimm.2006.11.025] [Citation(s) in RCA: 230] [Impact Index Per Article: 12.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2006] [Revised: 11/21/2006] [Accepted: 11/22/2006] [Indexed: 12/31/2022]
Abstract
The complexity of multicellular organisms is dependent on systems enabling cells to respond to specific stimuli. Cytokines and their receptors are one such system, whose perturbation can lead to a variety of disease states. This review represents an overview of our current understanding of the cytokine receptors, Janus kinases (Jaks), Signal transducers and activators of transcription (Stats) and Suppressors of cytokine signaling (Socs), focussing on their contribution to diseases of an immune or hematologic nature.
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Affiliation(s)
- Lynda A O'Sullivan
- School of Life & Environmental Sciences, Deakin University, 221 Burwood Highway, Burwood, Victoria 3125, Australia
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38
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Aggarwal BB, Sethi G, Ahn KS, Sandur SK, Pandey MK, Kunnumakkara AB, Sung B, Ichikawa H. Targeting signal-transducer-and-activator-of-transcription-3 for prevention and therapy of cancer: modern target but ancient solution. Ann N Y Acad Sci 2007; 1091:151-69. [PMID: 17341611 DOI: 10.1196/annals.1378.063] [Citation(s) in RCA: 331] [Impact Index Per Article: 18.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Abstract
Recent evidence indicates a convergence of molecular targets for both prevention and therapy of cancer. Signal-transducer-and-activator-of-transcription-3 (STAT3), a member of a family of six different transcription factors, is closely linked with tumorigenesis. Its role in cancer is indicated by numerous avenues of evidence, including the following: STAT3 is constitutively active in tumor cells; STAT3 is activated by growth factors (e.g., EGF, TGF-alpha, IL-6, hepatocyte growth factor) and oncogenic kinases (e.g., Src); STAT3 regulates the expression of genes that mediate proliferation (e.g., c-myc and cyclin D1), suppress apoptosis (e.g., Bcl-x(L) and survivin), or promote angiogenesis (e.g, VEGF); STAT3 activation has been linked with chemoresistance and radioresistance; and chemopreventive agents have been shown to suppress STAT3 activation. Thus inhibitors of STAT3 activation have potential for both prevention and therapy of cancer. Besides small peptides and oligonucleotides, numerous small molecules have been identified as blockers of STAT3 activation, including synthetic molecules (e.g., AG 490, decoy peptides, and oligonucleotides) and plant polyphenols (e.g., curcumin, resveratrol, flavopiridol, indirubin, magnolol, piceatannol, parthenolide, EGCG, and cucurbitacin). This article discusses these aspects of STAT3 in more detail.
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Affiliation(s)
- Bharat B Aggarwal
- Department of Experimental Therapeutics, The University of Texas M. D. Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, TX 77030, USA.
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Stärkel P, Saeger CD, Leclercq I, Horsmans Y. Role of signal transducer and activator of transcription 3 in liver fibrosis progression in chronic hepatitis C-infected patients. J Transl Med 2007; 87:173-81. [PMID: 17318196 DOI: 10.1038/labinvest.3700496] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022] Open
Abstract
In vitro and animal data suggest that hepatitis C virus (HCV) proteins might interfere with signal transducer and activator of transcription 3 (Stat3) signaling. It remains unknown whether Stat3 influences the apoptotic-proliferation balance and how this may relate to liver fibrosis progression in HCV-infected patients. We assessed Stat3 expression and DNA-binding as well as expression of its regulators protein inhibitor of activated Stat 3 (Pias3) and suppressor of cytokine signaling 3 (Socs3) in 65 HCV-infected livers at various stages of fibrosis progression. We then determined the level of expression of the proliferation markers cyclin D1 and proliferating cell nuclear antigen (PCNA) in conjunction with pro- and antiapoptotic markers Bax and Bcl-2 in the same liver samples. With the onset of fibrosis, Stat3 DNA-binding decreased and became almost undetectable in livers with bridging fibrosis or cirrhosis. Stat3 DNA-binding inversely correlated with Pias3 expression and Stat3-Pias3 interaction increased with the progression of fibrosis. Cyclin D1 and PCNA in hepatocytes decreased dramatically during fibrosis progression and levels highly correlated with Stat3 expression. In addition, an antiapoptotic profile due to upregulation of Bcl-2 principally in infiltrating inflammatory cells was observed with progressing fibrosis. In conclusion, fibrosis progression is characterized by a continuous decline in Stat3 DNA-binding activity related to overexpression and progressive interaction of Pias3-Stat3. The decrease in Stat3 activity correlated with reduced hepatocytes proliferation and a positive antiapoptotic balance in infiltrating inflammatory cells that are known mediators of cell damage in HCV.
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Affiliation(s)
- Peter Stärkel
- Department and Laboratory of Gastroenterology, St Luc University Hospital, Université Catholique de Louvain, Brussels, Belgium.
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Yang SF, Wang SN, Wu CF, Yeh YT, Chai CY, Chunag SC, Sheen MC, Lee KT. Altered p-STAT3 (tyr705) expression is associated with histological grading and intratumour microvessel density in hepatocellular carcinoma. J Clin Pathol 2006; 60:642-8. [PMID: 16901975 PMCID: PMC1955084 DOI: 10.1136/jcp.2006.036970] [Citation(s) in RCA: 66] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Abstract
BACKGROUND Constitutive activation of signal transducer and activator of transcription 3 at tyrosine residue 705 (p-STAT3 (tyr705)) has been associated with many types of human cancers. However, its potential roles and biological effects in hepatocellular carcinoma (HCC) are not well established. AIM To explore whether an altered p-STAT3 (tyr705) expression is associated with angiogenesis or proliferation and thereby plays a part in HCC development. METHODS Paraffin-wax-embedded sections from 69 patients with HCC were collected in this study. Using a semiquantitative immunohistochemical staining method, the expression patterns of p-STAT3 (tyr705) in both HCC lesions and the adjacent non-tumorous liver parenchyma were analysed. The results obtained were further correlated with intratumour microvessel density (MVD), Ki-67 expression, clinicopathological parameters and overall survival. RESULTS A strong p-STAT3 (tyr705) nuclear staining was observed in 49.3% of HCC lesions, but was reported only in 5.8% of the adjacent non-tumorous liver parenchyma (p<0.001). The expression of p-STAT3 (tyr705) in HCC lesions was significantly and positively correlated with the intratumour MVD (p = 0.002), but not with Ki-67 expression. No significant correlation of p-STAT3 (tyr705) was found in addition to histological grading (p = 0.019). Multivariate Cox regression analysis showed that p-STAT3 (tyr705) expression was a significant predictor of overall survival for HCC (p = 0.036), although the Kaplan-Meier survival curves showed no significant difference between the high and low p-STAT3 (tyr705) expression subgroups. CONCLUSIONS The results showed that p-STAT3 (tyr705) expression was closely correlated with histological grading and intratumour MVD in HCC. Thus, the potential role of p-STAT3 (tyr705) in HCC development may be through these correlations.
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Affiliation(s)
- Sheau-Fang Yang
- Department of Pathology, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
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Altered p-STAT3 (tyr705) expression is associated with histological grading and intratumour microvessel density in hepatocellular carcinoma. J Clin Pathol 2006. [PMID: 16901975 DOI: 10.1136/jcp.2006.036970.] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/03/2022]
Abstract
BACKGROUND Constitutive activation of signal transducer and activator of transcription 3 at tyrosine residue 705 (p-STAT3 (tyr705)) has been associated with many types of human cancers. However, its potential roles and biological effects in hepatocellular carcinoma (HCC) are not well established. AIM To explore whether an altered p-STAT3 (tyr705) expression is associated with angiogenesis or proliferation and thereby plays a part in HCC development. METHODS Paraffin-wax-embedded sections from 69 patients with HCC were collected in this study. Using a semiquantitative immunohistochemical staining method, the expression patterns of p-STAT3 (tyr705) in both HCC lesions and the adjacent non-tumorous liver parenchyma were analysed. The results obtained were further correlated with intratumour microvessel density (MVD), Ki-67 expression, clinicopathological parameters and overall survival. RESULTS A strong p-STAT3 (tyr705) nuclear staining was observed in 49.3% of HCC lesions, but was reported only in 5.8% of the adjacent non-tumorous liver parenchyma (p<0.001). The expression of p-STAT3 (tyr705) in HCC lesions was significantly and positively correlated with the intratumour MVD (p = 0.002), but not with Ki-67 expression. No significant correlation of p-STAT3 (tyr705) was found in addition to histological grading (p = 0.019). Multivariate Cox regression analysis showed that p-STAT3 (tyr705) expression was a significant predictor of overall survival for HCC (p = 0.036), although the Kaplan-Meier survival curves showed no significant difference between the high and low p-STAT3 (tyr705) expression subgroups. CONCLUSIONS The results showed that p-STAT3 (tyr705) expression was closely correlated with histological grading and intratumour MVD in HCC. Thus, the potential role of p-STAT3 (tyr705) in HCC development may be through these correlations.
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Lee TL, Yeh J, Van Waes C, Chen Z. Epigenetic modification of SOCS-1 differentially regulates STAT3 activation in response to interleukin-6 receptor and epidermal growth factor receptor signaling through JAK and/or MEK in head and neck squamous cell carcinomas. Mol Cancer Ther 2006; 5:8-19. [PMID: 16432158 DOI: 10.1158/1535-7163.mct-05-0069] [Citation(s) in RCA: 99] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Signal transducer and activator of transcription 3 (STAT3) has been reported to be activated by interleukin-6 receptor (IL-6R) or epidermal growth factor receptor (EGFR) in head and neck squamous cell carcinomas (HNSCC), which may have important implications for responsiveness to therapeutics targeted at EGFR, IL-6R, or intermediary kinases. Suppressor of cytokine signaling-1 (SOCS-1) has been implicated recently in the negative regulation of IL-6R/Janus-activated kinase (JAK)-mediated activation of STAT3, suggesting that SOCS-1 could affect alternative activation of STAT3 by EGFR, IL-6R, and associated kinases. We investigated whether epigenetic modification of SOCS-1 affects STAT3 activation in response to IL-6R-, EGFR-, JAK-, or mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MEK)-mediated signal activation. STAT3 was predominantly activated by IL-6R via Jak1/Jak2 in HNSCC lines UMSCC-9 and UMSCC-38 in association with transcriptional silencing of SOCS-1 by hypermethylation. In UMSCC-11A cells with unmethylated SOCS-1, STAT3 activation was regulated by both EGFR and IL-6R via a JAK-independent pathway involving MEK. Pharmacologic inhibitors of JAK and MEK and expression of SOCS-1 following demethylation or transient transfection inhibited STAT3 activation and cell proliferation and induced cell apoptosis in corresponding cell lines. Hypermethylation of SOCS-1 was found in about one-third of human HNSCC tissues, making it a potentially relevant marker for STAT-targeted therapy in HNSCC patients. We conclude that SOCS-1 methylation status can differentially affect STAT3 activation by IL-6R and EGFR through JAK or MEK in different HNSCC and response to pharmacologic antagonists. Identifying the potential factors and the regulatory pathways in STAT3 activation has important implications for the development and selection of molecularly targeted therapy in HNSCC.
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Affiliation(s)
- Tin Lap Lee
- Tumor Biology Section, Head and Neck Surgery Branch, National Institute on Deafness and Other Communication Disorders, NIH, 10/5D55, MSC-1419, Bethesda, MD 20892-1419, USA
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Abstract
Viral infection of mammalian cells rapidly triggers intracellular signalling events leading to interferon alpha/beta production and a cellular antiviral state. This 'host response' is our first line of immune defence against infection as it imposes several barriers to viral replication and spread. Hepatitis C virus (HCV) evades the host response through a complex combination of processes that include signalling interference, effector modulation and continual viral genetic variation. These evasion strategies support persistent infection and the spread of HCV. Defining the molecular mechanisms by which HCV regulates the host response is of crucial importance and may reveal targets for novel therapeutic strategies.
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Affiliation(s)
- Michael Gale
- Department of Microbiology, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, Texas 75390-9048, USA
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N/A. N/A. Shijie Huaren Xiaohua Zazhi 2005; 13:1725-1730. [DOI: 10.11569/wcjd.v13.i14.1725] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
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Street A, Macdonald A, McCormick C, Harris M. Hepatitis C virus NS5A-mediated activation of phosphoinositide 3-kinase results in stabilization of cellular beta-catenin and stimulation of beta-catenin-responsive transcription. J Virol 2005; 79:5006-16. [PMID: 15795286 PMCID: PMC1069556 DOI: 10.1128/jvi.79.8.5006-5016.2005] [Citation(s) in RCA: 119] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/07/2004] [Accepted: 01/03/2005] [Indexed: 01/04/2023] Open
Abstract
The hepatitis C virus (HCV) nonstructural NS5A protein has been shown to bind to and activate phosphoinositide 3-kinase (PI3K), resulting in activation of the downstream effector serine/threonine kinase Akt/protein kinase B. Here we present data pertaining to the effects of NS5A-mediated Akt activation on its downstream targets. Using a recombinant baculovirus to deliver the complete HCV polyprotein to human hepatoma cells in a tetracycline-regulable fashion, we confirm that expression of the complete HCV polyprotein also activates PI3K and Akt. We further show that this results in the inhibition of the Akt substrate Forkhead transcription factor and the stimulation of phosphorylation of a second key Akt substrate, glycogen synthase kinase-3beta (GSK-3beta). Phosphorylation of GSK-3beta results in its inactivation; consistent with this, we show that expression of the HCV polyprotein results in the accumulation of beta-catenin. Finally, we show that levels of beta-catenin-dependent transcription are also elevated in the presence of the HCV polyprotein. Given the prevalence of beta-catenin mutations in many human tumors, especially colon and hepatocellular carcinomas, these data implicate NS5A-mediated PI3K activation as a contributory factor in the increasingly common association between HCV infection and the development of hepatocellular carcinoma.
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Affiliation(s)
- Andrew Street
- School of Biochemistry and Microbiology, University of Leeds, Leeds LS2 9JT, United Kingdom
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