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1
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Yang J, Bai X, Liu G, Li X. A transcriptional regulatory network of HNF4α and HNF1α involved in human diseases and drug metabolism. Drug Metab Rev 2022; 54:361-385. [PMID: 35892182 DOI: 10.1080/03602532.2022.2103146] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/07/2023]
Abstract
HNF4α and HNF1α are core transcription factors involved in the development and progression of a variety of human diseases and drug metabolism. They play critical roles in maintaining the normal growth and function of multiple organs, mainly the liver, and in the metabolism of endogenous and exogenous substances. The twelve isoforms of HNF4α may exhibit different physiological functions, and HNF4α and HNF1α show varying or even opposing effects in different types of diseases, particularly cancer. Additionally, the regulation of CYP450, phase II drug-metabolizing enzymes, and drug transporters is affected by several factors. This article aims to review the role of HNF4α and HNF1α in human diseases and drug metabolism, including their structures and physiological functions, affected diseases, regulated drug metabolism genes, influencing factors, and related mechanisms. We also propose a transcriptional regulatory network of HNF4α and HNF1α that regulates the expression of target genes related to disease and drug metabolism.
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Affiliation(s)
- Jianxin Yang
- Research Center for High Altitude Medicine, Qinghai University Medical College, Xining, China
| | - Xue Bai
- Research Center for High Altitude Medicine, Qinghai University Medical College, Xining, China
| | - Guiqin Liu
- Research Center for High Altitude Medicine, Qinghai University Medical College, Xining, China
| | - Xiangyang Li
- Research Center for High Altitude Medicine, Qinghai University Medical College, Xining, China.,State Key Laboratory of Plateau Ecology and Agriculture, Qinghai University, Xining, China
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2
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Wu Y, Liu L, Zhao Y, Zhao R. Polysaccharides of vinegar-baked radix bupleuri promote the hepatic targeting effect of oxymatrine by regulating the protein expression of HNF4α, Mrp2, and OCT1. JOURNAL OF ETHNOPHARMACOLOGY 2021; 267:113471. [PMID: 33075440 DOI: 10.1016/j.jep.2020.113471] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/04/2020] [Revised: 09/26/2020] [Accepted: 10/11/2020] [Indexed: 06/11/2023]
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE Vinegar-baked Radix Bupleuri (VBRB) is a processed form of Bupleurum chinense DC. As a well-known meridian-guiding drug, it is traditionally used as a component of traditional Chinese medicine formulations indicated for the treatment of liver diseases. However, the liver targeting component in VBRB remains unclear. Therefore, this study aims to explore the efficacy and mechanism of PSS (polysaccharides in Vinegar-baked Radix Bupleuri) in enhancing liver targeting. MATERIALS AND METHODS Drug distribution of OM alone or combined with PSS was investigated in vivo. Relative uptake efficiency (RUE) and relative targeting efficiency (RTE) were calculated to evaluate liver targeting efficiency. The mRNA and protein expression of organic cation transporter 1 (OCT1), multi-drug resistance protein 2 (Mrp2), and hepatocyte nuclear factor 4α (HNF4α) in the liver were determined by q-PCR and Western blot. Then, AZT, the inhibitor of OCT1 and BI6015, the inhibitor of HNF4α were used to investigate regulatory mechanisms involved in the uptake of OM in the cell. At last, the role of PSS in the anti-hepatitis B virus (HBV) was explored on HepG2.2.15. RESULTS PSS increased the AUC of OM in the liver and increase the RUE and RTE in the liver which indicated a liver targeting enhancing effect. The mRNA and protein expression of OCT1 was increased while Mrp2 and HNF4α decreased. PSS could increase the uptake of OM in HepG2 by increasing the protein expression of HNF4α and OCT1, while inhibited Mrp2. Moreover, PSS combined with OM could enhance the anti-HBV effect of OM. CONCLUSION PSS enhanced the liver targeting efficiency and the underlying mechanism related to up-regulating the expression of OCT1 and HNF4α, while down-regulating of Mrp2. These results suggest that PSS may become a potential excipient and provide a new direction for new targeted research.
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Affiliation(s)
- Yayun Wu
- Guangdong Provincial Hospital of Chinese Medicine, The Second Clinical College of Guangzhou University of Chinese Medicine, Guangzhou University of Chinese Medicine, Guangdong, 510120, China
| | - Lijuan Liu
- Guangdong Provincial Hospital of Chinese Medicine, The Second Clinical College of Guangzhou University of Chinese Medicine, Guangzhou University of Chinese Medicine, Guangdong, 510120, China
| | - Ya Zhao
- Guangdong Provincial Hospital of Chinese Medicine, The Second Clinical College of Guangzhou University of Chinese Medicine, Guangzhou University of Chinese Medicine, Guangdong, 510120, China
| | - Ruizhi Zhao
- Guangdong Provincial Hospital of Chinese Medicine, The Second Clinical College of Guangzhou University of Chinese Medicine, Guangzhou University of Chinese Medicine, Guangdong, 510120, China; Guangdong Provincial Key Laboratory of Clinical Research on Traditional Chinese Medicine Syndrome, China.
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3
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Huang WG, Wang J, Liu YJ, Wang HX, Zhou SZ, Chen H, Yang FW, Li Y, Yi Y, He YH. Endoplasmic Reticulum Stress Increases Multidrug-resistance Protein 2 Expression and Mitigates Acute Liver Injury. Curr Mol Med 2020; 20:548-557. [PMID: 31976833 DOI: 10.2174/1566524020666200124102411] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2019] [Revised: 12/30/2019] [Accepted: 01/12/2020] [Indexed: 11/22/2022]
Abstract
BACKGROUND Multidrug-resistance protein (MRP) 2 is a key membrane transporter that is expressed on hepatocytes and regulated by nuclear factor kappa B (NF-κB). Interestingly, endoplasmic reticulum (ER) stress is closely associated with liver injury and the activation of NF-κB signaling. OBJECTIVE Here, we investigated the impact of ER stress on MRP2 expression and the functional involvement of MRP2 in acute liver injury. METHODS ER stress, MRP2 expression, and hepatocyte injury were analyzed in a carbon tetrachloride (CCl4)-induced mouse model of acute liver injury and in a thapsigargin (TG)-induced model of ER stress. RESULTS CCl4 and TG induced significant ER stress, MRP2 protein expression and NF- κB activation in mice and LO2 cells (P < 0.05). Pretreatment with ER stress inhibitor 4- phenyl butyric acid (PBA) significantly mitigated CCl4 and TG-induced ER stress and MRP2 protein expression (P < 0.05). Moreover, pretreatment with pyrrolidine dithiocarbamic acid (PDTC; NF-κB inhibitor) significantly inhibited CCl4-induced NF-κB activation and reduced MRP2 protein expression (1±0.097 vs. 0.623±0.054; P < 0.05). Furthermore, hepatic downregulation of MRP2 expression significantly increased CCl4- induced ER stress, apoptosis, and liver injury. CONCLUSION ER stress enhances intrahepatic MRP2 protein expression by activating NF-κB. This increase in MRP2 expression mitigates ER stress and acute liver injury.
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Affiliation(s)
- Wen-Ge Huang
- Department of Infectious Diseases, the Affiliated Hospital of Zunyi Medical University, Zunyi, 563003, Guizhou, China
| | - Jun Wang
- Department of Infectious Diseases, the Affiliated Hospital of Zunyi Medical University, Zunyi, 563003, Guizhou, China
| | - Yu-Juan Liu
- Department of Infectious Diseases, the Affiliated Hospital of Zunyi Medical University, Zunyi, 563003, Guizhou, China
| | - Hong-Xia Wang
- Department of Infectious Diseases, the Affiliated Hospital of Zunyi Medical University, Zunyi, 563003, Guizhou, China
| | - Si-Zhen Zhou
- Department of Infectious Diseases, the Affiliated Hospital of Zunyi Medical University, Zunyi, 563003, Guizhou, China
| | - Huan Chen
- Department of Infectious Diseases, the Affiliated Hospital of Zunyi Medical University, Zunyi, 563003, Guizhou, China
| | - Fang-Wan Yang
- Department of Infectious Diseases, the Affiliated Hospital of Zunyi Medical University, Zunyi, 563003, Guizhou, China
| | - Ying Li
- Department of Infectious Diseases, the Affiliated Hospital of Zunyi Medical University, Zunyi, 563003, Guizhou, China
| | - Yu Yi
- Department of Infectious Diseases, the Affiliated Hospital of Zunyi Medical University, Zunyi, 563003, Guizhou, China
| | - Yi-Huai He
- Department of Infectious Diseases, the Affiliated Hospital of Zunyi Medical University, Zunyi, 563003, Guizhou, China
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4
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Preya UH, Woo JH, Choi YS, Choi JH. Hepatocyte nuclear factor-1 beta protects endometriotic cells against apoptotic cell death by up-regulating the expression of antiapoptotic genes†. Biol Reprod 2019; 101:686-694. [PMID: 31322170 DOI: 10.1093/biolre/ioz127] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2019] [Revised: 06/07/2019] [Accepted: 07/11/2019] [Indexed: 12/17/2022] Open
Abstract
The overexpression of hepatocyte nuclear factor-1 beta (HNF1β) in endometriotic lesion has been demonstrated. However, the role of HNF1β in endometriosis remains largely unknown. Human endometriotic 12Z cells showed higher level of HNF1β when compared with normal endometrial HES cells. In human endometriotic 12Z cells, HNF1β knockdown increased susceptibility to apoptotic cell death by oxidative stress, while HNF1β overexpression suppressed apoptosis. In addition, HNF1β knockdown and overexpression significantly decreased and increased, respectively, the expression of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)-dependent antiapoptotic genes. Knockdown of the antiapoptotic genes significantly reduced the HNF1β-induced resistance against oxidative stress in 12Z cells. Furthermore, HNF1β regulated the transcriptional activity of NF-κB, and an NF-κB inhibitor suppressed the HNF1β-enhanced NF-κB-dependent antiapoptotic gene expression and the resistance of the 12Z cells against cell death. Taken together, these data suggest that HNF1β overexpression may protect endometriotic cells against oxidative damage by augmenting antiapoptotic gene expression.
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Affiliation(s)
- Umma Hafsa Preya
- College of Pharmacy, Kyung Hee University, Seoul, South Korea.,Department of Life and Nanopharmaceutical Sciences, Kyung Hee University, Seoul, South Korea
| | - Jeong-Hwa Woo
- College of Pharmacy, Kyung Hee University, Seoul, South Korea
| | - Youn Seok Choi
- Department of Obstetrics and Gynecology, School of Medicine, Catholic University of Daegu, Daegu, South Korea
| | - Jung-Hye Choi
- College of Pharmacy, Kyung Hee University, Seoul, South Korea.,Department of Life and Nanopharmaceutical Sciences, Kyung Hee University, Seoul, South Korea
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5
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Yeh MM, Bosch DE, Daoud SS. Role of hepatocyte nuclear factor 4-alpha in gastrointestinal and liver diseases. World J Gastroenterol 2019; 25:4074-4091. [PMID: 31435165 PMCID: PMC6700705 DOI: 10.3748/wjg.v25.i30.4074] [Citation(s) in RCA: 48] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/25/2019] [Revised: 07/15/2019] [Accepted: 07/19/2019] [Indexed: 02/06/2023] Open
Abstract
Hepatocyte nuclear factor 4-alpha (HNF4α) is a highly conserved member of nuclear receptor superfamily of ligand-dependent transcription factors that is expressed in liver and gastrointestinal organs (pancreas, stomach, and intestine). In liver, HNF4α is best known for its role as a master regulator of liver-specific gene expression and essential for adult and fetal liver function. Dysregulation of HNF4α expression has been associated with many human diseases such as ulcerative colitis, colon cancer, maturity-onset diabetes of the young, liver cirrhosis, and hepatocellular carcinoma. However, the precise role of HNF4α in the etiology of these human pathogenesis is not well understood. Limited information is known about the role of HNF4α isoforms in liver and gastrointestinal disease progression. There is, therefore, a critical need to know how disruption of the expression of these isoforms may impact on disease progression and phenotypes. In this review, we will update our current understanding on the role of HNF4α in human liver and gastrointestinal diseases. We further provide additional information on possible use of HNF4α as a target for potential therapeutic approaches.
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Affiliation(s)
- Matthew M Yeh
- Department of Pathology, University of Washington School of Medicine, Seattle, WA 98195, United States
| | - Dustin E Bosch
- Department of Medicine, University of Washington School of Medicine, Seattle, WA 98195, United States
| | - Sayed S Daoud
- Department of Pharmaceutical Sciences, Washington State University Health Sciences, Spokane, WA 99210, United States
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6
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Song Y, Yang X, Shen Y, Wang Y, Xia X, Zhang AM. STAT3 signaling pathway plays importantly genetic and functional roles in HCV infection. Mol Genet Genomic Med 2019; 7:e821. [PMID: 31219249 PMCID: PMC6687657 DOI: 10.1002/mgg3.821] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2019] [Revised: 05/29/2019] [Accepted: 05/31/2019] [Indexed: 12/12/2022] Open
Abstract
Background Hepatitis C virus (HCV) infection is an extensive health problem, which leads to serious liver diseases. Host genetic polymorphisms were associated with HCV infection, progression, and treatment effect of patients. Signal transducers and activators of transcription 3 (STAT3) signaling pathway was important to HCV infection, but no genetic association was studied between STAT3 signaling pathway and HCV infection. Methods To investigate the genetic and functional roles of the STAT3 signaling pathway, we collected 394 HCV patients and 395 normal controls to genotype 25 signal nucleotide polymorphisms (SNPs) of six genes (Interleukin 6 [IL6, OMIM 147620], Interleukin 6 receptor [IL6R, OMIM 147880], Hepatocyte nuclear factor 1 alpha [HNF1A, OMIM 142410], Hepatocyte nuclear factor 4 alpha [HNF4A, OMIM 600281], STAT3 [OMIM 102582], and ATP binding cassette subfamily C member 2 [ABCC2, OMIM 601107]). Then expression level of these genes were analyzed in HCV infected cells with or without IL6 transfection. Results Our results identified that the SNPs in STAT3 signaling pathway were associated with HCV infection or biochemical features of Yunnan HCV patients. Genotype AA of rs4075015 (IL6R) and GG of rs3212172 (HNF4A) increased the risk of HCV infection (p = 0.024 and 0.029), but the genotype AA of rs7553796 (IL6R) played a protective role in HCV infection (p = 0.0008). High‐density lipoprotein cholesterol (HDL‐C) and Glutamyl transpeptidase (GGT) level were associated with genotypes of rs4845617 (IL6R, p = 0.045) and rs1053023 (STAT3, p = 0.034), respectively. Cell assays suggested that IL6 transfection could suppress HCV proliferation. RNA and protein levels of the IL6R, HNF1A, STAT3, and ABCC2 genes significantly increased after HCV infection. Conclusion We identified STAT3 signaling pathway influenced HCV infection and biochemical characteristics of HCV patients through genetic and functional aspects.
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Affiliation(s)
- Yuzhu Song
- Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming, China.,Molecular Medicine Center of Yunnan Province, Kunming, China
| | - Xianyao Yang
- Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming, China
| | - Yunsong Shen
- Molecular Medicine Center of Yunnan Province, Kunming, China
| | - Yiqian Wang
- Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming, China
| | - Xueshan Xia
- Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming, China.,Molecular Medicine Center of Yunnan Province, Kunming, China
| | - A-Mei Zhang
- Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming, China.,Molecular Medicine Center of Yunnan Province, Kunming, China
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7
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Li Z, Liu Q. Hepatitis C virus regulates proprotein convertase subtilisin/kexin type 9 promoter activity. Biochem Biophys Res Commun 2018; 496:1229-1235. [PMID: 29397939 DOI: 10.1016/j.bbrc.2018.01.176] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2018] [Accepted: 01/29/2018] [Indexed: 12/11/2022]
Abstract
Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secretory serine protease mainly expressed in liver. Although PCSK9 has been shown to inhibit hepatitis C virus (HCV) entry and replication, whether HCV regulates PCSK9 transcription has not been well studied. PCSK9 promoter activity is modulated by numerous transcription factors including sterol-regulatory element binding protein (SREBP)-1a, -1c, -2, hepatocyte nuclear factor-1 (HNF-1), and forkhead box O3 (FoxO3). Since they are differently regulated by HCV, we studied the effects of these transcription factors on PCSK9 promoter activity in the context of HCV infection and replication. We demonstrated that PCSK9 promoter activity was up-regulated after HCV infection and in HCV genomic replicon cells. We also studied the effects of HCV proteins on the PCSK9 promoter activity. While HCV structural proteins core, E1, and E2 had no effect, NS2, NS3, NS3-4A, NS5A and NS5B enhanced, and p7 and NS4B decreased PCSK9 promoter activity. Furthermore, we showed that transcription factors SREBP-1c, HNF-1α and specificity protein 1 increased PCSK9 promoter activity in HCV replicon cells, whereas SREBP-1a, HNF-1β and FoxO3 had an inhibitory effect. These results demonstrated the molecular mechanisms of how HCV modulates PCSK9 promoter activity and advanced our understanding on the mutual interactions between HCV and PCSK9.
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Affiliation(s)
- Zhubing Li
- VIDO-InterVac, Vaccinology and Immunotherapeutics, University of Saskatchewan, Saskatoon, Saskatchewan, Canada
| | - Qiang Liu
- VIDO-InterVac, Vaccinology and Immunotherapeutics, Veterinary Microbiology, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.
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8
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Feng L, Liu L, Zhao Y, Zhao R. Saikosaponins A, C and D enhance liver-targeting effects of anticancer drugs by modulating drug transporters. Oncotarget 2017; 8:110092-110102. [PMID: 29299132 PMCID: PMC5746367 DOI: 10.18632/oncotarget.22639] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2017] [Accepted: 10/27/2017] [Indexed: 11/25/2022] Open
Abstract
Vinegar-baked Radix Bupleuri (VBRB) is clinically used to enhance the pharmacological activity of drugs used to treat liver diseases. Our previous study demonstrated that this effect is dependent on increased drug accumulation in the liver; however, the underlying mechanism remains unclear. We hypothesize that VBRB mediated its effects by altering drug transporters. Thus, the present study was designed to determine the effects of VBRB's main components, saikosaponin A, C, and D, on drug transporters. Transporter activity was determined by measuring the intracellular concentration of transporter substrates. Protein and mRNA levels were measured by Western blot and qPCR, respectively. Colchicine was used as the substrate for P-glycoprotein (Pgp) and multidrug resistance protein (MRP) 1, cisplatin was used as the substrate for Mrp2 and organic cation transporters 2 (Oct2), and verapamil and MK571 were used as inhibitors of Pgp and MRP1, respectively. Saikosaponin A, C, and D differentially affected transporter activity. All of the saikosaponins inhibited Pgp activity in Pgp over-expressing HEK293 cells and increased substrate uptake of OCT2 in OCT2 over-expressing HEK293. Saikosaponin C and D inhibited MRP2 activity in HEK293 cells and BRL 3A cell with high MRP2 expression; saikosaponin A increased colchicine accumulation in GSH-stimulated HEK293 cells, but decreased colchicine uptake in HEK293 cells. Saikosaponin D inhibited MRP1 activity in GSH-stimulated HEK293 cells, but marginally affected the uptake of colchicine in HEK293 cells. In conclusion, saikosaponins play a role in VBRB's induced liver targeting effect through affecting drug transporters with a transporter expression amount depending manner.
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Affiliation(s)
- Limin Feng
- Second Affiliated Hospital, Guangzhou University of Chinese Medicine, Guangzhou 510006, China
| | - Lijuan Liu
- Second Affiliated Hospital, Guangzhou University of Chinese Medicine, Guangzhou 510006, China
| | - Ya Zhao
- Second Affiliated Hospital, Guangzhou University of Chinese Medicine, Guangzhou 510006, China
| | - Ruizhi Zhao
- Second Affiliated Hospital, Guangzhou University of Chinese Medicine, Guangzhou 510006, China.,Guangdong Province Key Laboratory of Clinical Research on Traditional Chinese Medicine Syndrome, Guangzhou 510006, China
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9
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Pirro M, Bianconi V, Francisci D, Schiaroli E, Bagaglia F, Sahebkar A, Baldelli F. Hepatitis C virus and proprotein convertase subtilisin/kexin type 9: a detrimental interaction to increase viral infectivity and disrupt lipid metabolism. J Cell Mol Med 2017; 21:3150-3161. [PMID: 28722331 PMCID: PMC5706572 DOI: 10.1111/jcmm.13273] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/04/2017] [Accepted: 05/07/2017] [Indexed: 12/21/2022] Open
Abstract
From viral binding to the hepatocyte surface to extracellular virion release, the replication cycle of the hepatitis C virus (HCV) intersects at various levels with lipid metabolism; this leads to a derangement of the lipid profile and to increased viral infectivity. Accumulating evidence supports the crucial regulatory role of proprotein convertase subtilisin/kexin type 9 (PCSK9) in lipoprotein metabolism. Notably, a complex interaction between HCV and PCSK9 has been documented. Indeed, either increased or reduced circulating PCSK9 levels have been observed in HCV patients; this discrepancy might be related to several confounders, including HCV genotype, human immunodeficiency virus (HIV) coinfection and the ambiguous HCV‐mediated influence on PCSK9 transcription factors. On the other hand, PCSK9 may itself influence HCV infectivity, inasmuch as the expression of different hepatocyte surface entry proteins and receptors is regulated by PCSK9. The aim of this review is to summarize the current evidence about the complex interaction between HCV and liver lipoprotein metabolism, with a specific focus on PCSK9. The underlying assumption of this review is that the interconnections between HCV and PCSK9 may be central to explain viral infectivity.
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Affiliation(s)
- Matteo Pirro
- Unit of Internal Medicine, Department of Medicine, University of Perugia, Perugia, Italy
| | - Vanessa Bianconi
- Unit of Internal Medicine, Department of Medicine, University of Perugia, Perugia, Italy
| | - Daniela Francisci
- Unit of Infectious Diseases, Department of Medicine, University of Perugia, Perugia, Italy
| | - Elisabetta Schiaroli
- Unit of Infectious Diseases, Department of Medicine, University of Perugia, Perugia, Italy
| | - Francesco Bagaglia
- Unit of Internal Medicine, Department of Medicine, University of Perugia, Perugia, Italy
| | - Amirhossein Sahebkar
- Biotechnology Research Center, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Franco Baldelli
- Unit of Infectious Diseases, Department of Medicine, University of Perugia, Perugia, Italy
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10
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Piccolo P, Annunziata P, Soria LR, Attanasio S, Barbato A, Castello R, Carissimo A, Quagliata L, Terracciano LM, Brunetti-Pierri N. Down-regulation of hepatocyte nuclear factor-4α and defective zonation in livers expressing mutant Z α1-antitrypsin. Hepatology 2017; 66:124-135. [PMID: 28295475 DOI: 10.1002/hep.29160] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/18/2016] [Revised: 02/09/2017] [Accepted: 03/10/2017] [Indexed: 12/19/2022]
Abstract
UNLABELLED α1 -Antitrypsin (AAT) deficiency is one of the most common genetic disorders and the liver disease due to the Z mutant of AAT (ATZ) is a prototype of conformational disorder due to protein misfolding with consequent aberrant intermolecular protein aggregation. In the present study, we found that livers of PiZ transgenic mice expressing human ATZ have altered expression of a network of hepatocyte transcriptional factors, including hepatocyte nuclear factor-4α, that is early down-regulated and induces a transcriptional repression of ATZ expression. Reduced hepatocyte nuclear factor-4α was associated with activation of β-catenin, which regulates liver zonation. Livers of PiZ mice and human patients with AAT deficiency were both found to have a severe perturbation of liver zonation. Functionally, PiZ mice showed a severe defect of ureagenesis, as shown by increased baseline ammonia, and reduced urea production and survival after an ammonia challenge. Down-regulation of hepatocyte nuclear factor-4α expression and defective zonation in livers have not been recognized so far as features of the liver disease caused by ATZ and are likely involved in metabolic disturbances and in the increased risk of hepatocellular carcinoma in patients with AAT deficiency. CONCLUSION The findings of this study are consistent with the concept that abnormal AAT protein conformation and intrahepatic accumulation have broad effects on metabolic liver functions. (Hepatology 2017;66:124-135).
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Affiliation(s)
- Pasquale Piccolo
- Telethon Institute of Genetics and Medicine, Pozzuoli, Naples, Italy
| | | | - Leandro R Soria
- Telethon Institute of Genetics and Medicine, Pozzuoli, Naples, Italy
| | - Sergio Attanasio
- Telethon Institute of Genetics and Medicine, Pozzuoli, Naples, Italy
| | - Anna Barbato
- Telethon Institute of Genetics and Medicine, Pozzuoli, Naples, Italy
| | - Raffaele Castello
- Telethon Institute of Genetics and Medicine, Pozzuoli, Naples, Italy
| | | | - Luca Quagliata
- Molecular Pathology Division, Institute of Pathology, University of Basel, Basel, Switzerland
| | - Luigi M Terracciano
- Molecular Pathology Division, Institute of Pathology, University of Basel, Basel, Switzerland
| | - Nicola Brunetti-Pierri
- Telethon Institute of Genetics and Medicine, Pozzuoli, Naples, Italy.,Department of Translational Medicine, Federico II University, Naples, Italy
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11
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Levy G, Habib N, Guzzardi MA, Kitsberg D, Bomze D, Ezra E, Uygun BE, Uygun K, Trippler M, Schlaak JF, Shibolet O, Sklan EH, Cohen M, Timm J, Friedman N, Nahmias Y. Nuclear receptors control pro-viral and antiviral metabolic responses to hepatitis C virus infection. Nat Chem Biol 2016; 12:1037-1045. [PMID: 27723751 DOI: 10.1038/nchembio.2193] [Citation(s) in RCA: 41] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2016] [Accepted: 08/02/2016] [Indexed: 12/11/2022]
Abstract
Viruses lack the basic machinery needed to replicate and therefore must hijack the host's metabolism to propagate. Virus-induced metabolic changes have yet to be systematically studied in the context of host transcriptional regulation, and such studies shoul offer insight into host-pathogen metabolic interplay. In this work we identified hepatitis C virus (HCV)-responsive regulators by coupling system-wide metabolic-flux analysis with targeted perturbation of nuclear receptors in primary human hepatocytes. We found HCV-induced upregulation of glycolysis, ketogenesis and drug metabolism, with glycolysis controlled by activation of HNF4α, ketogenesis by PPARα and FXR, and drug metabolism by PXR. Pharmaceutical inhibition of HNF4α reversed HCV-induced glycolysis, blocking viral replication while increasing apoptosis in infected cells showing virus-induced dependence on glycolysis. In contrast, pharmaceutical inhibition of PPARα or FXR reversed HCV-induced ketogenesis but increased viral replication, demonstrating a novel host antiviral response. Our results show that virus-induced changes to a host's metabolism can be detrimental to its life cycle, thus revealing a biologically complex relationship between virus and host.
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Affiliation(s)
- Gahl Levy
- Grass Center for Bioengineering, Benin School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel.,Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Naomi Habib
- Grass Center for Bioengineering, Benin School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel.,Grass Center for Bioengineering, Benin School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel.,Klarman Cell Observatory, Broad Institute of Harvard and MIT, Cambridge, Massachusetts, USA
| | - Maria Angela Guzzardi
- Grass Center for Bioengineering, Benin School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel.,Institute of Clinical Physiology, National Research Council (CNR), Pisa, Italy
| | - Daniel Kitsberg
- Grass Center for Bioengineering, Benin School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel.,Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - David Bomze
- Grass Center for Bioengineering, Benin School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel.,Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Elishai Ezra
- Grass Center for Bioengineering, Benin School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel.,Faculty of Engineering, Jerusalem College of Technology, Jerusalem, Israel
| | - Basak E Uygun
- Center for Engineering in Medicine, Massachusetts General Hospital, Boston, Massachusetts, USA
| | - Korkut Uygun
- Center for Engineering in Medicine, Massachusetts General Hospital, Boston, Massachusetts, USA
| | - Martin Trippler
- Department of Gastroenterology and Hepatology, University Hospital, University Duisburg-Essen, Essen, Germany
| | - Joerg F Schlaak
- Department of Gastroenterology and Hepatology, University Hospital, University Duisburg-Essen, Essen, Germany
| | - Oren Shibolet
- Liver Unit, Department of Gastroenterology, Tel-Aviv Medical Center and Sackler Faculty of Medicine, Tel Aviv, Israel
| | - Ella H Sklan
- Department of Clinical Microbiology and Immunology, Sackler School of Medicine, Tel Aviv University, Tel Aviv, Israel
| | - Merav Cohen
- Grass Center for Bioengineering, Benin School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel.,Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Joerg Timm
- Institute for Virology, Medical Faculty, University of Düsseldorf, Düsseldorf, Germany
| | - Nir Friedman
- Grass Center for Bioengineering, Benin School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel.,Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Yaakov Nahmias
- Grass Center for Bioengineering, Benin School of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel.,Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
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12
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Crosstalk of HNF4 α with extracellular and intracellular signaling pathways in the regulation of hepatic metabolism of drugs and lipids. Acta Pharm Sin B 2016; 6:393-408. [PMID: 27709008 PMCID: PMC5045537 DOI: 10.1016/j.apsb.2016.07.003] [Citation(s) in RCA: 71] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2016] [Revised: 05/05/2016] [Accepted: 05/11/2016] [Indexed: 12/15/2022] Open
Abstract
The liver is essential for survival due to its critical role in the regulation of metabolic homeostasis. Metabolism of xenobiotics, such as environmental chemicals and drugs by the liver protects us from toxic effects of these xenobiotics, whereas metabolism of cholesterol, bile acids (BAs), lipids, and glucose provide key building blocks and nutrients to promote the growth or maintain the survival of the organism. As a well-established master regulator of liver development and function, hepatocyte nuclear factor 4 alpha (HNF4α) plays a critical role in regulating a large number of key genes essential for the metabolism of xenobiotics, metabolic wastes, and nutrients. The expression and activity of HNF4α is regulated by diverse hormonal and signaling pathways such as growth hormone, glucocorticoids, thyroid hormone, insulin, transforming growth factor-β, estrogen, and cytokines. HNF4α appears to play a central role in orchestrating the transduction of extracellular hormonal signaling and intracellular stress/nutritional signaling onto transcriptional changes in the liver. There have been a few reviews on the regulation of drug metabolism, lipid metabolism, cell proliferation, and inflammation by HNF4α. However, the knowledge on how the expression and transcriptional activity of HNF4α is modulated remains scattered. Herein I provide comprehensive review on the regulation of expression and transcriptional activity of HNF4α, and how HNF4α crosstalks with diverse extracellular and intracellular signaling pathways to regulate genes essential in liver pathophysiology.
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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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14
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Mandai M, Amano Y, Yamaguchi K, Matsumura N, Baba T, Konishi I. Ovarian clear cell carcinoma meets metabolism; HNF-1β confers survival benefits through the Warburg effect and ROS reduction. Oncotarget 2015; 6:30704-14. [PMID: 26375553 PMCID: PMC4741562 DOI: 10.18632/oncotarget.5228] [Citation(s) in RCA: 37] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2015] [Accepted: 08/10/2015] [Indexed: 12/21/2022] Open
Abstract
Ovarian clear cell carcinoma (OCCC) constitutes one of the subtypes of ovarian cancers, but it has unique clinical, histological and biological characteristics, one of which is chemo-resistance. It is also known to develop from endometriotic cyst, a benign ovarian tumor, at relatively high frequency. Recently, it is becoming well known that most of OCCCs express HNF1β, a transcription factor, which is closely associated with the development of liver, pancreas and kidney, as well as occurrence of familial forms of type 2 diabetes. Expression of HNF1β is now regarded as a hallmark of this tumor. Nevertheless, exact biological function of this gene in OCCC has not been clarified. We have shown in previous studies that microenvironment in endometriotic cysts contains severe oxidative stress and OCCC develops under such stressful environment as stress-resistant tumor, which may lead to chemo-resistance. We also showed that increased expression of HNF1β facilitates glucose uptake and glycolysis, which is known as Warburg effect. In the previous issue of this journal, by using comprehensive metabolome analysis, we report that HNF1β actually reduces and protects themselves from internal oxidative stress by dramatically changing cellular metabolism. In this article, we review the relevance and significance of cancer-specific metabolism and how they are associated with biological characteristics of OCCC via expression of HNF1β, along with future clinical implications of targeting cancer-specific metabolism.
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Affiliation(s)
- Masaki Mandai
- Department of Gynecology and Obstetrics, Kyoto University Graduate School of Medicine, Kyoto, Japan
- Department of Obstetrics and Gynecology, Kinki University Faculty of Medicine, Osaka-Sayama, Japan
| | - Yasuaki Amano
- Department of Gynecology and Obstetrics, Kyoto University Graduate School of Medicine, Kyoto, Japan
| | - Ken Yamaguchi
- Department of Gynecology and Obstetrics, Kyoto University Graduate School of Medicine, Kyoto, Japan
| | - Noriomi Matsumura
- Department of Gynecology and Obstetrics, Kyoto University Graduate School of Medicine, Kyoto, Japan
| | - Tsukasa Baba
- Department of Gynecology and Obstetrics, Kyoto University Graduate School of Medicine, Kyoto, Japan
| | - Ikuo Konishi
- Department of Gynecology and Obstetrics, Kyoto University Graduate School of Medicine, Kyoto, Japan
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15
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Ding CB, Zhao Y, Zhang JP, Peng ZG, Song DQ, Jiang JD. A zebrafish model for subgenomic hepatitis C virus replication. Int J Mol Med 2015; 35:791-7. [PMID: 25572289 DOI: 10.3892/ijmm.2015.2063] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2014] [Accepted: 12/15/2014] [Indexed: 12/19/2022] Open
Abstract
Persistent infection with hepatitis C virus (HCV) is a major risk factor in the development of hepatocellular carcinoma. The elucidation of the pathogenesis of HCV-associated liver disease is hampered by the absence of an appropriate small animal model. Zebrafish exhibits high genetic homology to mammals, and is easily manipulated experimentally. In this study, we describe the use of a zebrafish model for the analysis of HCV replication mechanisms. As the 5' untranslated region (UTR), the core protein, the non-structural protein 5B (NS5B) and the 3'UTR are essential for HCV replication, we constructed a HCV sub-replicon gene construct including the 4 gene sequences and the enhanced green fluorescent protein (EGFP) reporter gene; these genes were transcribed through the mouse hepatocyte nuclear factor 4 (mHNF4) promoter. By microinjection of the subgenomic replicon vector into zebrafish larvae, the virus was easily detected by observing EGFP fluorescence in the liver. The positive core and NS5B signals showed positive expression of the HCV gene construct in zebrafish by reverse transcription-polymerase chain reaction (RT-PCR) and western blot analysis. Importantly, the negative strand sequence of the HCV subgenomic RNA was detected by RT-PCR and hybridization in situ, demonstrating that the HCV sub-replicon has positive replication activity. Furthermore, the hybridization signal mainly appeared in the liver region of larvae, as detected by the sense probe of the core protein or NS5B, which confirmed that the sub-replicon amplification occurred in the zebrafish liver. The amplification of the sub-replicon caused alterations in the expression of certain genes, which is similar to HCV infection in human liver cells. To verify the use of this zebrafish model in drug evaluation, two drugs against HCV used in clinical practice, ribavirin and oxymatrine, were tested and these drugs showed significant inhibition of replication of the HCV sub-replicon in the larvae. In conclusion, this zebrafish model of HCV may prove to be a novel and simple in vivo model for the study of the mechanisms of HCV replication and may also prove useful in the disovery of new anti-HCV drugs.
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Affiliation(s)
- Cun-Bao Ding
- Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, P.R. China
| | - Ye Zhao
- Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, P.R. China
| | - Jing-Pu Zhang
- Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, P.R. China
| | - Zong-Gen Peng
- Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, P.R. China
| | - Dan-Qing Song
- Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, P.R. China
| | - Jian-Dong Jiang
- Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, P.R. China
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16
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van der Schoor LWE, Verkade HJ, Kuipers F, Jonker JW. New insights in the biology of ABC transporters ABCC2 and ABCC3: impact on drug disposition. Expert Opin Drug Metab Toxicol 2014; 11:273-93. [PMID: 25380746 DOI: 10.1517/17425255.2015.981152] [Citation(s) in RCA: 51] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
Abstract
INTRODUCTION For the elimination of environmental chemicals and metabolic waste products, the body is equipped with a range of broad specificity transporters that are present in excretory organs as well as in several epithelial blood-tissue barriers. AREAS COVERED ABCC2 and ABCC3 (also known as MRP2 and MRP3) mediate the transport of various conjugated organic anions, including many drugs, toxicants and endogenous compounds. This review focuses on the physiology of these transporters, their roles in drug disposition and how they affect drug sensitivity and toxicity. It also examines how ABCC2 and ABCC3 are coordinately regulated at the transcriptional level by members of the nuclear receptor (NR) family of ligand-modulated transcription factors and how this can be therapeutically exploited. EXPERT OPINION Mutations in both ABCC2 and ABCC3 have been associated with changes in drug disposition, sensitivity and toxicity. A defect in ABCC2 is associated with Dubin-Johnson syndrome, a recessively inherited disorder characterized by conjugated hyperbilirubinemia. Pharmacological manipulation of the activity of these transporters can potentially improve the pharmacokinetics and thus therapeutic activity of substrate drugs but also affect the physiological function of these transporters and consequently ameliorate associated disease states.
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Affiliation(s)
- Lori W E van der Schoor
- University of Groningen, University Medical Center Groningen, Center for Liver, Digestive and Metabolic Diseases, Department of Pediatrics , Hanzeplein 1, 9713 GZ Groningen , The Netherlands
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17
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Allegra S, Cusato J, De Nicolò A, Boglione L, Gatto A, Cariti G, Di Perri G, D'Avolio A. Role of pharmacogenetic in ribavirin outcome prediction and pharmacokinetics in an Italian cohort of HCV-1 and 4 patients. Biomed Pharmacother 2014; 69:47-55. [PMID: 25661337 DOI: 10.1016/j.biopha.2014.10.030] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2014] [Accepted: 10/27/2014] [Indexed: 02/07/2023] Open
Abstract
Ribavirin is phosphorylated by adenosine kinase 1 (AK1) and cytosolic 5'-nucleotidase 2 and it is transported into cells by concentrative nucleoside transporters (CNT) 2/3, coded by SLC28A2/3 genes, and equilibrative nucleoside transporters (ENT) 1/2, coded by SLC29A1/2 genes. We evaluated the association of some polymorphisms of IL28B, SLC28A2/3, SLC29A1, ABCB1, NT5C2, AK1, HNF4α genes and ribavirin treatment outcome and pharmacokinetics after 4weeks of therapy, in a cohort of HCV-1/4 Italian patients. Allelic discrimination was performed by real-time PCR; plasma concentrations were determined at the end of dosing interval (Ctrough) using an HPLC-UV method. Non response was negatively predicted by cryoglobulinemia and IL28B_rs12980275 AA genotype and positively by Metavir score; Metavir score, insulin resistance and SLC28A2_rs1060896 CA/AA and HNF4α_rs1884613 CC genotypes were negative predictive factors of SVR, whereas HCV viral load at baseline and IL28B_rs12980275 AA and rs8099917 TT genotypes positively predicted this outcome; RVR was negatively predicted by insulin resistance and positively by cryoglobulinemia and IL28B_rs12980275 AA genotype; Metavir score and insulin resistance were able to negatively predict EVR, whereas cryoglobulinemia and IL28B_rs12980275 AA genotype positively predicted it; at last, virological relapse was negatively predicted by IL28B_rs8099917 TT and AK1_rs1109374 TT genotypes, insulin resistance was a positive predictor factor. Concerning ribavirin pharmacokinetics, SLC28A2_rs11854488 TT was related to lower Ctrough levels; conversely patients with TC profile of SLC28A3_rs10868138 and SLC29A1_rs760370 GG genotype had higher ribavirin levels. These results might contribute to the clarification of mechanisms causing the individuality in the response to ribavirin containing therapy.
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Affiliation(s)
- Sarah Allegra
- Laboratory of Clinical Pharmacology and Pharmacogenetics(2), Unit of Infectious Diseases, University of Turin, Department of Medical Sciences, Amedeo di Savoia Hospital, Corso Svizzera 164, 10149 Turin, Italy
| | - Jessica Cusato
- Laboratory of Clinical Pharmacology and Pharmacogenetics(2), Unit of Infectious Diseases, University of Turin, Department of Medical Sciences, Amedeo di Savoia Hospital, Corso Svizzera 164, 10149 Turin, Italy.
| | - Amedeo De Nicolò
- Laboratory of Clinical Pharmacology and Pharmacogenetics(2), Unit of Infectious Diseases, University of Turin, Department of Medical Sciences, Amedeo di Savoia Hospital, Corso Svizzera 164, 10149 Turin, Italy
| | - Lucio Boglione
- Laboratory of Clinical Pharmacology and Pharmacogenetics(2), Unit of Infectious Diseases, University of Turin, Department of Medical Sciences, Amedeo di Savoia Hospital, Corso Svizzera 164, 10149 Turin, Italy
| | - Alberto Gatto
- Laboratory of Clinical Pharmacology and Pharmacogenetics(2), Unit of Infectious Diseases, University of Turin, Department of Medical Sciences, Amedeo di Savoia Hospital, Corso Svizzera 164, 10149 Turin, Italy
| | - Giuseppe Cariti
- Laboratory of Clinical Pharmacology and Pharmacogenetics(2), Unit of Infectious Diseases, University of Turin, Department of Medical Sciences, Amedeo di Savoia Hospital, Corso Svizzera 164, 10149 Turin, Italy
| | - Giovanni Di Perri
- Laboratory of Clinical Pharmacology and Pharmacogenetics(2), Unit of Infectious Diseases, University of Turin, Department of Medical Sciences, Amedeo di Savoia Hospital, Corso Svizzera 164, 10149 Turin, Italy
| | - Antonio D'Avolio
- Laboratory of Clinical Pharmacology and Pharmacogenetics(2), Unit of Infectious Diseases, University of Turin, Department of Medical Sciences, Amedeo di Savoia Hospital, Corso Svizzera 164, 10149 Turin, Italy
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18
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Foka P, Karamichali E, Dalagiorgou G, Serti E, Doumba PP, Pissas G, Kakkanas A, Kazazi D, Kochlios E, Gaitanou M, Koskinas J, Georgopoulou U, Mavromara P. Hepatitis C virus modulates lipid regulatory factor Angiopoietin-like 3 gene expression by repressing HNF-1α activity. J Hepatol 2014; 60:30-38. [PMID: 23978712 DOI: 10.1016/j.jhep.2013.08.016] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/16/2013] [Revised: 07/30/2013] [Accepted: 08/10/2013] [Indexed: 01/02/2023]
Abstract
BACKGROUND & AIMS HCV relies on host lipid metabolism to complete its life cycle and HCV core is crucial to this interaction. Liver secreted ANGPTL-3 is an LXR- and HNF-1α-regulated protein, which plays a key role in lipid metabolism by increasing plasma lipids via inhibition of lipase enzymes. Here we aimed to investigate the modulation of ANGPTL-3 by HCV core and identify the molecular mechanisms involved. METHODS qRT-PCR and ELISA were used to assess ANGPTL-3 mRNA and protein levels in HCV patients, the JFH-1 infectious system and liver cell lines. Transfections, chromatin immunoprecipitation and immunofluorescence delineated parts of the molecular mechanisms implicated in the core-mediated regulation of ANGPTL-3 gene expression. RESULTS ANGPTL-3 gene expression was decreased in HCV-infected patients and the JFH-1 infectious system. mRNA and promoter activity levels were down-regulated by core. The response was lost when an HNF-1α element in ANGPTL-3 promoter was mutated, while loss of HNF-1α DNA binding to this site was recorded in the presence of HCV core. HNF-1α mRNA and protein levels were not altered by core. However, trafficking between nucleus and cytoplasm was observed and then blocked by an inhibitor of the HNF-1α-specific kinase Mirk/Dyrk1B. Transactivation of LXR/RXR signalling could not restore core-mediated down-regulation of ANGPTL-3 promoter activity. CONCLUSIONS ANGPTL-3 is negatively regulated by HCV in vivo and in vitro. HCV core represses ANGPTL-3 expression through loss of HNF-1α binding activity and blockage of LXR/RXR transactivation. The putative ensuing increase in serum lipid clearance and uptake by the liver may sustain HCV virus replication and persistence.
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Affiliation(s)
- Pelagia Foka
- Molecular Virology Laboratory, Hellenic Pasteur Institute, Athens, Greece.
| | - Eirini Karamichali
- Molecular Virology Laboratory, Hellenic Pasteur Institute, Athens, Greece
| | | | - Elisavet Serti
- Molecular Virology Laboratory, Hellenic Pasteur Institute, Athens, Greece
| | - Polyxeni P Doumba
- 2nd Department of Internal Medicine, Medical School of Athens, Hippokration Hospital, Athens, Greece
| | - George Pissas
- Molecular Virology Laboratory, Hellenic Pasteur Institute, Athens, Greece
| | | | - Dorothea Kazazi
- Molecular Virology Laboratory, Hellenic Pasteur Institute, Athens, Greece
| | - Emmanouil Kochlios
- Molecular Virology Laboratory, Hellenic Pasteur Institute, Athens, Greece
| | - Maria Gaitanou
- Laboratory of Cellular and Molecular Neurobiology, Hellenic Pasteur Institute, Athens, Greece
| | - John Koskinas
- 2nd Department of Internal Medicine, Medical School of Athens, Hippokration Hospital, Athens, Greece
| | | | - Penelope Mavromara
- Molecular Virology Laboratory, Hellenic Pasteur Institute, Athens, Greece.
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Hepatocyte nuclear factor 4α and downstream secreted phospholipase A2 GXIIB regulate production of infectious hepatitis C virus. J Virol 2013; 88:612-27. [PMID: 24173221 DOI: 10.1128/jvi.02068-13] [Citation(s) in RCA: 33] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
Hepatitis C virus (HCV) is a major cause of chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma in humans. The life cycle of HCV is closely associated with the metabolism of lipids, especially very-low-density lipoprotein (VLDL) in hepatocytes. Hepatocyte nuclear factor 4α (HNF4α), the most abundant transcription factor in the liver, regulates the VLDL secretory pathway. However, the effects of HNF4α on the HCV life cycle are unclear. In this study, we investigated the regulatory effects of HNF4α on HCV assembly and secretion. HCV in HNF4α-deficient hepatocytes showed reduced assembly and secretion but unchanged entry and RNA replication. Bezafibrate, a chemical inhibitor of HNF4α, suppressed HCV assembly and secretion. HNF4α downregulation resulted in rearrangement of cytosolic lipid droplets (LDs), as evidenced by the aggregation of large LDs and distorted cytosolic distribution. Phospholipase A2 GXIIB (PLA2GXIIB), an HNF4α-regulated factor involved in VLDL secretion, was found to be crucial in HCV secretion. PLA2GXIIB expression was upregulated in hepatocytes harboring HCV subgenomic replicons or in HCV-infected hepatocytes. This upregulation was transcriptionally controlled in an HNF4α-dependent manner after HCV infection. Furthermore, PLA2GXIIB combined with microsomal triglyceride transfer protein was found to be responsible for the regulation of HNF4α-induced HCV infectivity. These results suggest that HNF4α and its downstream PLA2GXIIB are important factors affecting the late stage of the HCV life cycle and may serve as potential drug targets for the treatment of HCV infection.
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20
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O'Brien VP, Bokelmann K, Ramírez J, Jobst K, Ratain MJ, Brockmöller J, Tzvetkov MV. Hepatocyte nuclear factor 1 regulates the expression of the organic cation transporter 1 via binding to an evolutionary conserved region in intron 1 of the OCT1 gene. J Pharmacol Exp Ther 2013; 347:181-92. [PMID: 23922447 PMCID: PMC3781413 DOI: 10.1124/jpet.113.206359] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2013] [Accepted: 08/05/2013] [Indexed: 12/15/2022] Open
Abstract
The organic cation transporter 1 (OCT1), also known as solute carrier family 22 member 1, is strongly and specifically expressed in the human liver. Here we show that the hepatocyte nuclear factor 1 (HNF1) regulates OCT1 transcription and contributes to the strong, liver-specific expression of OCT1. Bioinformatic analyses revealed strong conservation of HNF1 binding motifs in an evolutionary conserved region (ECR) in intron 1 of the OCT1 gene. Electrophoretic mobility shift and chromatin immunoprecipitation assays confirmed the specific binding of HNF1 to the intron 1 ECR. In reporter gene assays performed in HepG2 cells, the intron 1 ECR increased SV40 promoter activity by 22-fold and OCT1 promoter activity by 13-fold. The increase was reversed when the HNF1 binding sites in the intron 1 ECR were mutated or the endogenous HNF1α expression was downregulated with small interfering RNA. Following HNF1α overexpression in Huh7 cells, the intron 1 ECR increased SV40 promoter activity by 11-fold and OCT1 promoter activity by 6-fold. Without HNF1α overexpression, the increases were only 3- and 2-fold, respectively. Finally, in human liver samples, high HNF1 expression was significantly correlated with high OCT1 expression (r = 0.48, P = 0.002, n = 40). In conclusion, HNF1 is a strong regulator of OCT1 expression. It remains to be determined whether genetic variants, disease conditions, or drugs that affect HNF1 activity may affect the pharmacokinetics and efficacy of OCT1-transported drugs such as morphine, tropisetron, ondansetron, tramadol, and metformin. Beyond OCT1, this study demonstrates the validity and usefulness of interspecies comparisons in the discovery of functionally relevant genomic sequences.
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Affiliation(s)
- Valerie P O'Brien
- Institute of Clinical Pharmacology, University Medical Center, Georg-August-Universität Göttingen, Germany (V.P.O., K.B., K.J., J.B., M.V.T.); and Department of Medicine, Section of Hematology/Oncology, The University of Chicago, Chicago, Illinois (J.R., M.J.R.)
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21
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Paracha UZ, Fatima K, Alqahtani M, Chaudhary A, Abuzenadah A, Damanhouri G, Qadri I. Oxidative stress and hepatitis C virus. Virol J 2013; 10:251. [PMID: 23923986 PMCID: PMC3751576 DOI: 10.1186/1743-422x-10-251] [Citation(s) in RCA: 107] [Impact Index Per Article: 8.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2013] [Accepted: 07/31/2013] [Indexed: 02/08/2023] Open
Abstract
The disproportionate imbalance between the systemic manifestation of reactive oxygen species and body's ability to detoxify the reactive intermediates is referred to as oxidative stress. Several biological processes as well as infectious agents, physiological or environmental stress, and perturbed antioxidant response can promote oxidative stress. Oxidative stress usually happens when cells are exposed to more electrically charged reactive oxygen species (ROS) such as H2O2 or O2-. The cells' ability to handle such pro-oxidant species is impeded by viral infections particularly within liver that plays an important role in metabolism and detoxification of harmful substances. During liver diseases (such as hepatocellular or cholestatic problems), the produced ROS are involved in transcriptional activation of a large number of cytokines and growth factors, and continued production of ROS and Reactive Nitrogen Species (RNS) feed into the vicious cycle. Many human viruses like HCV are evolved to manipulate this delicate pro- and antioxidant balance; thus generating the sustainable oxidative stress that not only causes hepatic damage but also stimulates the processes to reduce treatment of damage. In this review article, the oxidant and antioxidant pathways that are perturbed by HCV genes are discussed. In the first line of risk, the pathways of lipid metabolism present a clear danger in accumulation of viral induced ROS. Viral infection leads to decrease in cellular concentrations of glutathione (GSH) resulting in oxidation of important components of cells such as proteins, DNA and lipids as well as double strand breakage of DNA. These disorders have the tendency to lead the cells toward cirrhosis and hepatocellular carcinoma in adults due to constant insult. We have highlighted the importance of such pathways and revealed differences in the extent of oxidative stress caused by HCV infection.
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Affiliation(s)
| | - Kaneez Fatima
- IQ Institute of Infection and Immunity, Lahore, Punjab, Pakistan
| | - Mohammad Alqahtani
- Center of Excellence in Genomic Medicine, King Abdul Aziz University, PO Box 80216, Jeddah, 21589, Saudi Arabia
| | - Adeel Chaudhary
- Center of Excellence in Genomic Medicine, King Abdul Aziz University, PO Box 80216, Jeddah, 21589, Saudi Arabia
- King Fahd Medical Research Center, King Abdul Aziz University, PO Box 80216, Jeddah 21589, Saudi Arabia
| | - Adel Abuzenadah
- Faculty of Applied Medical Sciences, King Abdulaziz University, PO Box 80216, Jeddah 21589, Saudi Arabia
- King Fahd Medical Research Center, King Abdul Aziz University, PO Box 80216, Jeddah 21589, Saudi Arabia
| | - Ghazi Damanhouri
- King Fahd Medical Research Center, King Abdul Aziz University, PO Box 80216, Jeddah 21589, Saudi Arabia
| | - Ishtiaq Qadri
- King Fahd Medical Research Center, King Abdul Aziz University, PO Box 80216, Jeddah 21589, Saudi Arabia
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W Rivero C, Rosso N, Gentile E, Cuestas M, Tiribelli C, Oubiña JR, Mathet VL. Dissimilar expression of multidrug resistance mdr1 and bcrp by the replication of hepatitis C virus: role of the nonstructural 5A protein. J Viral Hepat 2013; 20:e127-30. [PMID: 23490381 DOI: 10.1111/jvh.12028] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/26/2012] [Accepted: 10/06/2012] [Indexed: 12/09/2022]
Abstract
Multidrug resistance associated with the overexpression of ATP-dependent binding cassette (ABC) proteins is widely accepted as an important cause of treatment failure in patients with neoplastic or infectious diseases. Some of them play also a pivotal role in detoxification processes. Herein, we investigated the effect of hepatitis C virus (HCV) replication and nonstructural 5A (NS5A) protein on the expression and functional activity of two ABC transport proteins: MDR1 and BCRP. RT-quantitative real-time polymerase chain reaction (qPCR) was carried out for mdr1 and bcrp mRNAs in both Huh7 cells expressing NS5A and Huh7.5 cells containing either full-length- or subgenomic-HCV replicon systems. The functional activity of these pumps was studied by performing a dye efflux assay with DiOC2 and Rhodamine 123. A dose-dependent down-regulation of mdr1 expression was documented in Huh7 cells expressing the NS5A protein, as well as in both replicon systems. In contrast, a significant increase of bcrp expression in both systems was recorded, which were in full agreement with the dye efflux assay results. These results warrant further in vivo studies in HCV patients with cholestasis and/or patients that are refractive to the pharmacotherapy due to the activity of these pumps.
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Affiliation(s)
- C W Rivero
- Facultad de Medicina, Instituto de Investigaciones en Microbiología y Parasitología Médica (IMPAM), UBA-CONICET, UBA, Buenos Aires, Argentina
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23
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Worley MJ, Welch WR, Berkowitz RS, Ng SW. Endometriosis-associated ovarian cancer: a review of pathogenesis. Int J Mol Sci 2013; 14:5367-79. [PMID: 23466883 PMCID: PMC3634491 DOI: 10.3390/ijms14035367] [Citation(s) in RCA: 111] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2013] [Revised: 02/21/2013] [Accepted: 02/26/2013] [Indexed: 12/20/2022] Open
Abstract
Endometriosis is classically defined as the presence of endometrial glands and stroma outside of the endometrial lining and uterine musculature. With an estimated frequency of 5%–10% among women of reproductive age, endometriosis is a common gynecologic disorder. While in itself a benign lesion, endometriosis shares several characteristics with invasive cancer, has been shown to undergo malignant transformation, and has been associated with an increased risk of epithelial ovarian carcinoma (EOC). Numerous epidemiologic studies have shown an increased risk of EOC among women with endometriosis. This is particularly true for women with endometrioid and clear cell ovarian carcinoma. However, the carcinogenic pathways by which endometriosis associated ovarian carcinoma (EAOC) develops remain poorly understood. Current molecular studies have sought to link endometriosis with EAOC through pathways related to oxidative stress, inflammation and hyperestrogenism. In addition, numerous studies have sought to identify an intermediary lesion between endometriosis and EAOC that may allow for the identification of endometriosis at greatest risk for malignant transformation or for the prevention of malignant transformation of this common gynecologic disorder. The objective of the current article is to review the current data regarding the molecular events associated with EAOC development from endometriosis, with a primary focus on malignancies of the endometrioid and clear cell histologic sub-types.
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Affiliation(s)
- Michael J. Worley
- Division of Gynecologic Oncology, Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women’s Hospital, Harvard Medical School, 75 Francis Street, Boston, MA 02115, USA; E-Mails: (M.J.W.J.); (R.S.B.)
| | - William R. Welch
- Department of Pathology, Brigham and Women’s Hospital, Harvard Medical School, 75 Francis Street, Boston, MA 02115, USA; E-Mail:
| | - Ross S. Berkowitz
- Division of Gynecologic Oncology, Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women’s Hospital, Harvard Medical School, 75 Francis Street, Boston, MA 02115, USA; E-Mails: (M.J.W.J.); (R.S.B.)
| | - Shu-Wing Ng
- Division of Gynecologic Oncology, Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women’s Hospital, Harvard Medical School, 75 Francis Street, Boston, MA 02115, USA; E-Mails: (M.J.W.J.); (R.S.B.)
- Author to whom correspondence should be addressed; E-Mail: ; Tel.: +1-617-278-0072; Fax: +1-617-975-0856
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Herraez E, Gonzalez-Sanchez E, Vaquero J, Romero MR, Serrano MA, Marin JJG, Briz O. Cisplatin-induced chemoresistance in colon cancer cells involves FXR-dependent and FXR-independent up-regulation of ABC proteins. Mol Pharm 2012; 9:2565-76. [PMID: 22800197 DOI: 10.1021/mp300178a] [Citation(s) in RCA: 53] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Export pumps often limit the usefulness of anticancer drugs. Here we investigated the effect of cisplatin on the expression of ABC proteins in human colon cancer cells. Short-term incubation of Caco-2 and LS174T cells with cisplatin resulted in up-regulation of several ABC pumps, in particular MRP2 and BCRP. In partially cisplatin-resistant cells (LS174T/R) obtained by long-term exposure to cisplatin, MRP2 and BCRP up-regulation was more marked. This was further enhanced when these cells were cultured under maintained stimulation with cisplatin. The MRP2 promoter (MRP2pr) was cloned, and partially deleted constructs linked to reporter genes were generated. Transfection of LS174T and LS174T/R cells with these constructs revealed the ability of cisplatin to activate MRP2pr. The intensity of this response was dependent on the conserved MRP2pr region. Basal MRP2pr activity was higher in LS174T/R cells, in which the expression of the transcription factors c/EBPβ, HNF1α, HNF3β, and HNF4α, but not PXR, p53, c-Myc, AP1, YB-1, NRF2, and RARα was enhanced. Up-regulation was particularly high for FXR (200-fold) and SHP (50-fold). In LS174T/R cells, GW4064 induced the expression of FGF19, SHP, OSTα/β, but not MRP2 and BCRP, although the sensitivity of these cells to cisplatin was further reduced. In LS174T cells, GW4064-induced chemoresistance was seen only after being transfected with FXR+RXR, when BCRP, but not MRP2, was up-regulated. Protection of LS174T cells against cisplatin was mimicked by transfection with BCRP. In conclusion, in colon cancer cells, cisplatin treatment enhances chemoresistance through FXR-dependent and FXR-independent mechanisms involving the expression of BCRP and MRP2, respectively.
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Affiliation(s)
- Elisa Herraez
- Laboratory of Experimental Hepatology and Drug Targeting (HEVEFARM), Biomedical Research Institute of Salamanca, University of Salamanca, Spain
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25
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Ai D, Chen C, Han S, Ganda A, Murphy AJ, Haeusler R, Thorp E, Accili D, Horton JD, Tall AR. Regulation of hepatic LDL receptors by mTORC1 and PCSK9 in mice. J Clin Invest 2012; 122:1262-70. [PMID: 22426206 DOI: 10.1172/jci61919] [Citation(s) in RCA: 137] [Impact Index Per Article: 10.5] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2011] [Accepted: 02/01/2012] [Indexed: 12/26/2022] Open
Abstract
Individuals with type 2 diabetes have an increased risk of atherosclerosis. One factor underlying this is dyslipidemia, which in hyperinsulinemic subjects with early type 2 diabetes is typically characterized by increased VLDL secretion but normal LDL cholesterol levels, possibly reflecting enhanced catabolism of LDL via hepatic LDLRs. Recent studies have also suggested that hepatic insulin signaling sustains LDLR levels. We therefore sought to elucidate the mechanisms linking hepatic insulin signaling to regulation of LDLR levels. In WT mice, insulin receptor knockdown by shRNA resulted in decreased hepatic mTORC1 signaling and LDLR protein levels. It also led to increased expression of PCSK9, a known post-transcriptional regulator of LDLR expression. Administration of the mTORC1 inhibitor rapamycin caused increased expression of PCSK9, decreased levels of hepatic LDLR protein, and increased levels of VLDL/LDL cholesterol in WT but not Pcsk9-/- mice. Conversely, mice with increased hepatic mTORC1 activity exhibited decreased expression of PCSK9 and increased levels of hepatic LDLR protein levels. Pcsk9 is regulated by the transcription factor HNF1α, and our further detailed analyses suggest that increased mTORC1 activity leads to activation of PKCδ, reduced activity of HNF4α and HNF1α, decreased PCSK9 expression, and ultimately increased hepatic LDLR protein levels, which result in decreased circulating LDL levels. We therefore suggest that PCSK9 inhibition could be an effective way to reduce the adverse side effect of increased LDL levels that is observed in transplant patients taking rapamycin as immunosuppressive therapy.
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Affiliation(s)
- Ding Ai
- Department of Medicine, Columbia University, New York, New York 10032, USA.
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26
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Wang D, Jiang Z, Shen Z, Wang H, Wang B, Shou W, Zheng H, Chu X, Shi J, Huang W. Functional evaluation of genetic and environmental regulators of p450 mRNA levels. PLoS One 2011; 6:e24900. [PMID: 21998633 PMCID: PMC3187744 DOI: 10.1371/journal.pone.0024900] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/23/2011] [Accepted: 08/19/2011] [Indexed: 11/19/2022] Open
Abstract
Variations in the activities of Cytochrome P450s are one of the major factors responsible for inter-individual differences in drug clearance rates, which may cause serious toxicity or inefficacy of therapeutic drugs. Various mRNA level is one of the key factors for different activity of the major P450 genes. Although both genetic and environmental regulators of P450 gene expression have been widely investigated, few studies have evaluated the functional importance of cis- and trans-regulatory factors and environmental factors in the modulation of inter-individual expression variations of the P450 genes. In this study, we measured the mRNA levels of seven major P450 genes (CYP1A1, CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP3A4 and CYP3A5) in 96 liver biopsy samples from Chinese population. Both trans-acting (mRNA levels and non-synonymous SNPs of putative regulator genes) and cis-acting (gene copy number and functional SNPs) factors were investigated to identify the determinants of the expression variations of these seven P450 genes. We found that expression variations of most P450 genes, regulator genes and housekeeping genes were positively correlated at the mRNA level. After partial correlation analysis using ACTB and GAPDH expression to eliminate the effect of global regulators, a UPGMA (Unweighted Pair Group Method with Arithmetic Mean) tree was constructed to reveal the effects of specific regulation networks potentially masked by global regulators. Combined with the functional analysis of regulators, our results suggested that expression variation at the mRNA level was mediated by several factors in a gene-specific manner. Cis-acting genetic variants might play key roles in the expression variation of CYP2D6 and CYP3A5, environmental inducers might play key roles in CYP1A1 and CYP1A2 variation and global regulators might play key roles in CYP2C9 variation. In addition, the functions of regulators that play less important roles in controlling expression variation for each P450 gene were determined.
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Affiliation(s)
- Dazhi Wang
- Rui Jin Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China
- Department of Genetics, Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Center, Shanghai, China
| | - Zhengwen Jiang
- Department of Genetics, Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Center, Shanghai, China
| | - Zhongyang Shen
- Organ Transplant Center, Tianjin First Central Hospital, Tianjin, China
| | - Hui Wang
- Organ Transplant Center, Tianjin First Central Hospital, Tianjin, China
| | - Beilan Wang
- Department of Genetics, Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Center, Shanghai, China
| | - Weihua Shou
- Rui Jin Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China
- Department of Genetics, Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Center, Shanghai, China
| | - Hong Zheng
- Organ Transplant Center, Tianjin First Central Hospital, Tianjin, China
| | - Xun Chu
- Rui Jin Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China
- Department of Genetics, Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Center, Shanghai, China
| | - Jinxiu Shi
- Department of Genetics, Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Center, Shanghai, China
| | - Wei Huang
- Rui Jin Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China
- Department of Genetics, Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Center, Shanghai, China
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Manzoor S, Idrees M, Ashraf J, Mehmood A, Butt S, Fatima K, Akbar H, Rehaman IU, Qadri I. Identification of ionotrophic purinergic receptors in Huh-7 cells and their response towards structural proteins of HCV genotype 3a. Virol J 2011; 8:431. [PMID: 21899776 PMCID: PMC3177911 DOI: 10.1186/1743-422x-8-431] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/26/2011] [Accepted: 09/08/2011] [Indexed: 01/13/2023] Open
Abstract
Hepatitis C virus (HCV) is a major health problem in developing countries including Pakistan. Chronic HCV infection results in progressive liver disease including fibrosis, cirrhosis, insulin resistance and eventually hepatocellular carcinoma (HCC). Ionotrophic purinergic (P2X) receptors are identified to involve in a spectrum of physiological and pathophysiological processes. However, the role of P2X receptors in HCV liver associated diseases still remains to be investigated. The current study was designed to identify the presence of P2X receptors in human liver cells. Furthermore, it investigates the response of P2X receptors towards HCV structural proteins (E1E2). To determine that how many isoforms of P2X receptors are expressed in human liver cells, human hepatoma cell line (Huh-7) was used. Transcripts (mRNA) of five different isoforms of P2X receptors were identified in Huh-7 cells. To examine the gene expression of identified isoforms of P2X receptors in presence of HCV structural proteins E1E2, Huh-7/E1E2 cell line (stably expressing HCV structural proteins E1E2) was used. The results showed significant increase (6.2 fold) in gene expression of P2X4 receptors in Huh-7/E1E2 cells as compared to control Huh-7 cells. The findings of present study confirmed the presence of transcripts of five different isoforms of P2X receptors in human liver cells and suggest that P2X4 receptors could be represented an important component of the purinergic signaling complex in HCV induced liver pathogenesis.
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Affiliation(s)
- Sobia Manzoor
- National Centre of Excellence in Molecular Biology, University of the Punjab, Lahore, Pakistan
- NUST Center of Virology and Immunology (NCVI), National University of Sciences and Technology (NUST), Islamabad 44000, Pakistan
| | - Muhammad Idrees
- National Centre of Excellence in Molecular Biology, University of the Punjab, Lahore, Pakistan
| | - Javed Ashraf
- Margalla Institute of Health Sciences, Quaid-e-Azam Avenue Gulrez Phase-III, Rawalpindi, Pakistan
| | - Azra Mehmood
- NUST Center of Virology and Immunology (NCVI), National University of Sciences and Technology (NUST), Islamabad 44000, Pakistan
| | - Sadia Butt
- National Centre of Excellence in Molecular Biology, University of the Punjab, Lahore, Pakistan
| | - Kaneez Fatima
- NUST Center of Virology and Immunology (NCVI), National University of Sciences and Technology (NUST), Islamabad 44000, Pakistan
| | - Haji Akbar
- National Centre of Excellence in Molecular Biology, University of the Punjab, Lahore, Pakistan
| | - Irshad U Rehaman
- National Centre of Excellence in Molecular Biology, University of the Punjab, Lahore, Pakistan
| | - Ishtiaq Qadri
- NUST Center of Virology and Immunology (NCVI), National University of Sciences and Technology (NUST), Islamabad 44000, Pakistan
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28
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Chandra V, Holla P, Ghosh D, Chakrabarti D, Padigaru M, Jameel S. The hepatitis E virus ORF3 protein regulates the expression of liver-specific genes by modulating localization of hepatocyte nuclear factor 4. PLoS One 2011; 6:e22412. [PMID: 21799848 PMCID: PMC3140526 DOI: 10.1371/journal.pone.0022412] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2011] [Accepted: 06/21/2011] [Indexed: 12/17/2022] Open
Abstract
The hepatitis E virus (HEV) is a small RNA virus and the cause of acute viral hepatitis E. The open reading frame 3 protein (pORF3) of HEV appears to be a pleiotropic regulatory protein that helps in the establishment, propagation and progression of viral infection. However, the global cellular effects of this protein remain to be explored. In the absence of traditional in vitro viral infection systems or efficient replicon systems, we made an adenovirus based ORF3 protein expression system to study its effects on host cell gene expression. We infected Huh7 hepatoma cells with recombinant adenoviruses expressing pORF3 and performed microarray-based gene expression analyses. Several genes down regulated in pORF3-expressing cells were found to be under regulation of the liver-enriched hepatocyte nuclear factor 4 (HNF4), which regulates hepatocyte-specific gene expression. While HNF4 localizes to the nucleus, its phosphorylation results in impaired nuclear localization of HNF4. Here we report that pORF3 increases HNF4 phosphorylation through the ERK and Akt kinases, which results in impaired nuclear translocation of HNF4 and subsequently the down modulation of HNF4-responsive genes in pORF3-expressing cells. We propose that modulation of several hepatocyte specific genes by pORF3 will create an environment favorable for viral replication and pathogenesis.
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Affiliation(s)
- Vivek Chandra
- Virology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi, India
| | - Prasida Holla
- Virology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi, India
| | - Dhrubaa Ghosh
- Virology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi, India
| | | | | | - Shahid Jameel
- Virology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi, India
- * E-mail:
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29
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Steuerwald NM, Parsons JC, Bennett K, Bates TC, Bonkovsky HL. Parallel microRNA and mRNA expression profiling of (genotype 1b) human hepatoma cells expressing hepatitis C virus. Liver Int 2010; 30:1490-504. [PMID: 20825557 DOI: 10.1111/j.1478-3231.2010.02321.x] [Citation(s) in RCA: 30] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/26/2022]
Abstract
BACKGROUND & AIMS MicroRNAs (miRNAs) are members of a class of small noncoding functional RNAs that modulate gene regulation at the post-transcriptional level in a sequence specific manner. miRNA dysfunction has been linked to the pathophysiology of human diseases including those resulting from viral infections. The objective of this study was to investigate changes in miRNA profiles that occur in hepatoma cells expressing hepatitis C virus (HCV) and identify anticorrelated mRNAs, which may be their regulatory targets. METHODS Microarrays were used to perform global miRNA and mRNA expression analysis. Fold changes and pairwise statistics were computed for the resulting datasets. Hierarchical cluster and pathway analyses were performed to assess the degree of differential expression and identify regulatory networks. Bioinformatics tools were used to integrate mRNA profiling results with miRNA target predictions. RESULTS Replication of the Con1 strain of HCV virus in hepatoma cells elicited extensive differential expression of both miRNAs and mRNAs. Forty-three differentially expressed miRNAs (P≤0.001) were identified by microarray analysis in HCV expressing cells. Six thousand eight hundred and fifteen differentially expressed mRNAs (P≤0.05) were identified. Computational analyses revealed anticorrelated miRNA:mRNA pairs for each target prediction algorithm used. Pathway analysis generated a filtered pathway with 120 entities, including seven major regulators and nine major targets potentially under the control of at least 11 miRNAs. CONCLUSIONS The expression of a number of anticorrelated miRNAs:mRNA pairs are affected by the presence of HCV. These miRNAs and their putative targets are attractive candidates for being involved in the pathogenesis and/or progression of HCV-induced chronic hepatitis.
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Affiliation(s)
- Nury M Steuerwald
- The Laboratory for Liver Digestive and Metabolic Disorders, Liver Biliary and Pancreatic Center, Carolinas Medical Center, Cannon Research Center, Charlotte, NC 28203, USA.
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30
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Gu X, Manautou JE. Regulation of hepatic ABCC transporters by xenobiotics and in disease states. Drug Metab Rev 2010; 42:482-538. [PMID: 20233023 DOI: 10.3109/03602531003654915] [Citation(s) in RCA: 51] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023]
Abstract
The subfamily of ABCC transporters consists of 13 members in mammals, including the multidrug resistance-associated proteins (MRPs), sulfonylurea receptors (SURs), and the cystic fibrosis transmembrane conductance regulator (CFTR). These proteins play roles in chemical detoxification, disposition, and normal cell physiology. ABCC transporters are expressed differentially in the liver and are regulated at the transcription and translation level. Their expression and function are also controlled by post-translational modification and membrane-trafficking events. These processes are tightly regulated. Information about alterations in the expression of hepatobiliary ABCC transporters could provide important insights into the pathogenesis of diseases and disposition of xenobiotics. In this review, we describe the regulation of hepatic ABCC transporters in humans and rodents by a variety of xenobiotics, under disease states and in genetically modified animal models deficient in transcription factors, transporters, and cell-signaling molecules.
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Affiliation(s)
- Xinsheng Gu
- Department of Pharmaceutical Sciences, University of Connecticut, Storrs, 06269, USA
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31
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de Boussac H, Ratajewski M, Sachrajda I, Köblös G, Tordai A, Pulaski L, Buday L, Váradi A, Arányi T. The ERK1/2-hepatocyte nuclear factor 4alpha axis regulates human ABCC6 gene expression in hepatocytes. J Biol Chem 2010; 285:22800-8. [PMID: 20463007 DOI: 10.1074/jbc.m110.105593] [Citation(s) in RCA: 33] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2023] Open
Abstract
ABCC6 mutations are responsible for the development of pseudoxanthoma elasticum, a rare recessive disease characterized by calcification of elastic fibers. Although ABCC6 is mainly expressed in the liver the disease has dermatologic, ocular, and cardiovascular symptoms. We investigated the transcriptional regulation of the gene and observed that hepatocyte growth factor (HGF) inhibits its expression in HepG2 cells via the activation of ERK1/2. Similarly, other factors activating the cascade also inhibited ABCC6 expression. We identified the ERK1/2 response element in the proximal promoter by luciferase reporter gene assays. This site overlapped with a region conferring the tissue-specific expression pattern to the gene and with a putative hepatocyte nuclear factor 4alpha (HNF4alpha) binding site. We demonstrated that HNF4alpha regulates the expression of ABCC6, acts through the putative binding site, and determines its cell type-specific expression. We also showed that HNF4alpha is inhibited by the activation of the ERK1/2 cascade. In conclusion we describe here the first regulatory pathway of ABCC6 expression showing that the ERK1/2-HNF4alpha axis has an important role in regulation of the gene.
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Affiliation(s)
- Hugues de Boussac
- Institute of Enzymology, Hungarian Academy of Sciences, Karolina ut 29, 1113 Budapest, Hungary
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32
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Safi SZ, Badshah Y, Waheed Y, Fatima K, Tahir S, Shinwari A, Qadri I. Distribution of hepatitis C virus genotypes, hepatic steatosis and their correlation with clinical and virological factors in Pakistan. ASIAN BIOMED 2010; 4:253-262. [DOI: 10.2478/abm-2010-0032] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Abstract
Background: Due to the inherently unstable nature of HCV, various genotypes have been identified. Steatosis is a histological feature in the progression of HCV-associated liver disease and has been shown to alter the host lipid metabolism. Objective: Assess the distribution of HCV genotypes in the two provinces of Pakistan, and determine the association of hepatic steatosis with altered clinical and virological factors in chronic HCV patients. Methods: One hundred twenty six chronic HCV patients (steatosis in 49 patients) were enrolled for qualitative analysis by PCR. Out of 126 ELISA and PCR positive samples, 119 (48 with hepatic steatosis) chronic HCV patients (mean age 42.0±13.3 years, mean body mass index (BMI) 24.2±4.1) were proved positive after PCR-based detection. Biochemical and virological factors such as HCV genotype, or glucose, in 119 CHC patients were determined and compared between patients with and without hepatic steatosis. Results: Out of 126 samples, 119 were HCV positive, where 58 (48.7%) were genotype 3a, 24 (20.2%) were 3b, 12 (10.1%) were 1a, eight (6.7%) were 2a, six (5.0%) were 1b, and one (0.8%) was 4. Furthermore, seven (5.9%) had a co-infection and three (2.5%) were untypable. BMI (p=0.004), genotype 3a (p<0.001), and triglycerides (p=0.002) were significantly associated with steatosis. It is noteworthy that cholesterol (p=0.281), glucose (p=0.305), lowdensity lipoprotein (p=0.101), high-density lipoprotein (p=0.129), alanine amino transferase (p=0.099), aspartate transaminase (p=0.177), bilirubin (p= 0.882), and age (p=0.846) showed non-significant association. Conclusion: Genotype 3a is the predominant genotype in Pakistan. Hepatic steatosis is quite frequent feature in HCV patients and strongly correlates with BMI, genotype 3a, and triglyceride contents in patients infected with HCV.
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Affiliation(s)
- Sher Zaman Safi
- NUST Center of Virology and Immunology (NCVI), National University of Sciences & Technology (NUST), Islamabad 44000, Pakistan
| | - Yasmin Badshah
- NUST Center of Virology and Immunology (NCVI), National University of Sciences & Technology (NUST), Islamabad 44000, Pakistan
| | - Yasir Waheed
- NUST Center of Virology and Immunology (NCVI), National University of Sciences & Technology (NUST), Islamabad 44000, Pakistan
| | - Kaneez Fatima
- NUST Center of Virology and Immunology (NCVI), National University of Sciences & Technology (NUST), Islamabad 44000, Pakistan
| | - Sadia Tahir
- NUST Center of Virology and Immunology (NCVI), National University of Sciences & Technology (NUST), Islamabad 44000, Pakistan
| | - Alamgir Shinwari
- NUST Center of Virology and Immunology (NCVI), National University of Sciences & Technology (NUST), Islamabad 44000, Pakistan
| | - Ishtiaq Qadri
- NUST Center of Virology and Immunology (NCVI), National University of Sciences & Technology (NUST), Islamabad 44000, Pakistan
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Mirandola S, Bowman D, Hussain MM, Alberti A. Hepatic steatosis in hepatitis C is a storage disease due to HCV interaction with microsomal triglyceride transfer protein (MTP). Nutr Metab (Lond) 2010; 7:13. [PMID: 20178560 PMCID: PMC2838899 DOI: 10.1186/1743-7075-7-13] [Citation(s) in RCA: 43] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/25/2009] [Accepted: 02/23/2010] [Indexed: 02/08/2023] Open
Abstract
Liver steatosis is a frequent histological feature in patients chronically infected with hepatitis C virus (HCV). The relationship between HCV and hepatic steatosis seems to be the result of both epigenetic and genetic factors. In vivo and in vitro studies have shown that HCV can alter intrahepatic lipid metabolism by affecting lipid synthesis, oxidative stress, lipid peroxidation, insulin resistance and the assembly and secretion of VLDL. Many studies suggest that HCV-related steatosis might be the result of a direct interaction between the virus and MTP. It has been demonstrated that MTP is critical for the secretion of HCV particles and that inhibition of its lipid transfer activity reduces HCV production. However, higher degrees of hepatic steatosis were found in chronic hepatitis C patients carrying the T allele of MTP -493G/T polymorphism that seems to be associated with increased MTP transcription. We propose here that liver steatosis in hepatitis C could be a storage disease induced by the effects of the virus and of its proteins on the intracellular lipid machinery and on MTP. Available data support the hypothesis that HCV may modulate MTP expression and activity through a number of mechanisms such as inhibition of its activity and transcriptional control. Initial up regulation could favour propagation of HCV while down regulation in chronic phase could cause impairment of triglyceride secretion and excessive lipid accumulation, with abnormal lipid droplets facilitating the "storage" of virus particles for persistent infection.
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Zhang WX, Chen B, Chen H, Cai Q, Cai WM. Co-regulation of mRNA level of UDP glucuronosyltransferase 1A9 and multi-drug resistance protein 2 in Chinese human liver. Clin Chim Acta 2010; 411:119-21. [PMID: 19833113 DOI: 10.1016/j.cca.2009.10.002] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/03/2009] [Revised: 10/05/2009] [Accepted: 10/06/2009] [Indexed: 11/22/2022]
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Rahman SM, Qadri I, Janssen RC, Friedman JE. Fenofibrate and PBA prevent fatty acid-induced loss of adiponectin receptor and pAMPK in human hepatoma cells and in hepatitis C virus-induced steatosis. J Lipid Res 2009; 50:2193-2202. [PMID: 19502591 PMCID: PMC2759825 DOI: 10.1194/jlr.m800633-jlr200] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2008] [Revised: 04/29/2009] [Indexed: 12/24/2022] Open
Abstract
Adiponectin receptors play a key role in steatosis and inflammation; however, very little is known about regulation of adiponectin receptors in liver. Here, we examined the effects of palmitate loading, endoplasmic reticulum (ER) stress, and the hypolipidemic agent fenofibrate on adiponectin receptor R2 (AdipoR2) levels and AMP-activated protein kinase (AMPK) in human hepatoma Huh7 cells and in Huh.8 cells, a model of hepatitis C-induced steatosis. Palmitate treatment reduced AdipoR2 protein and basal AMPK phosphorylation in Huh7 cells. Fenofibrate treatment preserved AdipoR2 and phosphorylated AMPK (pAMPK) levels in palmitate-treated cells accompanied by reduced triglyceride (TG) accumulation and less activation of ER stress markers CCAAT/enhancer binding (C/EBPbeta) and eukaryotic translation initiation factor 2 alpha. ER stress agents thapsigargin and tunicamycin suppressed AdipoR2 and pAMPK levels in Huh7 cells, while fenofibrate and the chemical chaperone 4-phenylbutyrate (PBA) prevented these changes. AdipoR2 levels were lower in Huh.8 cells and fenofibrate treatment increased AdipoR2 while reducing activation of c-Jun N-terminal kinase and C/EBPbeta expression without changing TG levels. Taken together, these results suggest that fatty acids and ER stress reduce AdipoR2 protein and pAMPK levels, while fenofibrate and PBA might be important therapeutic agents to correct lipid- and ER stress-mediated loss of AdipoR2 and pAMPK associated with nonalcoholic steatohepatitis.
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Fardel O, Le Vée M. Regulation of human hepatic drug transporter expression by pro-inflammatory cytokines. Expert Opin Drug Metab Toxicol 2009; 5:1469-81. [DOI: 10.1517/17425250903304056] [Citation(s) in RCA: 65] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
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Qadri I, Hu LJ, Iwahashi M, Al-Zuabi S, Quattrochi LC, Simon FR. Interaction of hepatocyte nuclear factors in transcriptional regulation of tissue specific hormonal expression of human multidrug resistance-associated protein 2 (abcc2). Toxicol Appl Pharmacol 2009; 234:281-292. [PMID: 19010343 DOI: 10.1016/j.taap.2008.10.005] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/22/2008] [Revised: 10/09/2008] [Accepted: 10/17/2008] [Indexed: 01/13/2023]
Abstract
Multidrug resistance-associated protein 2 (MRP2) (ABCC2) is an ATP-binding cassette membrane protein located primarily on apical surface of hepatocytes that mediates transport of conjugated xenobiotics and endogenous compounds into bile. MRP2 is highly expressed in hepatocytes, and at lower levels in small intestines, stomach and kidney. Previous reports have characterized mammalian MRP2 promoters, but none have established the molecular mechanism(s) involved in liver enriched expression. This study aims to investigate the mechanism of hepatic MRP2 regulation. A 2130 bp of MRP2 promoter was cloned from PAC-1 clone P108G1-7, to identify putative liver specific/hormone responsive functional DNA binding sites. Using deletion analysis, site specific mutagenesis and co-transfection studies, liver specific expression was determined. MRP2 promoter-LUC constructs were highly expressed in liver cell lines compared to non-liver cells. The region extending from -3 to+458 bp of MRP2 promoter starting from AUG contained the potential binding sites for CAATT box enhancer binding protein (C/EBP), hepatocytes nuclear factor 1, 3 and 4 (HNF1, HNF3, and HNF4. Only HNF1 and HNF4 co-transfection with MRP2 luciferase increased expression. Site specific mutational analysis of HNF1 binding site indicated an important role for HNF1alpha. HNF4alpha induction of MRP2 was independent of HNF1 binding site. C/EBP, HNF3, and HNF6 inhibited HNF1alpha while HNF4alpha induced MRP2 luciferase expression and glucocorticoids stimulated MRP2 expression. This study emphasizes the complex regulation of MRP2 with HNF1alpha and HNF4alpha playing a central role. The coordinated regulation of xenobiotic transporters and oxidative conjugation may determine the adaptive responses to cellular detoxification processes.
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Affiliation(s)
- Ishtiaq Qadri
- NUST Center of Virology and Immunology, National University of Science and Technology, Tamizudin Road, Rawalpindi, Pakistan.
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Kamata S, Kishimoto T, Kobayashi S, Miyazaki M. Expression and localization of ATP binding cassette (ABC) family of drug transporters in gastric hepatoid adenocarcinomas. Histopathology 2008; 52:747-54. [DOI: 10.1111/j.1365-2559.2008.03026.x] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/24/2023]
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Kamiyama Y, Matsubara T, Yoshinari K, Nagata K, Kamimura H, Yamazoe Y. Role of human hepatocyte nuclear factor 4alpha in the expression of drug-metabolizing enzymes and transporters in human hepatocytes assessed by use of small interfering RNA. Drug Metab Pharmacokinet 2007; 22:287-98. [PMID: 17827783 DOI: 10.2133/dmpk.22.287] [Citation(s) in RCA: 114] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
Hepatocyte nuclear factor 4alpha (HNF4alpha) is an important transcription factor in hepatic gene expression. Here, we have investigated the role of HNF4alpha in the expression of drug-metabolizing enzymes and transporters in human hepatocytes using an adenovirus expressing human HNF4alpha-small interfering RNA (hHNF4alpha-siRNA). The hHNF4alpha-siRNA effectively reduced the mRNA and nuclear protein levels of hHNF4alpha in a concentration-dependent manner. The hHNF4alpha-siRNA also decreased the mRNA levels of CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP3A4, UGT1A1, UGT1A9, SULT2A1, ABCB1, ABCB11, ABCC2, OATP1B1 and OCT1, as well as those of PXR and CAR. To discern the role of these nuclear receptors, we co-infected hepatocytes with hHNF4alpha-siRNA and PXR- or CAR-expressing adenovirus. The hHNF4alpha-siRNA-induced reductions of the enzyme and transporter mRNA levels were not restored except CYP2B6 mRNA levels, which were returned to the control level by overexpressing CAR. Furthermore, although hHNF4alpha-siRNA did not significantly affect the fold-induction of CYP2B6, CYP2C8, CYP2C9, or CYP3A4 mRNA levels following treatment with CYP inducers, the levels in hHNF4alpha-suppressed cells fell significantly compared to the control. These results suggest that HNF4alpha plays a dominant role in the expression of drug-metabolizing enzymes and transporters in human hepatocytes, and that HNF4alpha expression levels is a possible determinant for inter-individual variations in the expression of these enzymes and transporters.
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Affiliation(s)
- Yoshiteru Kamiyama
- Division of Drug Metabolism and Molecular Toxicology, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan
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