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1
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Yang YH, Wen R, Huang XM, Zhang T, Yang N, Liu CF, Zhang TN. HNF4A mitigates sepsis-associated lung injury by upregulating NCOR2/GR/STAB1 axis and promoting macrophage polarization towards M2 phenotype. Cell Death Dis 2025; 16:120. [PMID: 39979267 PMCID: PMC11842871 DOI: 10.1038/s41419-025-07452-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2024] [Revised: 01/15/2025] [Accepted: 02/12/2025] [Indexed: 02/22/2025]
Abstract
Sepsis can trigger systemic inflammation and lead to detrimental effects on several organs, with particular emphasis on the lungs. In sepsis-associated lung injury, macrophages assume a pivotal role, as their overactivation could facilitate the secretion of inflammatory factors and the imbalance of polarization. Hepatocyte nuclear factor 4 alpha (HNF4A) has been reported its potential involvement in the regulation of inflammatory response and macrophage polarization. This study discusses the role and mechanism of HNF4A in sepsis-induced lung damage. HNF4A exhibits a decrease in expression by analyzing the differentially expressed genes in the lungs of septic mice from the Gene Expression Omnibus dataset GSE15379. Then, we established a mouse sepsis model through a cecal ligation and puncture method and observed that the expression of HNF4A was reduced in both lung tissues and alveolar macrophages. To evaluate the function of HNF4A, we overexpressed HNF4A mediated by adenovirus vectors, which were injected into mice. We found that HNF4A overexpression resulted in a higher survival rate in septic mice and an amelioration of pulmonary damage. Meanwhile, HNF4A overexpression mitigated the infiltration of inflammatory cells and impeded the M1 polarization but facilitated the M2 polarization of macrophages in the lung tissues or the alveolar lavage fluid. In vitro, we treated bone marrow-derived macrophages with interleukin-4. Consistent results were obtained that HNF4A overexpression promoted the M2 polarization of macrophages. Mechanistically, we found that HNF4A transcriptionally regulate the expression of nuclear receptor coactivator 2 (NCOA2) through binding to its promoter region. NCOA2 interacted with glucocorticoid receptor (GR). Stabilin 1 (STAB1) was selected as a possible target by transcriptome sequencing analysis. Functional experiments confirmed STAB1 as a downstream target of the HNF4A/NCOA2/GR axis. Overall, this research investigated the potential impact of HNF4A on pulmonary injury in sepsis. It is suggested that one of the regulatory mechanisms involved in this association may be the NCOR2/GR/STAB1 axis.
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Affiliation(s)
- Yu-Hang Yang
- Department of Pediatrics, PICU, Shengjing Hospital of China Medical University, Shenyang, China
| | - Ri Wen
- Department of Pediatrics, PICU, Shengjing Hospital of China Medical University, Shenyang, China
| | - Xin-Mei Huang
- Department of Endocrinology, Shanghai Fifth People's Hospital, Fudan University, Shanghai, China
| | - Tao Zhang
- Department of Pediatrics, PICU, Shengjing Hospital of China Medical University, Shenyang, China
| | - Ni Yang
- Department of Pediatrics, PICU, Shengjing Hospital of China Medical University, Shenyang, China
| | - Chun-Feng Liu
- Department of Pediatrics, PICU, Shengjing Hospital of China Medical University, Shenyang, China.
| | - Tie-Ning Zhang
- Department of Pediatrics, PICU, Shengjing Hospital of China Medical University, Shenyang, China.
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2
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Ma X, Huang T, Chen X, Li Q, Liao M, Fu L, Huang J, Yuan K, Wang Z, Zeng Y. Molecular mechanisms in liver repair and regeneration: from physiology to therapeutics. Signal Transduct Target Ther 2025; 10:63. [PMID: 39920130 PMCID: PMC11806117 DOI: 10.1038/s41392-024-02104-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2023] [Revised: 09/02/2024] [Accepted: 12/12/2024] [Indexed: 02/09/2025] Open
Abstract
Liver repair and regeneration are crucial physiological responses to hepatic injury and are orchestrated through intricate cellular and molecular networks. This review systematically delineates advancements in the field, emphasizing the essential roles played by diverse liver cell types. Their coordinated actions, supported by complex crosstalk within the liver microenvironment, are pivotal to enhancing regenerative outcomes. Recent molecular investigations have elucidated key signaling pathways involved in liver injury and regeneration. Viewed through the lens of metabolic reprogramming, these pathways highlight how shifts in glucose, lipid, and amino acid metabolism support the cellular functions essential for liver repair and regeneration. An analysis of regenerative variability across pathological states reveals how disease conditions influence these dynamics, guiding the development of novel therapeutic strategies and advanced techniques to enhance liver repair and regeneration. Bridging laboratory findings with practical applications, recent clinical trials highlight the potential of optimizing liver regeneration strategies. These trials offer valuable insights into the effectiveness of novel therapies and underscore significant progress in translational research. In conclusion, this review intricately links molecular insights to therapeutic frontiers, systematically charting the trajectory from fundamental physiological mechanisms to innovative clinical applications in liver repair and regeneration.
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Affiliation(s)
- Xiao Ma
- Division of Liver Surgery, Department of General Surgery and Laboratory of Liver Surgery, and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China
| | - Tengda Huang
- Division of Liver Surgery, Department of General Surgery and Laboratory of Liver Surgery, and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China
| | - Xiangzheng Chen
- Division of Liver Surgery, Department of General Surgery and Laboratory of Liver Surgery, and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China
| | - Qian Li
- Division of Liver Surgery, Department of General Surgery and Laboratory of Liver Surgery, and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China
| | - Mingheng Liao
- Division of Liver Surgery, Department of General Surgery and Laboratory of Liver Surgery, and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China
| | - Li Fu
- Division of Liver Surgery, Department of General Surgery and Laboratory of Liver Surgery, and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China
| | - Jiwei Huang
- Division of Liver Surgery, Department of General Surgery and Laboratory of Liver Surgery, and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China
| | - Kefei Yuan
- Division of Liver Surgery, Department of General Surgery and Laboratory of Liver Surgery, and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China
| | - Zhen Wang
- Division of Liver Surgery, Department of General Surgery and Laboratory of Liver Surgery, and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China.
| | - Yong Zeng
- Division of Liver Surgery, Department of General Surgery and Laboratory of Liver Surgery, and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China.
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3
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Sun Y, Yuan X, Hu Z, Li Y. Harnessing nuclear receptors to modulate hepatic stellate cell activation for liver fibrosis resolution. Biochem Pharmacol 2025; 232:116730. [PMID: 39710274 DOI: 10.1016/j.bcp.2024.116730] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2024] [Revised: 12/04/2024] [Accepted: 12/19/2024] [Indexed: 12/24/2024]
Abstract
With the recent approval of Resmetirom as the first drug targeting nuclear receptors for metabolic dysfunction-associated steatohepatitis (MASH), there is promising way to treat MASH-associated liver fibrosis. However, liver fibrosis can arise from various pathogenic factors, and effective treatments for fibrosis due to other causes remain elusive. The activation of hepatic stellate cells (HSCs) represents a central link in the pathogenesis of hepatic fibrosis. Therefore, harnessing nuclear receptors to modulate HSC activation may be an effective approach to resolving the complex liver fibrosis caused by various factors. In this comprehensive review, we systematically explore the structure and physiological functions of nuclear receptors, shedding light on their multifaceted roles in HSC activation. Recent advancements in drug development targeting nuclear receptors are discussed, providing insights into their potential as rational and effective therapeutic targets for modulating HSC activation in the context of liver fibrosis. By elucidating the intricate interplay between nuclear receptors and HSC activation, this review contributes to the discovery of new nuclear receptor targets in HSCs for resolving hepatic fibrosis.
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Affiliation(s)
- Yaxin Sun
- Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China; Zhongshan Institute for Drug Discovery, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Zhongshan, China; University of Chinese Academy of Sciences, Beijing, China
| | - Xiaoyan Yuan
- Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China; Zhongshan Institute for Drug Discovery, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Zhongshan, China; University of Chinese Academy of Sciences, Beijing, China
| | - Zhenhua Hu
- Zhongshan Institute for Drug Discovery, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Zhongshan, China; Department of Health and Nursing, Nanfang College of Sun Yat-sen University, Guangzhou, China.
| | - Yuanyuan Li
- Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China; Zhongshan Institute for Drug Discovery, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Zhongshan, China; University of Chinese Academy of Sciences, Beijing, China.
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4
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Dubois V, Lefebvre P, Staels B, Eeckhoute J. Nuclear receptors: pathophysiological mechanisms and drug targets in liver disease. Gut 2024; 73:1562-1569. [PMID: 38862216 DOI: 10.1136/gutjnl-2023-331741] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/28/2024] [Accepted: 05/18/2024] [Indexed: 06/13/2024]
Abstract
Nuclear receptors (NRs) are ligand-dependent transcription factors required for liver development and function. As a consequence, NRs have emerged as attractive drug targets in a wide range of liver diseases. However, liver dysfunction and failure are linked to loss of hepatocyte identity characterised by deficient NR expression and activities. This might at least partly explain why several pharmacological NR modulators have proven insufficiently efficient to improve liver functionality in advanced stages of diseases such as metabolic dysfunction-associated steatotic liver disease (MASLD). In this perspective, we review the most recent advances in the hepatic NR field and discuss the contribution of multiomic approaches to our understanding of their role in the molecular organisation of an intricated transcriptional regulatory network, as well as in liver intercellular dialogues and interorgan cross-talks. We discuss the potential benefit of novel therapeutic approaches simultaneously targeting multiple NRs, which would not only reactivate the hepatic NR network and restore hepatocyte identity but also impact intercellular and interorgan interplays whose importance to control liver functions is further defined. Finally, we highlight the need of considering individual parameters such as sex and disease stage in the development of NR-based clinical strategies.
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Affiliation(s)
- Vanessa Dubois
- Basic and Translational Endocrinology (BaTE), Department of Basic and Applied Medical Sciences, Ghent University, Gent, Belgium
| | - Philippe Lefebvre
- Univ. Lille, Inserm, CHU Lille, Institut Pasteur de Lille, U1011-EGID, Lille, France
| | - Bart Staels
- Univ. Lille, Inserm, CHU Lille, Institut Pasteur de Lille, U1011-EGID, Lille, France
| | - Jerome Eeckhoute
- Univ. Lille, Inserm, CHU Lille, Institut Pasteur de Lille, U1011-EGID, Lille, France
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5
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Kotulkar M, Paine-Cabrera D, Robarts DR, Apte U. Regulation of hepatic xenosensor function by HNF4alpha. Toxicol Sci 2024; 200:346-356. [PMID: 38810120 DOI: 10.1093/toxsci/kfae069] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 05/31/2024] Open
Abstract
Nuclear receptors such as constitutive androstane receptor (CAR), pregnane X receptor (PXR), and peroxisome proliferator-activated receptor-alpha (PPARα), and transcription factors with nuclear receptor type activity such as aryl hydrocarbon receptor (AhR) function as xenobiotic sensors. Hepatocyte nuclear factor 4alpha (HNF4α) is a highly conserved orphan nuclear receptor essential for liver function. We tested the hypothesis that HNF4α is essential for the function of these 4 major xenosensors. Wild-type (WT) and hepatocyte-specific Hnf4a null (HNF4α-KO) mice were treated with the mouse-specific activators of AhR (TCDD, 30 µg/kg), CAR (TCPOBOP, 2.5 µg/g), PXR, (PCN, 100 µg/g), and PPARα (WY-14643, 1 mg/kg). Blood and liver tissue samples were collected to study receptor activation. TCDD (AhR agonist) treatment did not affect the liver-to-body weight ratio (LW/BW) in either WT or HNF4α-KO mice. Further, TCDD activated AhR in both WT and HNF4α-KO mice, confirmed by increase in expression of AhR target genes. TCPOBOP (CAR agonist) significantly increased the LW/BW ratio and CAR target gene expression in WT mice, but not in HNF4α-KO mice. PCN (a mouse PXR agonist) significantly increased LW/BW ratio in both WT and HNF4α-KO mice however, failed to induce PXR target genes in HNF4α-KO mice. The treatment of WY-14643 (PPARα agonist) increased LW/BW ratio and PPARα target gene expression in WT mice but not in HNF4α-KO mice. Together, these data indicate that the function of CAR, PXR, and PPARα but not of AhR was disrupted in HNF4α-KO mice. These results demonstrate that HNF4α function is critical for the activation of hepatic xenosensors, which are critical for toxicological responses.
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MESH Headings
- Animals
- Hepatocyte Nuclear Factor 4/metabolism
- Hepatocyte Nuclear Factor 4/genetics
- Liver/metabolism
- Liver/drug effects
- PPAR alpha/agonists
- PPAR alpha/metabolism
- PPAR alpha/genetics
- Mice, Knockout
- Constitutive Androstane Receptor
- Pregnane X Receptor/genetics
- Pregnane X Receptor/metabolism
- Receptors, Cytoplasmic and Nuclear/genetics
- Receptors, Cytoplasmic and Nuclear/agonists
- Receptors, Cytoplasmic and Nuclear/metabolism
- Mice
- Receptors, Steroid/genetics
- Receptors, Steroid/metabolism
- Receptors, Steroid/agonists
- Receptors, Aryl Hydrocarbon/agonists
- Receptors, Aryl Hydrocarbon/genetics
- Receptors, Aryl Hydrocarbon/metabolism
- Mice, Inbred C57BL
- Male
- Pyrimidines/pharmacology
- Polychlorinated Dibenzodioxins/toxicity
- Pyridines/pharmacology
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Affiliation(s)
- Manasi Kotulkar
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS 66160, United States
| | - Diego Paine-Cabrera
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS 66160, United States
| | - Dakota R Robarts
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS 66160, United States
| | - Udayan Apte
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS 66160, United States
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6
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Kotulkar M, Paine-Cabrera D, Apte U. Role of Hepatocyte Nuclear Factor 4 Alpha in Liver Cancer. Semin Liver Dis 2024; 44:383-393. [PMID: 38901435 DOI: 10.1055/a-2349-7236] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 06/22/2024]
Abstract
Liver cancer is the sixth most common cancer and the fourth leading cause of cancer-related deaths worldwide. Hepatocellular carcinoma (HCC) is the most prevalent primary liver cancer and the incidence of HCC is on the rise. Liver cancers in general and HCC in particular do not respond to chemotherapy. Radiological ablation, surgical resection, and liver transplantation are the only medical therapies currently available. Hepatocyte nuclear factor 4 α (HNF4α) is an orphan nuclear receptor expressed only in hepatocytes in the liver. HNF4α is considered the master regulator of hepatic differentiation because it regulates a significant number of genes involved in various liver-specific functions. In addition to maintaining hepatic differentiation, HNF4α also acts as a tumor suppressor by inhibiting hepatocyte proliferation by suppressing the expression of promitogenic genes and inhibiting epithelial to mesenchymal transition in hepatocytes. Loss of HNF4α expression and function is associated with rapid progression of chronic liver diseases that ultimately lead to liver cirrhosis and HCC, including metabolism-associated steatohepatitis, alcohol-associated liver disease, and hepatitis virus infection. This review summarizes the role of HNF4α in liver cancer pathogenesis and highlights its potential as a potential therapeutic target for HCC.
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Affiliation(s)
- Manasi Kotulkar
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas
| | - Diego Paine-Cabrera
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas
| | - Udayan Apte
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas
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7
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de Haan LR, van Golen RF, Heger M. Molecular Pathways Governing the Termination of Liver Regeneration. Pharmacol Rev 2024; 76:500-558. [PMID: 38697856 DOI: 10.1124/pharmrev.123.000955] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2023] [Revised: 01/24/2024] [Accepted: 02/08/2024] [Indexed: 05/05/2024] Open
Abstract
The liver has the unique capacity to regenerate, and up to 70% of the liver can be removed without detrimental consequences to the organism. Liver regeneration is a complex process involving multiple signaling networks and organs. Liver regeneration proceeds through three phases: the initiation phase, the growth phase, and the termination phase. Termination of liver regeneration occurs when the liver reaches a liver-to-body weight that is required for homeostasis, the so-called "hepatostat." The initiation and growth phases have been the subject of many studies. The molecular pathways that govern the termination phase, however, remain to be fully elucidated. This review summarizes the pathways and molecules that signal the cessation of liver regrowth after partial hepatectomy and answers the question, "What factors drive the hepatostat?" SIGNIFICANCE STATEMENT: Unraveling the pathways underlying the cessation of liver regeneration enables the identification of druggable targets that will allow us to gain pharmacological control over liver regeneration. For these purposes, it would be useful to understand why the regenerative capacity of the liver is hampered under certain pathological circumstances so as to artificially modulate the regenerative processes (e.g., by blocking the cessation pathways) to improve clinical outcomes and safeguard the patient's life.
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Affiliation(s)
- Lianne R de Haan
- Jiaxing Key Laboratory for Photonanomedicine and Experimental Therapeutics, Department of Pharmaceutics, College of Medicine, Jiaxing University, Jiaxing, China (L.R.d.H., M.H.); Department of Internal Medicine, Noordwest Ziekenhuisgroep, Alkmaar, The Netherlands (L.R.d.H.); Department of Gastroenterology and Hepatology, Leiden University Medical Center, Leiden, The Netherlands (R.F.v.G.); Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands (M.H.); and Membrane Biochemistry and Biophysics, Department of Chemistry, Faculty of Science, Utrecht University, Utrecht, The Netherlands (M.H.)
| | - Rowan F van Golen
- Jiaxing Key Laboratory for Photonanomedicine and Experimental Therapeutics, Department of Pharmaceutics, College of Medicine, Jiaxing University, Jiaxing, China (L.R.d.H., M.H.); Department of Internal Medicine, Noordwest Ziekenhuisgroep, Alkmaar, The Netherlands (L.R.d.H.); Department of Gastroenterology and Hepatology, Leiden University Medical Center, Leiden, The Netherlands (R.F.v.G.); Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands (M.H.); and Membrane Biochemistry and Biophysics, Department of Chemistry, Faculty of Science, Utrecht University, Utrecht, The Netherlands (M.H.)
| | - Michal Heger
- Jiaxing Key Laboratory for Photonanomedicine and Experimental Therapeutics, Department of Pharmaceutics, College of Medicine, Jiaxing University, Jiaxing, China (L.R.d.H., M.H.); Department of Internal Medicine, Noordwest Ziekenhuisgroep, Alkmaar, The Netherlands (L.R.d.H.); Department of Gastroenterology and Hepatology, Leiden University Medical Center, Leiden, The Netherlands (R.F.v.G.); Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands (M.H.); and Membrane Biochemistry and Biophysics, Department of Chemistry, Faculty of Science, Utrecht University, Utrecht, The Netherlands (M.H.)
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8
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Matsumoto S, Kikuchi A. Wnt/β-catenin signaling pathway in liver biology and tumorigenesis. In Vitro Cell Dev Biol Anim 2024; 60:466-481. [PMID: 38379098 DOI: 10.1007/s11626-024-00858-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2023] [Accepted: 01/16/2024] [Indexed: 02/22/2024]
Abstract
The Wnt/β-catenin pathway is an evolutionarily conserved signaling pathway that controls fundamental physiological and pathological processes by regulating cell proliferation and differentiation. The Wnt/β-catenin pathway enables liver homeostasis by inducing differentiation and contributes to liver-specific features such as metabolic zonation and regeneration. In contrast, abnormalities in the Wnt/β-catenin pathway promote the development and progression of hepatocellular carcinoma (HCC). Similarly, hepatoblastoma, the most common childhood liver cancer, is frequently associated with β-catenin mutations, which activate Wnt/β-catenin signaling. HCCs with activation of the Wnt/β-catenin pathway have unique gene expression patterns and pathological and clinical features. Accordingly, they are highly differentiated with retaining hepatocyte-like characteristics and tumorigenic. Activation of the Wnt/β-catenin pathway in HCC also alters the state of immune cells, causing "immune evasion" with inducing resistance to immune checkpoint inhibitors, which have recently become widely used to treat HCC. Activated Wnt/β-catenin signaling exhibits these phenomena in liver tumorigenesis through the expression of downstream target genes, and the molecular basis is still poorly understood. In this review, we describe the physiological roles of Wnt/b-catenin signaling and then discuss their characteristic changes by the abnormal activation of Wnt/b-catenin signaling. Clarification of the mechanism would contribute to the development of therapeutic agents in the future.
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Affiliation(s)
- Shinji Matsumoto
- Departments of Molecular Biology and Biochemistry, Graduate School of Medicine, Osaka University, 2-2 Yamada-Oka, Suita, Osaka, 565-0871, Japan.
- Integrated Frontier Research for Medical Science Division, Institute for Open and Transdisciplinary Research Initiatives (OTRI), 2-2 Yamada-Oka, Suita, Osaka, 565-0871, Japan.
| | - Akira Kikuchi
- Departments of Molecular Biology and Biochemistry, Graduate School of Medicine, Osaka University, 2-2 Yamada-Oka, Suita, Osaka, 565-0871, Japan
- Center of Infectious Disease Education and Research (CiDER), Osaka University, 2-8 Yamada-Oka, Suita, Osaka, 565-0871, Japan
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9
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Thakur A, Park K, Cullum R, Fuglerud BM, Khoshnoodi M, Drissler S, Stephan TL, Lotto J, Kim D, Gonzalez FJ, Hoodless PA. HNF4A guides the MLL4 complex to establish and maintain H3K4me1 at gene regulatory elements. Commun Biol 2024; 7:144. [PMID: 38297077 PMCID: PMC10830483 DOI: 10.1038/s42003-024-05835-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2022] [Accepted: 01/18/2024] [Indexed: 02/02/2024] Open
Abstract
Hepatocyte nuclear factor 4A (HNF4A/NR2a1), a transcriptional regulator of hepatocyte identity, controls genes that are crucial for liver functions, primarily through binding to enhancers. In mammalian cells, active and primed enhancers are marked by monomethylation of histone 3 (H3) at lysine 4 (K4) (H3K4me1) in a cell type-specific manner. How this modification is established and maintained at enhancers in connection with transcription factors (TFs) remains unknown. Using analysis of genome-wide histone modifications, TF binding, chromatin accessibility and gene expression, we show that HNF4A is essential for an active chromatin state. Using HNF4A loss and gain of function experiments in vivo and in cell lines in vitro, we show that HNF4A affects H3K4me1, H3K27ac and chromatin accessibility, highlighting its contribution to the establishment and maintenance of a transcriptionally permissive epigenetic state. Mechanistically, HNF4A interacts with the mixed-lineage leukaemia 4 (MLL4) complex facilitating recruitment to HNF4A-bound regions. Our findings indicate that HNF4A enriches H3K4me1, H3K27ac and establishes chromatin opening at transcriptional regulatory regions.
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Affiliation(s)
- Avinash Thakur
- Terry Fox Laboratory, BC Cancer, Vancouver, V5Z 1L3, Canada
- Department of Medical Genetics, University of British Columbia, Vancouver, V6T 1Z4, Canada
| | - Kwangjin Park
- Terry Fox Laboratory, BC Cancer, Vancouver, V5Z 1L3, Canada
- Department of Medical Genetics, University of British Columbia, Vancouver, V6T 1Z4, Canada
| | - Rebecca Cullum
- Terry Fox Laboratory, BC Cancer, Vancouver, V5Z 1L3, Canada
| | - Bettina M Fuglerud
- Terry Fox Laboratory, BC Cancer, Vancouver, V5Z 1L3, Canada
- Department of Medical Genetics, University of British Columbia, Vancouver, V6T 1Z4, Canada
| | | | - Sibyl Drissler
- Terry Fox Laboratory, BC Cancer, Vancouver, V5Z 1L3, Canada
- Cell and Developmental Biology Program, University of British Columbia, Vancouver, V6T 1Z4, Canada
| | - Tabea L Stephan
- Terry Fox Laboratory, BC Cancer, Vancouver, V5Z 1L3, Canada
- Cell and Developmental Biology Program, University of British Columbia, Vancouver, V6T 1Z4, Canada
| | - Jeremy Lotto
- Terry Fox Laboratory, BC Cancer, Vancouver, V5Z 1L3, Canada
- Cell and Developmental Biology Program, University of British Columbia, Vancouver, V6T 1Z4, Canada
| | - Donghwan Kim
- Center of Cancer Research, National Cancer Institute, Bethesda, 2089, USA
| | - Frank J Gonzalez
- Center of Cancer Research, National Cancer Institute, Bethesda, 2089, USA
| | - Pamela A Hoodless
- Terry Fox Laboratory, BC Cancer, Vancouver, V5Z 1L3, Canada.
- Department of Medical Genetics, University of British Columbia, Vancouver, V6T 1Z4, Canada.
- Cell and Developmental Biology Program, University of British Columbia, Vancouver, V6T 1Z4, Canada.
- School of Biomedical Engineering, University of British Columbia, Vancouver, V6T 1Z4, Canada.
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10
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Deans JR, Deol P, Titova N, Radi SH, Vuong LM, Evans JR, Pan S, Fahrmann J, Yang J, Hammock BD, Fiehn O, Fekry B, Eckel-Mahan K, Sladek FM. HNF4α isoforms regulate the circadian balance between carbohydrate and lipid metabolism in the liver. Front Endocrinol (Lausanne) 2023; 14:1266527. [PMID: 38111711 PMCID: PMC10726135 DOI: 10.3389/fendo.2023.1266527] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/25/2023] [Accepted: 11/06/2023] [Indexed: 12/20/2023] Open
Abstract
Hepatocyte Nuclear Factor 4α (HNF4α), a master regulator of hepatocyte differentiation, is regulated by two promoters (P1 and P2) which drive the expression of different isoforms. P1-HNF4α is the major isoform in the adult liver while P2-HNF4α is thought to be expressed only in fetal liver and liver cancer. Here, we show that P2-HNF4α is indeed expressed in the normal adult liver at Zeitgeber time (ZT)9 and ZT21. Using exon swap mice that express only P2-HNF4α we show that this isoform orchestrates a distinct transcriptome and metabolome via unique chromatin and protein-protein interactions, including with different clock proteins at different times of the day leading to subtle differences in circadian gene regulation. Furthermore, deletion of the Clock gene alters the circadian oscillation of P2- (but not P1-)HNF4α RNA, revealing a complex feedback loop between the HNF4α isoforms and the hepatic clock. Finally, we demonstrate that while P1-HNF4α drives gluconeogenesis, P2-HNF4α drives ketogenesis and is required for elevated levels of ketone bodies in female mice. Taken together, we propose that the highly conserved two-promoter structure of the Hnf4a gene is an evolutionarily conserved mechanism to maintain the balance between gluconeogenesis and ketogenesis in the liver in a circadian fashion.
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Affiliation(s)
- Jonathan R. Deans
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, Riverside, CA, United States
- Genetics, Genomics and Bioinformatics Graduate Program, University of California, Riverside, Riverside, CA, United States
| | - Poonamjot Deol
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, Riverside, CA, United States
| | - Nina Titova
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, Riverside, CA, United States
| | - Sarah H. Radi
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, Riverside, CA, United States
- Biochemistry and Molecular Biology Graduate Program, University of California, Riverside, Riverside, CA, United States
| | - Linh M. Vuong
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, Riverside, CA, United States
| | - Jane R. Evans
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, Riverside, CA, United States
| | - Songqin Pan
- Proteomics Core, Institute for Integrative Genome Biology, University of California, Riverside, Riverside, CA, United States
| | - Johannes Fahrmann
- National Institutes of Health West Coast Metabolomics Center, University of California, Davis, Davis, CA, United States
| | - Jun Yang
- Department of Entomology and Nematology & UCD Comprehensive Cancer Center, University of California, Davis, Davis, CA, United States
| | - Bruce D. Hammock
- Department of Entomology and Nematology & UCD Comprehensive Cancer Center, University of California, Davis, Davis, CA, United States
| | - Oliver Fiehn
- National Institutes of Health West Coast Metabolomics Center, University of California, Davis, Davis, CA, United States
| | - Baharan Fekry
- Department of Biochemistry and Molecular Biology, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, TX, United States
| | - Kristin Eckel-Mahan
- Department of Biochemistry and Molecular Biology, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, TX, United States
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, TX, United States
| | - Frances M. Sladek
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, Riverside, CA, United States
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11
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Namachivayam A, Valsala Gopalakrishnan A. Effect of Lauric acid against ethanol-induced hepatotoxicity by modulating oxidative stress/apoptosis signalling and HNF4α in Wistar albino rats. Heliyon 2023; 9:e21267. [PMID: 37908709 PMCID: PMC10613920 DOI: 10.1016/j.heliyon.2023.e21267] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2023] [Revised: 10/12/2023] [Accepted: 10/18/2023] [Indexed: 11/02/2023] Open
Abstract
Ethanol (EtOH) is most widely used in alcoholic beverages to prepare alcohol. As EtOH is mainly metabolised in the liver, the excessive consumption of EtOH forms a primary toxic metabolic product called acetaldehyde, as the gradual increase in acetaldehyde leads to liver injury, as reported. Lauric acid (LA) is rich in antioxidant, antifungal, antibacterial, anticancer, and antiviral properties. LA is an edible component highly present in coconut oil. However, no report on LA protective effects against the EtOH-instigated hepatotoxicity exists. Therefore, the experiment is carried out to investigate the potency effects of LA on EtOH-instigated hepatotoxicity in thirty male albino rats. Rats were divided into five groups (n-6): control DMSO alone, EtOH -intoxicated, EtOH + LA 180 mg/kg, EtOH + LA 360 mg/kg, and LA alone were administered orally using oral gavage. The study measured body weight every weekend in all rat groups. The rats were sacrificed and assessed for serum markers (alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase), antioxidant activity (superoxide dismutase, reduced glutathione, glutathione peroxidase), lipid peroxidation (malondialdehyde), histopathological, cytokine levels (TNF-α, IL-1β and IL-6), protein expression (caspase 3 and caspase 8 and Bcl-2 and HNF4α) were evaluated after the 56-days study period. The impact of EtOH intoxication reduces the rat's body weight by 90 g, upregulates the liver enzyme markers, depletes the antioxidant levels, produces malondialdehyde, changes the histoarchitecture (periportal inflammation and hepatocyte damage), downregulates the Bcl-2 expressions and HNF4α, and elevates the expression of cytokines and apoptotic markers. LA alleviated EtOH-induced liver toxicity by significant (p < 0.05) modulation of biochemical levels, caspase-8/3 signalling, reducing pro-inflammatory cytokines, and restoring the normal histoarchitecture, upregulating the Bcl-2 and HNF4α Expressions. In conclusion, LA treatment can protect the liver against EtOH-induced hepatotoxicity, evidenced by alleviating Oxidative stress, lipid peroxidation, inflammation, apoptosis, and upregulation of HNF4α.
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Affiliation(s)
- Arunraj Namachivayam
- Department of Biomedical Sciences, School of Bio-Sciences and Technology, Vellore Institute of Technology (VIT), Vellore 632014, India
| | - Abilash Valsala Gopalakrishnan
- Department of Biomedical Sciences, School of Bio-Sciences and Technology, Vellore Institute of Technology (VIT), Vellore 632014, India
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12
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Shah A, Huck I, Duncan K, Gansemer ER, Liu K, Adajar RC, Apte U, Stamnes MA, Rutkowski DT. Interference with the HNF4-dependent gene regulatory network diminishes endoplasmic reticulum stress in hepatocytes. Hepatol Commun 2023; 7:e0278. [PMID: 37820274 PMCID: PMC10578741 DOI: 10.1097/hc9.0000000000000278] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/21/2023] [Accepted: 08/08/2023] [Indexed: 10/13/2023] Open
Abstract
BACKGROUND In all eukaryotic cell types, the unfolded protein response (UPR) upregulates factors that promote protein folding and misfolded protein clearance to help alleviate endoplasmic reticulum (ER) stress. Yet, ER stress in the liver is uniquely accompanied by the suppression of metabolic genes, the coordination and purpose of which are largely unknown. METHODS Here, we combined in silico machine learning, in vivo liver-specific deletion of the master regulator of hepatocyte differentiation HNF4α, and in vitro manipulation of hepatocyte differentiation state to determine how the UPR regulates hepatocyte identity and toward what end. RESULTS Machine learning identified a cluster of correlated genes that were profoundly suppressed by persistent ER stress in the liver. These genes, which encode diverse functions including metabolism, coagulation, drug detoxification, and bile synthesis, are likely targets of the master regulator of hepatocyte differentiation HNF4α. The response of these genes to ER stress was phenocopied by liver-specific deletion of HNF4α. Strikingly, while deletion of HNF4α exacerbated liver injury in response to an ER stress challenge, it also diminished UPR activation and partially preserved ER ultrastructure, suggesting attenuated ER stress. Conversely, pharmacological maintenance of hepatocyte identity in vitro enhanced sensitivity to stress. CONCLUSIONS Together, our findings suggest that the UPR regulates hepatocyte identity through HNF4α to protect ER homeostasis even at the expense of liver function.
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Affiliation(s)
- Anit Shah
- Department of Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, Iowa, USA
| | - Ian Huck
- Department of Pharmacology, Toxicology, and Therapeutics, Kansas University Medical Center, Kansas City, Kansas, USA
| | - Kaylia Duncan
- Department of Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, Iowa, USA
| | - Erica R. Gansemer
- Department of Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, Iowa, USA
| | - Kaihua Liu
- Department of Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, Iowa, USA
| | - Reed C. Adajar
- Department of Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, Iowa, USA
| | - Udayan Apte
- Department of Pharmacology, Toxicology, and Therapeutics, Kansas University Medical Center, Kansas City, Kansas, USA
| | - Mark A. Stamnes
- Department of Molecular Physiology and Biophysics, University of Iowa Carver College of Medicine, Iowa City, Iowa, USA
| | - D. Thomas Rutkowski
- Department of Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, Iowa, USA
- Department of Internal Medicine, University of Iowa Carver College of Medicine, Iowa City, Iowa, USA
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa Carver College of Medicine, Iowa City, Iowa, USA
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13
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Kotulkar M, Cabrera DP, Robarts D, Apte U. Regulation of Hepatic Xenosensor Function by HNF4alpha. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.10.11.561888. [PMID: 37873133 PMCID: PMC10592787 DOI: 10.1101/2023.10.11.561888] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 10/25/2023]
Abstract
Nuclear receptors including Aryl hydrocarbon Receptor (AhR), Constitutive Androstane Receptor (CAR), Pregnane X Receptor (PXR), and Peroxisome Proliferator-Activated Receptor-alpha (PPARα) function as xenobiotic sensors. Hepatocyte nuclear factor 4alpha (HNF4α) is a highly conserved orphan nuclear receptor essential for liver function. We tested the hypothesis that HNF4α is essential for function of these four major xenosensors. Wild-type (WT) and hepatocyte-specific HNF4α knockout (HNF4α-KO) mice were treated with the mouse-specific activators of AhR (TCDD, 30 µg/kg), CAR (TCPOBOP, 2.5 µg/g), PXR, (PCN, 100 µg/g), and PPARα (WY-14643, 1 mg/kg). Blood and liver tissue samples were collected to study nuclear receptor activation. TCDD (AhR agonist) treatment did not affect the liver-to-body weight ratio (LW/BW) in either WT or HNF4α-KO mice. Further, TCDD activated AhR in both WT and HNF4-KO mice, confirmed by increase in expression of its target genes. TCPOBOP (CAR agonist) significantly increased the LW/BW ratio and CAR target gene expression in WT mice, but not in HNF4α-KO mice. PCN (a mouse PXR agonist) significantly increased LW/BW ratio in both WT and HNF4α-KO mice however, it failed to induce PXR target genes in HNF4 KO mice. The treatment of WY-14643 (PPARα agonist) increased LW/BW ratio and PPARα target gene expression in WT mice but not in HNF4α-KO mice. Together, these data indicate that the function of CAR, PXR, and PPARα but not of AhR was disrupted in HNF4α-KO mice. These results demonstrate that HNF4α function is critical for the activation of hepatic xenosensors, which are critical for toxicological responses.
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14
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Kotulkar M, Paine-Cabrera D, Abernathy S, Robarts DR, Parkes WS, Lin-Rahardja K, Numata S, Lebofsky M, Jaeschke H, Apte U. Role of HNF4alpha-cMyc interaction in liver regeneration and recovery after acetaminophen-induced acute liver injury. Hepatology 2023; 78:1106-1117. [PMID: 37021787 PMCID: PMC10523339 DOI: 10.1097/hep.0000000000000367] [Citation(s) in RCA: 24] [Impact Index Per Article: 12.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/01/2022] [Accepted: 03/01/2023] [Indexed: 04/07/2023]
Abstract
BACKGROUND AND AIMS Overdose of acetaminophen (APAP) is the major cause of acute liver failure in the western world. We report a novel signaling interaction between hepatocyte nuclear factor 4 alpha (HNF4α) cMyc and nuclear factor erythroid 2-related factor 2 (Nrf2) during liver injury and regeneration after APAP overdose. APPROACH AND RESULTS APAP-induced liver injury and regeneration were studied in male C57BL/6J (WT) mice, hepatocyte-specific HNF4α knockout mice (HNF4α-KO), and HNF4α-cMyc double knockout mice (DKO). C57BL/6J mice treated with 300 mg/kg maintained nuclear HNF4α expression and exhibited liver regeneration, resulting in recovery. However, treatment with 600-mg/kg APAP, where liver regeneration was inhibited and recovery was delayed, showed a rapid decline in HNF4α expression. HNF4α-KO mice developed significantly higher liver injury due to delayed glutathione recovery after APAP overdose. HNF4α-KO mice also exhibited significant induction of cMyc, and the deletion of cMyc in HNF4α-KO mice (DKO mice) reduced the APAP-induced liver injury. The DKO mice had significantly faster glutathione replenishment due to rapid induction in Gclc and Gclm genes. Coimmunoprecipitation and ChIP analyses revealed that HNF4α interacts with Nrf2 and affects its DNA binding. Furthermore, DKO mice showed significantly faster initiation of cell proliferation resulting in rapid liver regeneration and recovery. CONCLUSIONS These data show that HNF4α interacts with Nrf2 and promotes glutathione replenishment aiding in recovery from APAP-induced liver injury, a process inhibited by cMyc. These studies indicate that maintaining the HNF4α function is critical for regeneration and recovery after APAP overdose.
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Affiliation(s)
- Manasi Kotulkar
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas, USA
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15
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Elias G, Schonfeld M, Saleh S, Parrish M, Barmanova M, Weinman SA, Tikhanovich I. Sepsis-induced endothelial dysfunction drives acute-on-chronic liver failure through Angiopoietin-2-HGF-C/EBPβ pathway. Hepatology 2023; 78:803-819. [PMID: 36943063 PMCID: PMC10440279 DOI: 10.1097/hep.0000000000000354] [Citation(s) in RCA: 8] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/19/2022] [Accepted: 02/13/2023] [Indexed: 03/23/2023]
Abstract
BACKGROUND AND AIMS Acute-on-chronic liver failure (ACLF) is an acute liver and multisystem failure in patients with previously stable cirrhosis. A common cause of ACLF is sepsis secondary to bacterial infection. Sepsis-associated ACLF involves a loss of differentiated liver function in the absence of direct liver injury, and its mechanism is unknown. We aimed to study the mechanism of sepsis-associated ACLF using a novel mouse model. APPROACH AND RESULTS Sepsis-associated ACLF was induced by cecal ligation and puncture procedure (CLP) in mice treated with thioacetamide (TAA). The combination of TAA and CLP resulted in a significant decrease in liver synthetic function and high mortality. These changes were associated with reduced metabolic gene expression and increased CCAAT enhancer binding protein beta (C/EBPβ) transcriptional activity. We found that C/EBPβ binding to its target gene promoters was increased. In humans, C/EBPβ chromatin binding was similarly increased in the ACLF group compared with control cirrhosis. Hepatocyte-specific Cebpb knockout mice had reduced mortality and increased gene expression of hepatocyte differentiation markers in TAA/CLP mice, suggesting that C/EBPβ promotes liver failure in these mice. C/EBPβ activation was associated with endothelial dysfunction, characterized by reduced Angiopoietin-1/Angiopoietin-2 ratio and increased endothelial production of HGF. Angiopoietin-1 supplementation or Hgf knockdown reduced hepatocyte C/EBPβ accumulation, restored liver function, and reduced mortality, suggesting that endothelial dysfunction induced by sepsis drives ACLF through HGF-C/EBPβ pathway. CONCLUSIONS The transcription factor C/EBPβ is activated in both mouse and human ACLF and is a potential therapeutic target to prevent liver failure in patients with sepsis and cirrhosis.
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Affiliation(s)
- Grant Elias
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Michael Schonfeld
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Sara Saleh
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Mark Parrish
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Marina Barmanova
- Liver Center, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Steven A Weinman
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
- Kansas City VA Medical Center, Kansas City, MO, USA
| | - Irina Tikhanovich
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
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16
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Deng Y, Han X, Chen H, Zhao C, Chen Y, Zhou J, de The H, Zhu J, Yuan H. Ypel5 regulates liver development and function in zebrafish. J Mol Cell Biol 2023; 15:mjad019. [PMID: 36948605 PMCID: PMC10588938 DOI: 10.1093/jmcb/mjad019] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2022] [Revised: 03/16/2023] [Accepted: 03/20/2023] [Indexed: 03/24/2023] Open
Abstract
YPEL5 is a member of the Yippee-like (YPEL) gene family that is evolutionarily conserved in eukaryotic species. To date, the physiological function of YPEL5 has not been assessed due to a paucity of genetic animal models. Here, using CRISPR/Cas9-mediated genome editing, we generated a stable ypel5-/- mutant zebrafish line. Disruption of ypel5 expression leads to liver enlargement associated with hepatic cell proliferation. Meanwhile, hepatic metabolism and function are dysregulated in ypel5-/- mutant zebrafish, as revealed by metabolomic and transcriptomic analyses. Mechanistically, Hnf4a is identified as a crucial downstream mediator that is positively regulated by Ypel5. Zebrafish hnf4a overexpression could largely rescue ypel5 deficiency-induced hepatic defects. Furthermore, PPARα signaling mediates the regulation of Hnf4a by Ypel5 through directly binding to the transcriptional enhancer of the Hnf4a gene. Herein, this work demonstrates an essential role of Ypel5 in hepatocyte proliferation and function and provides the first in vivo evidence for a physiological role of the ypel5 gene in vertebrates.
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Affiliation(s)
- Yun Deng
- Shanghai Institute of Hematology, State Key Laboratory of Medical Genomics, National Research Center for Translational Medicine at Shanghai, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
- CNRS-LIA Hematology and Cancer, Sino-French Research Center for Life Sciences and Genomics, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Xiao Han
- Shanghai Institute of Hematology, State Key Laboratory of Medical Genomics, National Research Center for Translational Medicine at Shanghai, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
- CNRS-LIA Hematology and Cancer, Sino-French Research Center for Life Sciences and Genomics, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Huiqiao Chen
- Department of Hematology, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310011, China
| | - Chaoxian Zhao
- Shanghai Institute of Hematology, State Key Laboratory of Medical Genomics, National Research Center for Translational Medicine at Shanghai, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
- CNRS-LIA Hematology and Cancer, Sino-French Research Center for Life Sciences and Genomics, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Yi Chen
- Shanghai Institute of Hematology, State Key Laboratory of Medical Genomics, National Research Center for Translational Medicine at Shanghai, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Jun Zhou
- Shanghai Institute of Hematology, State Key Laboratory of Medical Genomics, National Research Center for Translational Medicine at Shanghai, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
- CNRS-LIA Hematology and Cancer, Sino-French Research Center for Life Sciences and Genomics, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Hugues de The
- CNRS-LIA Hematology and Cancer, Sino-French Research Center for Life Sciences and Genomics, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
- Université de Paris 7/INSERM/CNRS UMR 944/7212, Equipe Labellisée Ligue Nationale Contre le Cancer, Hôpital St. Louis, Paris 75010, France
| | - Jun Zhu
- CNRS-LIA Hematology and Cancer, Sino-French Research Center for Life Sciences and Genomics, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
- Université de Paris 7/INSERM/CNRS UMR 944/7212, Equipe Labellisée Ligue Nationale Contre le Cancer, Hôpital St. Louis, Paris 75010, France
| | - Hao Yuan
- Shanghai Institute of Hematology, State Key Laboratory of Medical Genomics, National Research Center for Translational Medicine at Shanghai, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
- CNRS-LIA Hematology and Cancer, Sino-French Research Center for Life Sciences and Genomics, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
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17
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Abstract
Hepatocyte nuclear factor 4 α (HNF4α) is a highly conserved member of the nuclear receptor superfamily expressed at high levels in the liver, kidney, pancreas, and gut. In the liver, HNF4α is exclusively expressed in hepatocytes, where it is indispensable for embryonic and postnatal liver development and for normal liver function in adults. It is considered a master regulator of hepatic differentiation because it regulates a significant number of genes involved in hepatocyte-specific functions. Loss of HNF4α expression and function is associated with the progression of chronic liver disease. Further, HNF4α is a target of chemical-induced liver injury. In this review, we discuss the role of HNF4α in liver pathophysiology and highlight its potential use as a therapeutic target for liver diseases.
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Affiliation(s)
- Manasi Kotulkar
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas
| | - Dakota R Robarts
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas
| | - Udayan Apte
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas
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18
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Shah A, Huck I, Duncan K, Gansemer ER, Apte U, Stamnes MA, Rutkowski DT. Interference with the HNF4-dependent gene regulatory network diminishes ER stress in hepatocytes. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.02.09.527889. [PMID: 36798396 PMCID: PMC9934629 DOI: 10.1101/2023.02.09.527889] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 02/11/2023]
Abstract
In all eukaryotic cell types, the unfolded protein response (UPR) upregulates factors that promote protein folding and misfolded protein clearance to help alleviate endoplasmic reticulum (ER) stress. Yet ER stress in the liver is uniquely accompanied by the suppression of metabolic genes, the coordination and purpose of which is largely unknown. Here, we used unsupervised machine learning to identify a cluster of correlated genes that were profoundly suppressed by persistent ER stress in the liver. These genes, which encode diverse functions including metabolism, coagulation, drug detoxification, and bile synthesis, are likely targets of the master regulator of hepatocyte differentiation HNF4α. The response of these genes to ER stress was phenocopied by liver-specific deletion of HNF4 α. Strikingly, while deletion of HNF4α exacerbated liver injury in response to an ER stress challenge, it also diminished UPR activation and partially preserved ER ultrastructure, suggesting attenuated ER stress. Conversely, pharmacological maintenance of hepatocyte identity in vitro enhanced sensitivity to stress. Several pathways potentially link HNF4α to ER stress sensitivity, including control of expression of the tunicamycin transporter MFSD2A; modulation of IRE1/XBP1 signaling; and regulation of Pyruvate Dehydrogenase. Together, these findings suggest that HNF4α activity is linked to hepatic ER homeostasis through multiple mechanisms.
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Affiliation(s)
- Anit Shah
- Department of Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, IA
| | - Ian Huck
- Department of Pharmacology, Toxicology, and Therapeutics, Kansas University Medical Center, Kansas City, KS
| | - Kaylia Duncan
- Department of Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, IA
| | - Erica R. Gansemer
- Department of Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, IA
| | - Udayan Apte
- Department of Pharmacology, Toxicology, and Therapeutics, Kansas University Medical Center, Kansas City, KS
| | - Mark A. Stamnes
- Department of Molecular Physiology and Biophysics, University of Iowa Carver College of Medicine, Iowa City, IA
| | - D. Thomas Rutkowski
- Department of Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, IA
- Department of Internal Medicine, University of Iowa Carver College of Medicine, Iowa City, IA
- Department of Fraternal Order of Eagles Diabetes Research Center, University of Iowa Carver College of Medicine, Iowa City, IA
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19
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Robarts DR, Paine-Cabrera D, Kotulkar M, Venneman KK, Gunewardena S, Corton JC, Lau C, Foquet L, Bial G, Apte U. Identifying Human Specific Adverse Outcome Pathways of Per- and Polyfluoroalkyl Substances Using Liver-Chimeric Humanized Mice. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.02.01.526711. [PMID: 36778348 PMCID: PMC9915685 DOI: 10.1101/2023.02.01.526711] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Background Per- and polyfluoroalkyl substances (PFAS) are persistent organic pollutants with myriad adverse effects. While perfluorooctanoic acid (PFOA) and perfluorooctane sulfonic acid (PFOS) are the most common contaminants, levels of replacement PFAS, such as perfluoro-2-methyl-3-oxahexanoic acid (GenX), are increasing. In rodents, PFOA, PFOS, and GenX have several adverse effects on the liver, including nonalcoholic fatty liver disease. Objective We aimed to determine human-relevant mechanisms of PFAS induced adverse hepatic effects using FRG liver-chimeric humanized mice with livers repopulated with functional human hepatocytes. Methods Male humanized mice were treated with 0.067 mg/L of PFOA, 0.145 mg/L of PFOS, or 1 mg/L of GenX in drinking water for 28 days. Liver and serum were collected for pathology and clinical chemistry, respectively. RNA-sequencing coupled with pathway analysis was used to determine molecular mechanisms. Results PFOS caused a significant decrease in total serum cholesterol and LDL/VLDL, whereas GenX caused a significant elevation in LDL/VLDL with no change in total cholesterol and HDL. PFOA had no significant changes in serum LDL/VLDL and total cholesterol. All three PFAS induced significant hepatocyte proliferation. RNA-sequencing with alignment to the human genome showed a total of 240, 162, and 619 differentially expressed genes after PFOA, PFOS, and GenX exposure, respectively. Upstream regulator analysis revealed inhibition of NR1D1, a transcriptional repressor important in circadian rhythm, as the major common molecular change in all PFAS treatments. PFAS treated mice had significant nuclear localization of NR1D1. In silico modeling showed PFOA, PFOS, and GenX potentially interact with the DNA-binding domain of NR1D1. Discussion These data implicate PFAS in circadian rhythm disruption via inhibition of NR1D1. These studies show that FRG humanized mice are a useful tool for studying the adverse outcome pathways of environmental pollutants on human hepatocytes in situ.
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Affiliation(s)
- Dakota R. Robarts
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS
| | - Diego Paine-Cabrera
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS
| | - Manasi Kotulkar
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS
| | - Kaitlyn K. Venneman
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS
| | - Sumedha Gunewardena
- Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, KS
| | - J. Christopher Corton
- Center for Computational Toxicology and Exposure, Office of Research and Development, U.S. EPA, Research Triangle Park, NC
| | - Christopher Lau
- Center for Public Health and Environmental Assessment, Office of Research and Development, US EPA, Research Triangle Park, NC
| | | | | | - Udayan Apte
- Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS
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20
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Fekry B, Ribas-Latre A, Drunen RV, Santos RB, Shivshankar S, Dai Y, Zhao Z, Yoo SH, Chen Z, Sun K, Sladek FM, Younes M, Eckel-Mahan K. Hepatic circadian and differentiation factors control liver susceptibility for fatty liver disease and tumorigenesis. FASEB J 2022; 36:e22482. [PMID: 35947136 PMCID: PMC10062014 DOI: 10.1096/fj.202101398r] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2021] [Revised: 07/06/2022] [Accepted: 07/21/2022] [Indexed: 11/11/2022]
Abstract
Hepatocellular carcinoma (HCC) is a leading cause of cancer deaths, and the most common primary liver malignancy to present in the clinic. With the exception of liver transplant, treatment options for advanced HCC are limited, but improved tumor stratification could open the door to new treatment options. Previously, we demonstrated that the circadian regulator Aryl Hydrocarbon-Like Receptor Like 1 (ARNTL, or Bmal1) and the liver-enriched nuclear factor 4 alpha (HNF4α) are robustly co-expressed in healthy liver but incompatible in the context of HCC. Faulty circadian expression of HNF4α- either by isoform switching, or loss of expression- results in an increased risk for HCC, while BMAL1 gain-of-function in HNF4α-positive HCC results in apoptosis and tumor regression. We hypothesize that the transcriptional programs of HNF4α and BMAL1 are antagonistic in liver disease and HCC. Here, we study this antagonism by generating a mouse model with inducible loss of hepatic HNF4α and BMAL1 expression. The results reveal that simultaneous loss of HNF4α and BMAL1 is protective against fatty liver and HCC in carcinogen-induced liver injury and in the "STAM" model of liver disease. Furthermore, our results suggest that targeting Bmal1 expression in the absence of HNF4α inhibits HCC growth and progression. Specifically, pharmacological suppression of Bmal1 in HNF4α-deficient, BMAL1-positive HCC with REV-ERB agonist SR9009 impairs tumor cell proliferation and migration in a REV-ERB-dependent manner, while having no effect on healthy hepatocytes. Collectively, our results suggest that stratification of HCC based on HNF4α and BMAL1 expression may provide a new perspective on HCC properties and potential targeted therapeutics.
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Affiliation(s)
- Baharan Fekry
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA
| | - Aleix Ribas-Latre
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA
| | - Rachel Van Drunen
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA
| | - Rafael Bravo Santos
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA
| | - Samay Shivshankar
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA
| | - Yulin Dai
- Center for Precision Health, School of Biomedical Informatics, The University of Texas Health Science Center, Houston, Texas, USA
| | - Zhongming Zhao
- Center for Precision Health, School of Biomedical Informatics, The University of Texas Health Science Center, Houston, Texas, USA.,Human Genetics Center, School of Public Health, The University of Texas Health Science Center, Houston, Texas, USA
| | - Seung-Hee Yoo
- Department of Biochemistry and Molecular Biology, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA
| | - Zheng Chen
- Department of Biochemistry and Molecular Biology, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA
| | - Kai Sun
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA.,Department of Integrative Biology and Pharmacology, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA
| | - Frances M Sladek
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, California, USA
| | - Mamoun Younes
- Department of Pathology, George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, USA
| | - Kristin Eckel-Mahan
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA.,Department of Integrative Biology and Pharmacology, McGovern Medical School at the University of Texas Health Science Center, Houston, Texas, USA
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21
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Gunewardena S, Huck I, Walesky C, Robarts D, Weinman S, Apte U. Progressive loss of hepatocyte nuclear factor 4 alpha activity in chronic liver diseases in humans. Hepatology 2022; 76:372-386. [PMID: 35006629 PMCID: PMC9762158 DOI: 10.1002/hep.32326] [Citation(s) in RCA: 29] [Impact Index Per Article: 9.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/11/2021] [Revised: 12/15/2021] [Accepted: 12/20/2021] [Indexed: 12/16/2022]
Abstract
BACKGROUND AND AIMS Hepatocyte nuclear factor 4 alpha (HNF4α) is indispensable for hepatocyte differentiation and critical for maintaining liver health. Here, we demonstrate that loss of HNF4α activity is a crucial step in the pathogenesis of chronic liver diseases (CLDs) that lead to development of HCC. APPROACH AND RESULTS We developed an HNF4α target gene signature, which can accurately determine HNF4α activity, and performed an exhaustive in silico analysis using hierarchical and K-means clustering, survival, and rank-order analysis of 30 independent data sets containing over 3500 individual samples. The association of changes in HNF4α activity to CLD progression of various etiologies, including HCV- and HBV-induced liver cirrhosis (LC), NAFLD/NASH, and HCC, was determined. Results revealed a step-wise reduction in HNF4α activity with each progressive stage of pathogenesis. Cluster analysis of LC gene expression data sets using the HNF4α signature showed that loss of HNF4α activity was associated with progression of Child-Pugh class, faster decompensation, incidence of HCC, and lower survival with and without HCC. A moderate decrease in HNF4α activity was observed in NAFLD from normal liver, but a further significant decline was observed in patients from NAFLD to NASH. In HCC, loss of HNF4α activity was associated with advanced disease, increased inflammatory changes, portal vein thrombosis, and substantially lower survival. CONCLUSIONS In conclusion, these data indicate that loss of HNF4α function is a common event in the pathogenesis of CLDs leading to HCC and is important from both diagnostic and therapeutic standpoints.
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Affiliation(s)
- Sumedha Gunewardena
- Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, KS
| | - Ian Huck
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS
| | - Chad Walesky
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS
| | - Dakota Robarts
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS
| | - Steven Weinman
- Department of Medicine, University of Kansas Medical Center, Kansas City, KS
| | - Udayan Apte
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS
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22
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Li G, Li X, Yang L, Wang S, Dai Y, Fekry B, Veillon L, Tan L, Berdeaux R, Eckel-Mahan K, Lorenzi PL, Zhao Z, Lehner R, Sun K. Adipose tissue-specific ablation of Ces1d causes metabolic dysregulation in mice. Life Sci Alliance 2022; 5:e202101209. [PMID: 35459739 PMCID: PMC9034061 DOI: 10.26508/lsa.202101209] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2021] [Revised: 04/08/2022] [Accepted: 04/11/2022] [Indexed: 01/25/2023] Open
Abstract
Carboxylesterase 1d (Ces1d) is a crucial enzyme with a wide range of activities in multiple tissues. It has been reported to localize predominantly in ER. Here, we found that Ces1d levels are significantly increased in obese patients with type 2 diabetes. Intriguingly, a high level of Ces1d translocates onto lipid droplets where it digests the lipids to produce a unique set of fatty acids. We further revealed that adipose tissue-specific Ces1d knock-out (FKO) mice gained more body weight with increased fat mass during a high fat-diet challenge. The FKO mice exhibited impaired glucose and lipid metabolism and developed exacerbated liver steatosis. Mechanistically, deficiency of Ces1d induced abnormally large lipid droplet deposition in the adipocytes, causing ectopic accumulation of triglycerides in other peripheral tissues. Furthermore, loss of Ces1d diminished the circulating free fatty acids serving as signaling molecules to trigger the epigenetic regulations of energy metabolism via lipid-sensing transcriptional factors, such as HNF4α. The metabolic disorders induced an unhealthy microenvironment in the metabolically active tissues, ultimately leading to systemic insulin resistance.
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Affiliation(s)
- Gang Li
- Center for Metabolic and Degenerative Diseases, The Brown Foundation Institute of Molecular Medicine for the Prevention of Human Diseases, University of Texas Health Science Center at Houston, Houston, TX, USA
| | - Xin Li
- Center for Metabolic and Degenerative Diseases, The Brown Foundation Institute of Molecular Medicine for the Prevention of Human Diseases, University of Texas Health Science Center at Houston, Houston, TX, USA
| | - Li Yang
- Center for Metabolic and Degenerative Diseases, The Brown Foundation Institute of Molecular Medicine for the Prevention of Human Diseases, University of Texas Health Science Center at Houston, Houston, TX, USA
| | - Shuyue Wang
- Center for Metabolic and Degenerative Diseases, The Brown Foundation Institute of Molecular Medicine for the Prevention of Human Diseases, University of Texas Health Science Center at Houston, Houston, TX, USA
| | - Yulin Dai
- Center for Precision Health, School of Biomedical Informatics, University of Texas Health Science Center at Houston, Houston, TX, USA
| | - Baharan Fekry
- Center for Metabolic and Degenerative Diseases, The Brown Foundation Institute of Molecular Medicine for the Prevention of Human Diseases, University of Texas Health Science Center at Houston, Houston, TX, USA
| | - Lucas Veillon
- Metabolomic Core Facility, Department of Bioinformatics and Computational Biology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Lin Tan
- Metabolomic Core Facility, Department of Bioinformatics and Computational Biology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Rebecca Berdeaux
- Center for Metabolic and Degenerative Diseases, The Brown Foundation Institute of Molecular Medicine for the Prevention of Human Diseases, University of Texas Health Science Center at Houston, Houston, TX, USA
- Department of Integrative Biology and Pharmacology, The University of Texas Health Science Center at Houston, Houston, TX, USA
- Program in Biochemistry and Cell Biology, MD Anderson Cancer Center-UTHealth Graduate School of Biomedical Sciences, Houston, TX, USA
| | - Kristin Eckel-Mahan
- Center for Metabolic and Degenerative Diseases, The Brown Foundation Institute of Molecular Medicine for the Prevention of Human Diseases, University of Texas Health Science Center at Houston, Houston, TX, USA
- Department of Integrative Biology and Pharmacology, The University of Texas Health Science Center at Houston, Houston, TX, USA
- Program in Biochemistry and Cell Biology, MD Anderson Cancer Center-UTHealth Graduate School of Biomedical Sciences, Houston, TX, USA
| | - Philip L Lorenzi
- Metabolomic Core Facility, Department of Bioinformatics and Computational Biology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Zhongming Zhao
- Center for Precision Health, School of Biomedical Informatics, University of Texas Health Science Center at Houston, Houston, TX, USA
| | - Richard Lehner
- Group on Molecular and Cell Biology of Lipids, Department of Pediatrics, University of Alberta, Edmonton, Canada
| | - Kai Sun
- Center for Metabolic and Degenerative Diseases, The Brown Foundation Institute of Molecular Medicine for the Prevention of Human Diseases, University of Texas Health Science Center at Houston, Houston, TX, USA
- Department of Integrative Biology and Pharmacology, The University of Texas Health Science Center at Houston, Houston, TX, USA
- Program in Biochemistry and Cell Biology, MD Anderson Cancer Center-UTHealth Graduate School of Biomedical Sciences, Houston, TX, USA
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23
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Robarts DR, Venneman KK, Gunewardena S, Apte U. GenX induces fibroinflammatory gene expression in primary human hepatocytes. Toxicology 2022; 477:153259. [PMID: 35850385 PMCID: PMC9741548 DOI: 10.1016/j.tox.2022.153259] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/16/2022] [Revised: 07/12/2022] [Accepted: 07/14/2022] [Indexed: 01/09/2023]
Abstract
The toxicity induced by the persistent organic pollutants per- and polyfluoroalkyl substances (PFAS) is dependent on the length of their polyfluorinated tail. Long-chain PFASs have significantly longer half-lives and profound toxic effects compared to their short-chain counterparts. Recently, production of a short-chain PFAS substitute called ammonium 2,3,3,3-tetrafluoro-2-(heptafluoropropoxy) propanoate, also known as GenX, has significantly increased. However, the adverse health effects of GenX are not completely known. In this study, we investigated the dose-dependent effects of GenX on primary human hepatocytes (PHH). Freshly isolated PHH were treated with either 0.1, 10, or 100 μM of GenX for 48 and 96 h; then, global transcriptomic changes were determined using Human Clariom™ D arrays. GenX-induced transcriptional changes were similar at 0.1 and 10 μM doses but were significantly different at the 100 μM dose. Genes involved in lipid, monocarboxylic acid, and ketone metabolism were significantly altered following exposure of PHH at all doses. However, at the 100 μM dose, GenX caused changes in genes involved in cell proliferation, inflammation and fibrosis. A correlation analysis of concentration and differential gene expression revealed that 576 genes positively (R > 0.99) and 375 genes negatively (R < -0.99) correlated with GenX concentration. The upstream regulator analysis indicated HIF1α was inhibited at the lower doses but were activated at the higher dose. Additionally, VEGF, PPARα, STAT3, and SMAD4 signaling was induced at the 100 µM dose. These data indicate that at lower doses GenX can interfere with metabolic pathways and at higher doses can induce fibroinflammatory changes in human hepatocytes.
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Affiliation(s)
- Dakota R Robarts
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA.
| | - Kaitlyn K Venneman
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA.
| | - Sumedha Gunewardena
- Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, KS, USA.
| | - Udayan Apte
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA.
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24
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Schonfeld M, Averilla J, Gunewardena S, Weinman SA, Tikhanovich I. Male-Specific Activation of Lysine Demethylases 5B and 5C Mediates Alcohol-Induced Liver Injury and Hepatocyte Dedifferentiation. Hepatol Commun 2022; 6:1373-1391. [PMID: 35084807 PMCID: PMC9134811 DOI: 10.1002/hep4.1895] [Citation(s) in RCA: 14] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/29/2021] [Revised: 12/13/2021] [Accepted: 12/22/2021] [Indexed: 12/11/2022] Open
Abstract
Alcohol-associated liver disease (ALD) is a major cause of alcohol-related mortality. Sex differences in sensitivity to ALD are well described, but these are often disregarded in studies of ALD development. We aimed to define sex-specific pathways in liver exposed to alcohol. Mice were fed the Lieber-DeCarli alcohol liquid diet or a combination of a high-fat diet with alcohol in water. Single-cell RNA sequencing (scRNA-Seq) was performed on liver cells from male and female mice. Mice were treated with adeno-associated virus (AAV)-short hairpin (sh)Control or AAV-sh lysine demethylase 5b (shKdm5b) and/or AAV-shKdm5c vectors. Changes after Kdm5b/5c knockdown were assessed by RNA-Seq and histone H3 lysine K4 (H3K4)me3 chromatin immunoprecipitation-Seq analysis. Using scRNA-Seq analysis, we found several sex-specific pathways induced by alcohol, including pathways related to lipid metabolism and hepatocyte differentiation. Bioinformatic analysis suggested that two epigenetic regulators, H3K4-specific lysine demethylases KDM5B and KDM5C, contribute to sex differences in alcohol effects. We found that in alcohol-fed male mice, KDM5B and KDM5C are involved in hepatocyte nuclear factor 4 alpha (Hnf4a) down-regulation, hepatocyte dedifferentiation, and an increase in fatty acid synthesis. This effect is mediated by alcohol-induced KDM5B and KDM5C recruitment to Hnf4a and other gene promoters in male but not in female mice. Kdm5b and Kdm5c knockdown or KDM5-inhibitor treatment prevented alcohol-induced lipid accumulation and restored levels of Hnf4a and other hepatocyte differentiation genes in male mice. In addition, Kdm5b knockdown prevented hepatocellular carcinoma development in male mice by up-regulating Hnf4a and decreasing tumor cell proliferation. Conclusion: Alcohol specifically activates KDM5 demethylases in male mice to promote alcohol-induced hepatocyte dedifferentiation and tumor development.
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Affiliation(s)
- Michael Schonfeld
- Department of Internal MedicineUniversity of Kansas Medical CenterKansas CityKSUSA
| | - Janice Averilla
- Department of Internal MedicineUniversity of Kansas Medical CenterKansas CityKSUSA
| | - Sumedha Gunewardena
- Department of Molecular and Integrative PhysiologyUniversity of Kansas Medical CenterKansas CityKSUSA
| | - Steven A. Weinman
- Department of Internal MedicineUniversity of Kansas Medical CenterKansas CityKSUSA
- Liver CenterUniversity of Kansas Medical CenterKansas CityKSUSA
- Kansas City VA Medical CenterKansas CityMOUSA
| | - Irina Tikhanovich
- Department of Internal MedicineUniversity of Kansas Medical CenterKansas CityKSUSA
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25
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Liu P, Jiang L, Kong W, Xie Q, Li P, Liu X, Zhang J, Liu M, Wang Z, Zhu L, Yang H, Zhou Y, Zou J, Liu X, Liu L. PXR activation impairs hepatic glucose metabolism partly via inhibiting the HNF4 α-GLUT2 pathway. Acta Pharm Sin B 2022; 12:2391-2405. [PMID: 35646519 PMCID: PMC9136535 DOI: 10.1016/j.apsb.2021.09.031] [Citation(s) in RCA: 19] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/14/2021] [Revised: 09/05/2021] [Accepted: 09/16/2021] [Indexed: 01/20/2023] Open
Abstract
Drug-induced hyperglycemia/diabetes is a global issue. Some drugs induce hyperglycemia by activating the pregnane X receptor (PXR), but the mechanism is unclear. Here, we report that PXR activation induces hyperglycemia by impairing hepatic glucose metabolism due to inhibition of the hepatocyte nuclear factor 4-alpha (HNF4α)‒glucose transporter 2 (GLUT2) pathway. The PXR agonists atorvastatin and rifampicin significantly downregulated GLUT2 and HNF4α expression, and impaired glucose uptake and utilization in HepG2 cells. Overexpression of PXR downregulated GLUT2 and HNF4α expression, while silencing PXR upregulated HNF4α and GLUT2 expression. Silencing HNF4α decreased GLUT2 expression, while overexpressing HNF4α increased GLUT2 expression and glucose uptake. Silencing PXR or overexpressing HNF4α reversed the atorvastatin-induced decrease in GLUT2 expression and glucose uptake. In human primary hepatocytes, atorvastatin downregulated GLUT2 and HNF4α mRNA expression, which could be attenuated by silencing PXR. Silencing HNF4α downregulated GLUT2 mRNA expression. These findings were reproduced with mouse primary hepatocytes. Hnf4α plasmid increased Slc2a2 promoter activity. Hnf4α silencing or pregnenolone-16α-carbonitrile (PCN) suppressed the Slc2a2 promoter activity by decreasing HNF4α recruitment to the Slc2a2 promoter. Liver-specific Hnf4α deletion and PCN impaired glucose tolerance and hepatic glucose uptake, and decreased the expression of hepatic HNF4α and GLUT2. In conclusion, PXR activation impaired hepatic glucose metabolism partly by inhibiting the HNF4α‒GLUT2 pathway. These results highlight the molecular mechanisms by which PXR activators induce hyperglycemia/diabetes.
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26
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Robarts DR, McGreal SR, Umbaugh DS, Parkes WS, Kotulkar M, Abernathy S, Lee N, Jaeschke H, Gunewardena S, Whelan SA, Hanover JA, Zachara NE, Slawson C, Apte U. Regulation of Liver Regeneration by Hepatocyte O-GlcNAcylation in Mice. Cell Mol Gastroenterol Hepatol 2022; 13:1510-1529. [PMID: 35093590 PMCID: PMC9043307 DOI: 10.1016/j.jcmgh.2022.01.014] [Citation(s) in RCA: 21] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/07/2021] [Revised: 01/18/2022] [Accepted: 01/19/2022] [Indexed: 12/15/2022]
Abstract
BACKGROUND & AIMS The liver has a unique capacity to regenerate after injury in a highly orchestrated and regulated manner. Here, we report that O-GlcNAcylation, an intracellular post-translational modification regulated by 2 enzymes, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), is a critical termination signal for liver regeneration following partial hepatectomy (PHX). METHODS We studied liver regeneration after PHX on hepatocyte specific OGT and OGA knockout mice (OGT-KO and OGA-KO), which caused a significant decrease (OGT-KO) and increase (OGA-KO) in hepatic O-GlcNAcylation, respectively. RESULTS OGA-KO mice had normal regeneration, but the OGT-KO mice exhibited substantial defects in termination of liver regeneration with increased liver injury, sustained cell proliferation resulting in significant hepatomegaly, hepatic dysplasia, and appearance of small nodules at 28 days after PHX. This was accompanied by a sustained increase in expression of cyclins along with significant induction in pro-inflammatory and pro-fibrotic gene expression in the OGT-KO livers. RNA-sequencing studies revealed inactivation of hepatocyte nuclear 4 alpha (HNF4α), the master regulator of hepatic differentiation and a known termination signal, in OGT-KO mice at 28 days after PHX, which was confirmed by both Western blot and immunohistochemistry analysis. Furthermore, a significant decrease in HNFα target genes was observed in OGT-KO mice, indicating a lack of hepatocyte differentiation following decreased hepatic O-GlcNAcylation. Immunoprecipitation experiments revealed HNF4α is O-GlcNAcylated in normal differentiated hepatocytes. CONCLUSIONS These studies show that O-GlcNAcylation plays a critical role in the termination of liver regeneration via regulation of HNF4α in hepatocytes.
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Affiliation(s)
- Dakota R Robarts
- Department of Pharmacology, Toxicology and Therapeutics, Kansas City, Kansas
| | - Steven R McGreal
- Department of Pharmacology, Toxicology and Therapeutics, Kansas City, Kansas
| | - David S Umbaugh
- Department of Pharmacology, Toxicology and Therapeutics, Kansas City, Kansas
| | - Wendena S Parkes
- Department of Pharmacology, Toxicology and Therapeutics, Kansas City, Kansas
| | - Manasi Kotulkar
- Department of Pharmacology, Toxicology and Therapeutics, Kansas City, Kansas
| | - Sarah Abernathy
- Department of Pharmacology, Toxicology and Therapeutics, Kansas City, Kansas
| | - Norman Lee
- Department of Chemistry, Boston University, Boston, Massachusetts
| | - Hartmut Jaeschke
- Department of Pharmacology, Toxicology and Therapeutics, Kansas City, Kansas
| | | | - Stephen A Whelan
- Department of Chemistry, Boston University, Boston, Massachusetts
| | - John A Hanover
- Laboratory of Cell Biochemistry and Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland
| | - Natasha E Zachara
- Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland
| | - Chad Slawson
- Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas
| | - Udayan Apte
- Department of Pharmacology, Toxicology and Therapeutics, Kansas City, Kansas.
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27
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Goel C, Monga SP, Nejak-Bowen K. Role and Regulation of Wnt/β-Catenin in Hepatic Perivenous Zonation and Physiological Homeostasis. THE AMERICAN JOURNAL OF PATHOLOGY 2022; 192:4-17. [PMID: 34924168 PMCID: PMC8747012 DOI: 10.1016/j.ajpath.2021.09.007] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/17/2021] [Revised: 09/02/2021] [Accepted: 09/22/2021] [Indexed: 01/03/2023]
Abstract
Metabolic heterogeneity or functional zonation is a key characteristic of the liver that allows different metabolic pathways to be spatially regulated within the hepatic system and together contribute to whole body homeostasis. These metabolic pathways are segregated along the portocentral axis of the liver lobule into three hepatic zones: periportal, intermediate or midzonal, and perivenous. The liver performs complementary or opposing metabolic functions within different hepatic zones while synergistic functions are regulated by overlapping zones, thereby maintaining the overall physiological stability. The Wnt/β-catenin signaling pathway is well known for its role in liver growth, development, and regeneration. In addition, the Wnt/β-catenin pathway plays a fundamental and dominant role in hepatic zonation and signals to orchestrate various functions of liver metabolism and pathophysiology. The β-catenin protein is the central player in the Wnt/β-catenin signaling cascade, and its activation is crucial for metabolic patterning of the liver. However, dysregulation of Wnt/β-catenin signaling is also implicated in different liver pathologies, including those associated with metabolic syndrome. β-Catenin is preferentially localized in the central region of the hepatic lobule surrounding the central vein and regulates multiple functions of this region. This review outlines the role of Wnt/β-catenin signaling pathway in controlling the different metabolic processes surrounding the central vein and its relation to liver homeostasis and dysfunction.
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Affiliation(s)
- Chhavi Goel
- Department of Pathology, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania
| | - Satdarshan P Monga
- Department of Pathology, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania; Department of Medicine, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania; Pittsburgh Liver Research Center, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania
| | - Kari Nejak-Bowen
- Department of Pathology, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania; Pittsburgh Liver Research Center, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania.
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28
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Bellido Molias F, Sim A, Leong KW, An O, Song Y, Ng VHE, Lim MWJ, Ying C, Teo JXJ, Göke J, Chen L. Antisense RNAs Influence Promoter Usage of Their Counterpart Sense Genes in Cancer. Cancer Res 2021; 81:5849-5861. [PMID: 34649947 PMCID: PMC9397637 DOI: 10.1158/0008-5472.can-21-1859] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/11/2021] [Revised: 08/19/2021] [Accepted: 10/11/2021] [Indexed: 01/07/2023]
Abstract
Multiple noncoding natural antisense transcripts (ncNAT) are known to modulate key biological events such as cell growth or differentiation. However, the actual impact of ncNATs on cancer progression remains largely unknown. In this study, we identified a complete list of differentially expressed ncNATs in hepatocellular carcinoma. Among them, a previously undescribed ncNAT HNF4A-AS1L suppressed cancer cell growth by regulating its sense gene HNF4A, a well-known cancer driver, through a promoter-specific mechanism. HNF4A-AS1L selectively activated the HNF4A P1 promoter via HNF1A, which upregulated expression of tumor suppressor P1-driven isoforms, while having no effect on the oncogenic P2 promoter. RNA-seq data from 23 tissue and cancer types identified approximately 100 ncNATs whose expression correlated specifically with the activity of one promoter of their associated sense gene. Silencing of two of these ncNATs ENSG00000259357 and ENSG00000255031 (antisense to CERS2 and CHKA, respectively) altered the promoter usage of CERS2 and CHKA. Altogether, these results demonstrate that promoter-specific regulation is a mechanism used by ncNATs for context-specific control of alternative isoform expression of their counterpart sense genes. SIGNIFICANCE: This study characterizes a previously unexplored role of ncNATs in regulation of isoform expression of associated sense genes, highlighting a mechanism of alternative promoter usage in cancer.
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Affiliation(s)
| | - Andre Sim
- Computational and Systems Biology, Genome Institute of Singapore, Singapore
| | - Ka Wai Leong
- Cancer Science Institute of Singapore, National University of Singapore, Singapore
| | - Omer An
- Cancer Science Institute of Singapore, National University of Singapore, Singapore
| | - Yangyang Song
- Cancer Science Institute of Singapore, National University of Singapore, Singapore
| | - Vanessa Hui En Ng
- Cancer Science Institute of Singapore, National University of Singapore, Singapore
| | - Max Wei Jie Lim
- Cancer Science Institute of Singapore, National University of Singapore, Singapore
| | - Chen Ying
- Computational and Systems Biology, Genome Institute of Singapore, Singapore
| | - Jasmin Xin Jia Teo
- Cancer Science Institute of Singapore, National University of Singapore, Singapore
| | - Jonathan Göke
- Computational and Systems Biology, Genome Institute of Singapore, Singapore.,Corresponding Authors: Leilei Chen, National University of Singapore, Center for Translational Medicine (MD6), 14 Medical Drive, #12-01, S117599 Singapore. Phone: 65-6516-8435; Fax: 65-6516-1873; E-mail: ; and Jonathan Göke,
| | - Leilei Chen
- Cancer Science Institute of Singapore, National University of Singapore, Singapore.,Department of Anatomy, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.,NUS Centre for Cancer Research, Yong Loo Lin School of Medicine, National University Singapore, Singapore.,Corresponding Authors: Leilei Chen, National University of Singapore, Center for Translational Medicine (MD6), 14 Medical Drive, #12-01, S117599 Singapore. Phone: 65-6516-8435; Fax: 65-6516-1873; E-mail: ; and Jonathan Göke,
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Gamboa CM, Wang Y, Xu H, Kalemba K, Wondisford FE, Sabaawy HE. Optimized 3D Culture of Hepatic Cells for Liver Organoid Metabolic Assays. Cells 2021; 10:cells10123280. [PMID: 34943788 PMCID: PMC8699701 DOI: 10.3390/cells10123280] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2021] [Revised: 11/16/2021] [Accepted: 11/19/2021] [Indexed: 12/25/2022] Open
Abstract
The liver is among the principal organs for glucose homeostasis and metabolism. Studies of liver metabolism are limited by the inability to expand primary hepatocytes in vitro while maintaining their metabolic functions. Human hepatic three-dimensional (3D) organoids have been established using defined factors, yet hepatic organoids from adult donors showed impaired expansion. We examined conditions to facilitate the expansion of adult donor-derived hepatic organoids (HepAOs) and HepG2 cells in organoid cultures (HepGOs) using combinations of growth factors and small molecules. The expansion dynamics, gluconeogenic and HNF4α expression, and albumin secretion are assessed. The conditions tested allow the generation of HepAOs and HepGOs in 3D cultures. Nevertheless, gluconeogenic gene expression varies greatly between conditions. The organoid expansion rates are limited when including the TGFβ inhibitor A8301, while are relatively higher with Forskolin (FSK) and Oncostatin M (OSM). Notably, expanded HepGOs grown in the optimized condition maintain detectable gluconeogenic expression in a spatiotemporal distribution at 8 weeks. We present optimized conditions by limiting A8301 and incorporating FSK and OSM to allow the expansion of HepAOs from adult donors and HepGOs with gluconeogenic competence. These models increase the repertoire of human hepatic cellular tools available for use in liver metabolic assays.
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Affiliation(s)
- Christian Moya Gamboa
- Rutgers Cancer Institute of New Jersey, Rutgers University, New Brunswick, NJ 08901, USA;
| | - Yujue Wang
- Department of Medicine, Robert Wood Johnson Medical School, Rutgers University, New Brunswick, NJ 08901, USA; (Y.W.); (H.X.); (K.K.)
| | - Huiting Xu
- Department of Medicine, Robert Wood Johnson Medical School, Rutgers University, New Brunswick, NJ 08901, USA; (Y.W.); (H.X.); (K.K.)
| | - Katarzyna Kalemba
- Department of Medicine, Robert Wood Johnson Medical School, Rutgers University, New Brunswick, NJ 08901, USA; (Y.W.); (H.X.); (K.K.)
| | - Fredric E. Wondisford
- Rutgers Cancer Institute of New Jersey, Rutgers University, New Brunswick, NJ 08901, USA;
- Department of Medicine, Robert Wood Johnson Medical School, Rutgers University, New Brunswick, NJ 08901, USA; (Y.W.); (H.X.); (K.K.)
- Correspondence: (F.E.W.); (H.E.S.); Tel.: +1-732-235-9838 (F.E.W.); +1-732-235-8081 (H.E.S.)
| | - Hatem E. Sabaawy
- Rutgers Cancer Institute of New Jersey, Rutgers University, New Brunswick, NJ 08901, USA;
- Department of Medicine, Robert Wood Johnson Medical School, Rutgers University, New Brunswick, NJ 08901, USA; (Y.W.); (H.X.); (K.K.)
- Department of Pathology and Laboratory Medicine, RBHS-Robert Wood Johnson Medical School, Rutgers University, New Brunswick, NJ 08901, USA
- Correspondence: (F.E.W.); (H.E.S.); Tel.: +1-732-235-9838 (F.E.W.); +1-732-235-8081 (H.E.S.)
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Teeli AS, Łuczyńska K, Haque E, Gayas MA, Winiarczyk D, Taniguchi H. Disruption of Tumor Suppressors HNF4α/HNF1α Causes Tumorigenesis in Liver. Cancers (Basel) 2021; 13:cancers13215357. [PMID: 34771521 PMCID: PMC8582545 DOI: 10.3390/cancers13215357] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/13/2021] [Revised: 10/14/2021] [Accepted: 10/18/2021] [Indexed: 12/18/2022] Open
Abstract
The hepatocyte nuclear factor-4α (HNF4α) and hepatocyte nuclear factor-1α (HNF1α) are transcription factors that influence the development and maintenance of homeostasis in a variety of tissues, including the liver. As such, disruptions in their transcriptional networks can herald a number of pathologies, such as tumorigenesis. Largely considered tumor suppressants in liver cancer, these transcription factors regulate key events of inflammation, epithelial-mesenchymal transition, metabolic reprogramming, and the differentiation status of the cell. High-throughput analysis of cancer cell genomes has identified a number of hotspot mutations in HNF1α and HNF4α in liver cancer. Such results also showcase HNF1α and HNF4α as important therapeutic targets helping us step into the era of personalized medicine. In this review, we update current findings on the roles of HNF1α and HNF4α in liver cancer development and progression. It covers the molecular mechanisms of HNF1α and HNF4α dysregulation and also highlights the potential of HNF4α as a therapeutic target in liver cancer.
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Affiliation(s)
- Aamir Salam Teeli
- Institute of Genetics and Animal Biotechnology of the Polish Academy of Sciences, 05-552 Jastrzebiec, Poland; (A.S.T.); (K.Ł.); (E.H.); (D.W.)
| | - Kamila Łuczyńska
- Institute of Genetics and Animal Biotechnology of the Polish Academy of Sciences, 05-552 Jastrzebiec, Poland; (A.S.T.); (K.Ł.); (E.H.); (D.W.)
| | - Effi Haque
- Institute of Genetics and Animal Biotechnology of the Polish Academy of Sciences, 05-552 Jastrzebiec, Poland; (A.S.T.); (K.Ł.); (E.H.); (D.W.)
| | - Mohmmad Abrar Gayas
- Department of Surgery and Radiology, Faculty of Veterinary Sciences and Animal Husbandry, SKUAST-K, Jammu 19000, India;
| | - Dawid Winiarczyk
- Institute of Genetics and Animal Biotechnology of the Polish Academy of Sciences, 05-552 Jastrzebiec, Poland; (A.S.T.); (K.Ł.); (E.H.); (D.W.)
| | - Hiroaki Taniguchi
- Institute of Genetics and Animal Biotechnology of the Polish Academy of Sciences, 05-552 Jastrzebiec, Poland; (A.S.T.); (K.Ł.); (E.H.); (D.W.)
- Correspondence:
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Diaz-Aragon R, Coard MC, Amirneni S, Faccioli L, Haep N, Malizio MR, Motomura T, Kocas-Kilicarslan ZN, Ostrowska A, Florentino RM, Frau C. Therapeutic Potential of HNF4α in End-stage Liver Disease. Organogenesis 2021; 17:126-135. [PMID: 35114889 DOI: 10.1080/15476278.2021.1994273] [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: 10/25/2022] Open
Abstract
The prevalence of end-stage liver disease (ESLD) in the US is increasing at an alarming rate. It can be caused by several factors; however, one of the most common routes begins with nonalcoholic fatty liver disease (NAFLD). ESLD is diagnosed by the presence of irreversible damage to the liver. Currently, the only definitive treatment for ESLD is orthotopic liver transplantation (OLT). Nevertheless, OLT is limited due to a shortage of donor livers. Several promising alternative treatment options are under investigation. Researchers have focused on the effect of liver-enriched transcription factors (LETFs) on disease progression. Specifically, hepatocyte nuclear factor 4-alpha (HNF4α) has been reported to reset the liver transcription network and possibly play a role in the regression of fibrosis and cirrhosis. In this review, we describe the function of HNF4α, along with its regulation at various levels. In addition, we summarize the role of HNF4α in ESLD and its potential as a therapeutic target in the treatment of ESLD.
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Affiliation(s)
- Ricardo Diaz-Aragon
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
| | - Michael C Coard
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
| | - Sriram Amirneni
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
| | - Lanuza Faccioli
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
| | - Nils Haep
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
| | - Michelle R Malizio
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
| | - Takashi Motomura
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
| | | | - Alina Ostrowska
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA.,Pittsburgh Liver Research Center, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Rodrigo M Florentino
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
| | - Carla Frau
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
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Schonfeld M, O’Neil M, Villar MT, Artigues A, Averilla J, Gunewardena S, Weinman SA, Tikhanovich I. A Western diet with alcohol in drinking water recapitulates features of alcohol-associated liver disease in mice. Alcohol Clin Exp Res 2021; 45:1980-1993. [PMID: 34523155 PMCID: PMC9006178 DOI: 10.1111/acer.14700] [Citation(s) in RCA: 22] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2021] [Revised: 08/17/2021] [Accepted: 08/18/2021] [Indexed: 12/13/2022]
Abstract
BACKGROUND Mouse models of alcohol-associated liver disease vary greatly in their ease of implementation and the pathology they produce. Effects range from steatosis and mild inflammation with the Lieber-DeCarli liquid diet to severe inflammation, fibrosis, and pyroptosis seen with the Tsukamoto-French intragastric feeding model. Implementation of all of these models is limited by the labor-intensive nature of the protocols and the specialized skills necessary for successful intragastric feeding. We thus sought to develop a new model to reproduce features of alcohol-induced inflammation and fibrosis with minimal operational requirements. METHODS Over a 16-week period, mice were fed ad libitum with a pelleted high-fat Western diet (WD; 40% calories from fat) and alcohol added to the drinking water. We found the optimal alcohol consumption to be that at which the alcohol concentration was 20% for 4 days and 10% for 3 days per week. Control mice received WD pellets with water alone. RESULTS Alcohol consumption was 18 to 20 g/kg/day in males and 20 to 22 g/kg/day in females. Mice in the alcohol groups developed elevated serum transaminase levels after 12 weeks in males and 10 weeks in females. At 16 weeks, both males and females developed liver inflammation, steatosis, and pericellular fibrosis. Control mice on WD without alcohol had mild steatosis only. Alcohol-fed mice showed reduced HNF4α mRNA and protein expression. HNF4α is a master regulator of hepatocyte differentiation, down-regulation of which is a known driver of hepatocellular failure in alcoholic hepatitis. CONCLUSION A simple-to-administer, 16-week WD alcohol model recapitulates the inflammatory, fibrotic, and gene expression aspects of human alcohol-associated steatohepatitis.
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Affiliation(s)
- Michael Schonfeld
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Maura O’Neil
- Department of Pathology, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
- Liver Center, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Maria T Villar
- Department of Biochemistry, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Antonio Artigues
- Department of Biochemistry, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Janice Averilla
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Sumedha Gunewardena
- Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
| | - Steven A. Weinman
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
- Liver Center, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
- Kansas City VA Medical Center, Kansas City, MO, USA
| | - Irina Tikhanovich
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
- Liver Center, University of Kansas Medical Center, Kansas City, KS 66160, U.S.A
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Huck I, Morris EM, Thyfault J, Apte U. Hepatocyte-Specific Hepatocyte Nuclear Factor 4 Alpha (HNF4) Deletion Decreases Resting Energy Expenditure by Disrupting Lipid and Carbohydrate Homeostasis. Gene Expr 2021; 20:157-168. [PMID: 33691903 PMCID: PMC8201658 DOI: 10.3727/105221621x16153933463538] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023]
Abstract
Hepatocyte nuclear factor 4 alpha (HNF4) is required for hepatocyte differentiation and regulates expression of genes involved in lipid and carbohydrate metabolism including those that control VLDL secretion and gluconeogenesis. Whereas previous studies have focused on specific genes regulated by HNF4 in metabolism, its overall role in whole-body energy utilization has not been studied. In this study, we used indirect calorimetry to determine the effect of hepatocyte-specific HNF4 deletion (HNF4-KO) in mice on whole-body energy expenditure (EE) and substrate utilization in fed, fasted, and high-fat diet (HFD) conditions. HNF4-KO had reduced resting EE during fed conditions and higher rates of carbohydrate oxidation with fasting. HNF4-KO mice exhibited decreased body mass caused by fat mass depletion despite no change in energy intake and evidence of positive energy balance. HNF4-KO mice were able to upregulate lipid oxidation during HFD, suggesting that their metabolic flexibility was intact. However, only hepatocyte-specific HNF4-KO mice exhibited significant reduction in basal metabolic rate and spontaneous activity during HFD. Consistent with previous studies, hepatic gene expression in HNF4-KO supports decreased gluconeogenesis and decreased VLDL export and hepatic -oxidation in HNF4-KO livers across all feeding conditions. Together, our data suggest that deletion of hepatic HNF4 increases dependence on dietary carbohydrates and endogenous lipids for energy during fed and fasted conditions by inhibiting hepatic gluconeogenesis, hepatic lipid export, and intestinal lipid absorption resulting in decreased whole-body energy expenditure. These data clarify the role of hepatic HNF4 on systemic metabolism and energy homeostasis.
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Affiliation(s)
- Ian Huck
- *Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA
| | - E. Matthew Morris
- †Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, KS, USA
| | - John Thyfault
- †Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, KS, USA
- ‡Research Service, Kansas City VA Medical Center, Kansas City, KS, USA
| | - Udayan Apte
- *Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA
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Bhushan B, Molina L, Koral K, Stoops JW, Mars WM, Banerjee S, Orr A, Paranjpe S, Monga SP, Locker J, Michalopoulos GK. Yes-Associated Protein Is Crucial for Constitutive Androstane Receptor-Driven Hepatocyte Proliferation But Not for Induction of Drug Metabolism Genes in Mice. Hepatology 2021; 73:2005-2022. [PMID: 32794202 PMCID: PMC7885729 DOI: 10.1002/hep.31521] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/16/2020] [Revised: 07/14/2020] [Accepted: 07/16/2020] [Indexed: 12/11/2022]
Abstract
BACKGROUND AND AIMS Constitutive androstane receptor (CAR) agonists, such as 1,4-bis [2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP), are known to cause robust hepatocyte proliferation and hepatomegaly in mice along with induction of drug metabolism genes without any associated liver injury. Yes-associated protein (Yap) is a key transcription regulator that tightly controls organ size, including that of liver. Our and other previous studies suggested increased nuclear localization and activation of Yap after TCPOBOP treatment in mice and the potential role of Yap in CAR-driven proliferative response. Here, we investigated a direct role of Yap in CAR-driven hepatomegaly and hepatocyte proliferation using hepatocyte-specific Yap-knockout (KO) mice. APPROACH AND RESULTS Adeno-associated virus 8-thyroxine binding globulin promoter-Cre recombinase vector was injected to Yap-floxed mice for achieving hepatocyte-specific Yap deletion followed by TCPOBOP treatment. Yap deletion did not decrease protein expression of CAR or CAR-driven induction of drug metabolism genes (including cytochrome P450 [Cyp] 2b10, Cyp2c55, and UDP-glucuronosyltransferase 1a1 [Ugt1a1]). However, Yap deletion substantially reduced TCPOBOP-induced hepatocyte proliferation. TCPOBOP-driven cell cycle activation was disrupted in Yap-KO mice because of delayed (and decreased) induction of cyclin D1 and higher expression of p21, resulting in decreased phosphorylation of retinoblastoma protein. Furthermore, the induction of other cyclins, which are sequentially involved in progression through cell cycle (including cyclin E1, A2, and B1), and important mitotic regulators (such as Aurora B kinase and polo-like kinase 1) was remarkably reduced in Yap-KO mice. Microarray analysis revealed that 26% of TCPOBOP-responsive genes that were mainly related to proliferation, but not to drug metabolism, were altered by Yap deletion. Yap regulated these proliferation genes through alerting expression of Myc and forkhead box protein M1, two critical transcriptional regulators of CAR-mediated hepatocyte proliferation. CONCLUSIONS Our study revealed an important role of Yap signaling in CAR-driven hepatocyte proliferation; however, CAR-driven induction of drug metabolism genes was independent of Yap.
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Affiliation(s)
- Bharat Bhushan
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
| | - Laura Molina
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
| | - Kelly Koral
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
| | - John W. Stoops
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
| | - Wendy M. Mars
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
| | - Swati Banerjee
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
| | - Anne Orr
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
| | - Shirish Paranjpe
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
| | - Satdarshan P. Monga
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
| | - Joseph Locker
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
| | - George K. Michalopoulos
- Department of Pathology and Pittsburgh Liver Research CenterSchool of MedicineUniversity of PittsburghPittsburghPAUSA
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Pradhan-Sundd T, Liu S, Singh S, Poddar M, Ko S, Bell A, Franks J, Huck I, Stolz D, Apte U, Ranganathan S, Nejak-Bowen K, Monga SP. Dual β-Catenin and γ-Catenin Loss in Hepatocytes Impacts Their Polarity through Altered Transforming Growth Factor-β and Hepatocyte Nuclear Factor 4α Signaling. THE AMERICAN JOURNAL OF PATHOLOGY 2021; 191:885-901. [PMID: 33662348 DOI: 10.1016/j.ajpath.2021.02.008] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/19/2020] [Revised: 02/03/2021] [Accepted: 02/12/2021] [Indexed: 12/24/2022]
Abstract
Hepatocytes are highly polarized epithelia. Loss of hepatocyte polarity is associated with various liver diseases, including cholestasis. However, the molecular underpinnings of hepatocyte polarization remain poorly understood. Loss of β-catenin at adherens junctions is compensated by γ-catenin and dual loss of both catenins in double knockouts (DKOs) in mice liver leads to progressive intrahepatic cholestasis. However, the clinical relevance of this observation, and further phenotypic characterization of the phenotype, is important. Herein, simultaneous loss of β-catenin and γ-catenin was identified in a subset of liver samples from patients of progressive familial intrahepatic cholestasis and primary sclerosing cholangitis. Hepatocytes in DKO mice exhibited defects in apical-basolateral localization of polarity proteins, impaired bile canaliculi formation, and loss of microvilli. Loss of polarity in DKO livers manifested as epithelial-mesenchymal transition, increased hepatocyte proliferation, and suppression of hepatocyte differentiation, which was associated with up-regulation of transforming growth factor-β signaling and repression of hepatocyte nuclear factor 4α expression and activity. In conclusion, concomitant loss of the two catenins in the liver may play a pathogenic role in subsets of cholangiopathies. The findings also support a previously unknown role of β-catenin and γ-catenin in the maintenance of hepatocyte polarity. Improved understanding of the regulation of hepatocyte polarization processes by β-catenin and γ-catenin may potentially benefit development of new therapies for cholestasis.
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Affiliation(s)
- Tirthadipa Pradhan-Sundd
- Pittsburgh Heart, Lung and Blood Vascular Medicine Institute, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania; Department of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania; Pittsburgh Liver Research Center, University of Pittsburgh School of Medicine and University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania.
| | - Silvia Liu
- Pittsburgh Liver Research Center, University of Pittsburgh School of Medicine and University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania; Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania
| | - Sucha Singh
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania
| | - Minakshi Poddar
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania
| | - Sungjin Ko
- Pittsburgh Liver Research Center, University of Pittsburgh School of Medicine and University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania; Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania
| | - Aaron Bell
- Pittsburgh Liver Research Center, University of Pittsburgh School of Medicine and University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania; Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania
| | - Jonathan Franks
- Department of Cell Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania
| | - Ian Huck
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas
| | - Donna Stolz
- Department of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania; Pittsburgh Liver Research Center, University of Pittsburgh School of Medicine and University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania; Department of Cell Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania
| | - Udayan Apte
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas
| | - Sarangarajan Ranganathan
- Pittsburgh Liver Research Center, University of Pittsburgh School of Medicine and University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania; Department of Pediatrics, University of Pittsburgh Medical Center Children's Hospital of Pittsburgh, Pittsburgh, Pennsylvania
| | - Kari Nejak-Bowen
- Pittsburgh Liver Research Center, University of Pittsburgh School of Medicine and University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania; Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania
| | - Satdarshan P Monga
- Department of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania; Pittsburgh Liver Research Center, University of Pittsburgh School of Medicine and University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania; Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania.
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Control of Cell Identity by the Nuclear Receptor HNF4 in Organ Pathophysiology. Cells 2020; 9:cells9102185. [PMID: 32998360 PMCID: PMC7600215 DOI: 10.3390/cells9102185] [Citation(s) in RCA: 42] [Impact Index Per Article: 8.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2020] [Revised: 09/25/2020] [Accepted: 09/26/2020] [Indexed: 12/14/2022] Open
Abstract
Hepatocyte Nuclear Factor 4 (HNF4) is a transcription factor (TF) belonging to the nuclear receptor family whose expression and activities are restricted to a limited number of organs including the liver and gastrointestinal tract. In this review, we present robust evidence pointing to HNF4 as a master regulator of cellular differentiation during development and a safekeeper of acquired cell identity in adult organs. Importantly, we discuss that transient loss of HNF4 may represent a protective mechanism upon acute organ injury, while prolonged impairment of HNF4 activities could contribute to organ dysfunction. In this context, we describe in detail mechanisms involved in the pathophysiological control of cell identity by HNF4, including how HNF4 works as part of cell-specific TF networks and how its expression/activities are disrupted in injured organs.
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A negative reciprocal regulatory axis between cyclin D1 and HNF4α modulates cell cycle progression and metabolism in the liver. Proc Natl Acad Sci U S A 2020; 117:17177-17186. [PMID: 32631996 DOI: 10.1073/pnas.2002898117] [Citation(s) in RCA: 43] [Impact Index Per Article: 8.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/24/2022] Open
Abstract
Hepatocyte nuclear factor 4α (HNF4α) is a master regulator of liver function and a tumor suppressor in hepatocellular carcinoma (HCC). In this study, we explore the reciprocal negative regulation of HNF4α and cyclin D1, a key cell cycle protein in the liver. Transcriptomic analysis of cultured hepatocyte and HCC cells found that cyclin D1 knockdown induced the expression of a large network of HNF4α-regulated genes. Chromatin immunoprecipitation-sequencing (ChIP-seq) demonstrated that cyclin D1 inhibits the binding of HNF4α to thousands of targets in the liver, thereby diminishing the expression of associated genes that regulate diverse metabolic activities. Conversely, acute HNF4α deletion in the liver induces cyclin D1 and hepatocyte cell cycle progression; concurrent cyclin D1 ablation blocked this proliferation, suggesting that HNF4α maintains proliferative quiescence in the liver, at least, in part, via repression of cyclin D1. Acute cyclin D1 deletion in the regenerating liver markedly inhibited hepatocyte proliferation after partial hepatectomy, confirming its pivotal role in cell cycle progression in this in vivo model, and enhanced the expression of HNF4α target proteins. Hepatocyte cyclin D1 gene ablation caused markedly increased postprandial liver glycogen levels (in a HNF4α-dependent fashion), indicating that the cyclin D1-HNF4α axis regulates glucose metabolism in response to feeding. In AML12 hepatocytes, cyclin D1 depletion led to increased glucose uptake, which was negated if HNF4α was depleted simultaneously, and markedly elevated glycogen synthesis. To summarize, mutual repression by cyclin D1 and HNF4α coordinately controls the cell cycle machinery and metabolism in the liver.
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Lee S, Zhou P, Whyte S, Shin S. Adeno-Associated Virus Serotype 8-Mediated Genetic Labeling of Cholangiocytes in the Neonatal Murine Liver. Pharmaceutics 2020; 12:pharmaceutics12040351. [PMID: 32295003 PMCID: PMC7238059 DOI: 10.3390/pharmaceutics12040351] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/24/2020] [Revised: 04/08/2020] [Accepted: 04/10/2020] [Indexed: 12/13/2022] Open
Abstract
Determination of the cellular tropism of viral vectors is imperative for designing precise gene therapy. It has been widely accepted that transduction of hepatocytes using adeno-associated virus serotype 8 (AAV8) is a promising approach to correct inborn errors in neonates, but the type of neonatal hepatic cells transduced by AAV8 has not been thoroughly investigated. To address this question, we used a reporter mouse that carries Cre recombinase (Cre)-inducible yellow fluorescent protein (YFP). Our analysis primarily focused on cholangiocytes, given their pivotal roles in normal liver function and disease. We treated RosaYFP/+ mice at postnatal day 2 (P2) with AAV8-cytomegalovirus (CMV) promoter-Cre and analyzed livers at P10 and P56. The vast majority of HNF4α+ hepatocytes were labeled with YFP at both time points, and 11.6% and 24.4% of CK19+ cholangiocytes were marked at P10 and P56, respectively. We also detected YFP+ cells devoid of hepatocyte and cholangiocyte markers, and a subset of these cells expressed the endothelial and fibroblast marker CD34. Next, we used the hepatocyte-specific thyroxine-binding globulin (TBG) promoter. Surprisingly, AAV8-TBG-Cre marked 6.8% and 30.9% of cholangiocytes at P10 and P56, respectively. These results suggest that AAV8 can be a useful tool for targeting cholangiocytes in neonatal livers.
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Affiliation(s)
- Sanghoon Lee
- Division of Pediatric General and Thoracic Surgery, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45229, USA
- Department of Surgery, University of Cincinnati College of Medicine, Cincinnati, OH 45229, USA
| | - Ping Zhou
- Division of Pediatric General and Thoracic Surgery, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45229, USA
- Department of Surgery, University of Cincinnati College of Medicine, Cincinnati, OH 45229, USA
| | - Senyo Whyte
- Division of Pediatric General and Thoracic Surgery, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45229, USA
- Department of Surgery, University of Cincinnati College of Medicine, Cincinnati, OH 45229, USA
- Medical Scientist Training Program, University of Cincinnati College of Medicine, Cincinnati, OH 45229, USA
- Molecular and Developmental Biology Graduate Program, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Soona Shin
- Division of Pediatric General and Thoracic Surgery, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45229, USA
- Department of Surgery, University of Cincinnati College of Medicine, Cincinnati, OH 45229, USA
- Correspondence:
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Nuclear receptor HNF4α performs a tumor suppressor function in prostate cancer via its induction of p21-driven cellular senescence. Oncogene 2019; 39:1572-1589. [PMID: 31695151 PMCID: PMC7018660 DOI: 10.1038/s41388-019-1080-3] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2018] [Revised: 10/17/2019] [Accepted: 10/17/2019] [Indexed: 12/20/2022]
Abstract
Hepatocyte nuclear factor 4α (HNF4α, NR2A1) is a highly conserved member of the nuclear receptor superfamily. Recent advances reveal that it is a key transcriptional regulator of genes, broadly involved in xenobiotic and drug metabolism and also cancers of gastrointestinal tract. However, the exact functional roles of HNF4α in prostate cancer progression are still not fully understood. In this study, we determined the functional significance of HNF4α in prostate cancer. Our results showed that HNF4α exhibited a reduced expression pattern in clinical prostate cancer tissues, prostate cancer cell lines and xenograft model of castration-relapse prostate cancer. Stable HNF4α knockdown not only could promote cell proliferation and suppress doxorubicin (Dox)-induced cellular senescence in prostate cancer cells, but also confer resistance to paclitaxel treatment and enhance colony formation capacity and in vivo tumorigenicity of prostate cancer cells. On the contrary, ectopic overexpression of HNF4α could significantly inhibit the cell proliferation of prostate cancer cells, induce cell-cycle arrest at G2/M phase and trigger the cellular senescence in prostate cancer cells by activation of p21 signal pathway in a p53-independent manner via its direct transactivation of CDKN1A. Together, our results show that HNF4α performs a tumor suppressor function in prostate cancer via a mechanism of p21-driven cellular senescence.
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40
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Wang AW, Wang YJ, Zahm AM, Morgan AR, Wangensteen KJ, Kaestner KH. The Dynamic Chromatin Architecture of the Regenerating Liver. Cell Mol Gastroenterol Hepatol 2019; 9:121-143. [PMID: 31629814 PMCID: PMC6909351 DOI: 10.1016/j.jcmgh.2019.09.006] [Citation(s) in RCA: 41] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/21/2019] [Revised: 09/19/2019] [Accepted: 09/23/2019] [Indexed: 12/12/2022]
Abstract
BACKGROUND & AIMS The adult liver is the main detoxification organ and routinely is exposed to environmental insults but retains the ability to restore its mass and function upon tissue damage. However, extensive injury can lead to liver failure, and chronic injury causes fibrosis, cirrhosis, and hepatocellular carcinoma. Currently, the transcriptional regulation of organ repair in the adult liver is incompletely understood. METHODS We isolated nuclei from quiescent as well as repopulating hepatocytes in a mouse model of hereditary tyrosinemia, which recapitulates the injury and repopulation seen in toxic liver injury in human beings. We then performed the assay for transposase accessible chromatin with high-throughput sequencing specifically in repopulating hepatocytes to identify differentially accessible chromatin regions and nucleosome positioning. In addition, we used motif analysis to predict differential transcription factor occupancy and validated the in silico results with chromatin immunoprecipitation followed by sequencing for hepatocyte nuclear factor 4α (HNF4α) and CCCTC-binding factor (CTCF). RESULTS Chromatin accessibility in repopulating hepatocytes was increased in the regulatory regions of genes promoting proliferation and decreased in the regulatory regions of genes involved in metabolism. The epigenetic changes at promoters and liver enhancers correspond with the regulation of gene expression, with enhancers of many liver function genes showing a less accessible state during the regenerative process. Moreover, increased CTCF occupancy at promoters and decreased HNF4α binding at enhancers implicate these factors as key drivers of the transcriptomic changes in replicating hepatocytes that enable liver repopulation. CONCLUSIONS Our analysis of hepatocyte-specific epigenomic changes during liver repopulation identified CTCF and HNF4α as key regulators of hepatocyte proliferation and regulation of metabolic programs. Thus, liver repopulation in the setting of toxic injury makes use of both general transcription factors (CTCF) for promoter activation, and reduced binding by a hepatocyte-enriched factor (HNF4α) to temporarily limit enhancer activity. All sequencing data in this study were deposited to the Gene Expression Omnibus database and can be downloaded with accession number GSE109466.
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Affiliation(s)
- Amber W Wang
- Department of Genetics, University of Pennsylvania, Philadelphia, Pennsylvania
| | - Yue J Wang
- Department of Genetics, University of Pennsylvania, Philadelphia, Pennsylvania; Department of Biomedical Sciences, Florida State University College of Medicine, Tallahassee, Florida
| | - Adam M Zahm
- Department of Genetics, University of Pennsylvania, Philadelphia, Pennsylvania
| | - Ashleigh R Morgan
- Department of Genetics, University of Pennsylvania, Philadelphia, Pennsylvania
| | - Kirk J Wangensteen
- Department of Genetics, University of Pennsylvania, Philadelphia, Pennsylvania; Department of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania
| | - Klaus H Kaestner
- Department of Genetics, University of Pennsylvania, Philadelphia, Pennsylvania.
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41
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Thakur A, Wong JCH, Wang EY, Lotto J, Kim D, Cheng JC, Mingay M, Cullum R, Moudgil V, Ahmed N, Tsai SH, Wei W, Walsh CP, Stephan T, Bilenky M, Fuglerud BM, Karimi MM, Gonzalez FJ, Hirst M, Hoodless PA. Hepatocyte Nuclear Factor 4-Alpha Is Essential for the Active Epigenetic State at Enhancers in Mouse Liver. Hepatology 2019; 70:1360-1376. [PMID: 30933372 PMCID: PMC6773525 DOI: 10.1002/hep.30631] [Citation(s) in RCA: 49] [Impact Index Per Article: 8.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/29/2018] [Accepted: 03/27/2019] [Indexed: 12/11/2022]
Abstract
Cell-fate determination is influenced by interactions between master transcription factors (TFs) and cis-regulatory elements. Hepatocyte nuclear factor 4 alpha (HNF4A), a liver-enriched TF, acts as a master controller in specification of hepatic progenitor cells by regulating a network of TFs to control onset of hepatocyte cell fate. Using analysis of genome-wide histone modifications, DNA methylation, and hydroxymethylation in mouse hepatocytes, we show that HNF4A occupies active enhancers in hepatocytes and is essential for active histone and DNA signatures, especially acetylation of lysine 27 of histone 3 (H3K27ac) and 5-hydroxymethylcytosine (5hmC). In mice lacking HNF4A protein in hepatocytes, we observed a decrease in both H3K27ac and hydroxymethylation at regions bound by HNF4A. Mechanistically, HNF4A-associated hydroxymethylation (5hmC) requires its interaction with ten-eleven translocation methylcytosine dioxygenase 3 (TET3), a protein responsible for oxidation from 5mC to 5hmC. Furthermore, HNF4A regulates TET3 expression in liver by directly binding to an enhancer region. Conclusion: In conclusion, we identified that HNF4A is required for the active epigenetic state at enhancers that amplifies transcription of genes in hepatocytes.
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Affiliation(s)
- Avinash Thakur
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3,Department of Medical Genetics, University of British Columbia, Vancouver, Canada, V6T 1Z4
| | - Jasper C. H. Wong
- Department of Microbiology and Immunology, Michael Smith Laboratories Centre for High-Throughput Biology, University of British Columbia, Vancouver, British Columbia, Canada
| | - Evan Y. Wang
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3
| | - Jeremy Lotto
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3
| | - Donghwan Kim
- Center of Cancer Research, National Cancer Institute, Bethesda MD 2089
| | - Jung-Chien Cheng
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3
| | - Matthew Mingay
- Department of Microbiology and Immunology, Michael Smith Laboratories Centre for High-Throughput Biology, University of British Columbia, Vancouver, British Columbia, Canada
| | - Rebecca Cullum
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3
| | - Vaishali Moudgil
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3
| | - Nafeel Ahmed
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3
| | - Shu-Huei Tsai
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3
| | - Wei Wei
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3
| | - Colum P. Walsh
- Genomic Medicine Research Group, Centre for Molecular Biosciences, Biomedical Sciences Research Institute, Ulster University, Coleraine, BT52 1SA, UK
| | - Tabea Stephan
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3
| | - Misha Bilenky
- Genome Sciences Centre, BC Cancer, Vancouver, British Columbia, Canada
| | - Bettina M. Fuglerud
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3,Department of Biosciences, University of Oslo, Oslo, Norway, 0316
| | | | - Frank J. Gonzalez
- Center of Cancer Research, National Cancer Institute, Bethesda MD 2089
| | - Martin Hirst
- Department of Microbiology and Immunology, Michael Smith Laboratories Centre for High-Throughput Biology, University of British Columbia, Vancouver, British Columbia, Canada,Genome Sciences Centre, BC Cancer, Vancouver, British Columbia, Canada
| | - Pamela A. Hoodless
- Terry Fox Laboratory, BC Cancer, Vancouver, British Columbia, Canada, V5Z 1L3,Department of Medical Genetics, University of British Columbia, Vancouver, Canada, V6T 1Z4,School of Biomedical Engineering, University of British Columbia, Vancouver, Canada, V6T 1Z4
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42
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Fekry B, Ribas-Latre A, Baumgartner C, Mohamed AMT, Kolonin MG, Sladek FM, Younes M, Eckel-Mahan KL. HNF4α-Deficient Fatty Liver Provides a Permissive Environment for Sex-Independent Hepatocellular Carcinoma. Cancer Res 2019; 79:5860-5873. [PMID: 31575546 DOI: 10.1158/0008-5472.can-19-1277] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2019] [Revised: 08/02/2019] [Accepted: 09/23/2019] [Indexed: 02/07/2023]
Abstract
The incidence of hepatocellular carcinoma (HCC) is on the rise worldwide. Although the incidence of HCC in males is considerably higher than in females, the projected rates of HCC incidence are increasing for both sexes. A recently appreciated risk factor for HCC is the growing problem of nonalcoholic fatty liver disease, which is usually associated with obesity and the metabolic syndrome. In this study, we showed that under conditions of fatty liver, female mice were more likely to develop HCC than expected from previous models. Using an inducible knockout model of the tumor-suppressive isoform of hepatocyte nuclear factor 4 alpha ("P1-HNF4α") in the liver in combination with prolonged high fat (HF) diet, we found that HCC developed equally in male and female mice as early as 38 weeks of age. Similar sex-independent HCC occurred in the "STAM" model of mice, in which severe hyperglycemia and HF feeding results in rapid hepatic lipid deposition, fibrosis, and ultimately HCC. In both sexes, reduced P1-HNF4α activity, which also occurs under chronic HF diet feeding, increased hepatic lipid deposition and produced a greatly augmented circadian rhythm in IL6, a factor previously linked with higher HCC incidence in males. Loss of HNF4α combined with HF feeding induced epithelial-mesenchymal transition in an IL6-dependent manner. Collectively, these data provide a mechanism-based working hypothesis that could explain the rising incidence of aggressive HCC. SIGNIFICANCE: This study provides a mechanism for the growing incidence of hepatocellular carcinoma in both men and women, which is linked to nonalcoholic fatty liver disease.
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Affiliation(s)
- Baharan Fekry
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, Texas
| | - Aleix Ribas-Latre
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, Texas
| | - Corrine Baumgartner
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, Texas
| | - Alaa M T Mohamed
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, Texas
| | - Mikhail G Kolonin
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, Texas.,Department of Integrative Biology and Pharmacology, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, Texas
| | - Frances M Sladek
- Department of Molecular, Cell and Systems Biology, University of California Riverside, Riverside, California
| | - Mamoun Younes
- Department of Pathology and Laboratory Medicine, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, Texas
| | - Kristin L Eckel-Mahan
- Institute of Molecular Medicine, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, Texas. .,Department of Integrative Biology and Pharmacology, McGovern Medical School at the University of Texas Health Science Center (UT Health), Houston, Texas
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43
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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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Huck I, Gunewardena S, Espanol-Suner R, Willenbring H, Apte U. Hepatocyte Nuclear Factor 4 Alpha Activation Is Essential for Termination of Liver Regeneration in Mice. Hepatology 2019; 70:666-681. [PMID: 30520062 PMCID: PMC6551324 DOI: 10.1002/hep.30405] [Citation(s) in RCA: 72] [Impact Index Per Article: 12.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/03/2018] [Accepted: 11/23/2018] [Indexed: 12/14/2022]
Abstract
Hepatocyte nuclear factor 4 alpha (HNF4α) is critical for hepatic differentiation. Recent studies have highlighted its role in inhibition of hepatocyte proliferation and tumor suppression. However, the role of HNF4α in liver regeneration (LR) is not known. We hypothesized that hepatocytes modulate HNF4α activity when navigating between differentiated and proliferative states during LR. Western blotting analysis revealed a rapid decline in nuclear and cytoplasmic HNF4α protein levels, accompanied with decreased target gene expression, within 1 hour after two-thirds partial hepatectomy (post-PH) in C57BL/6J mice. HNF4α protein expression did not recover to pre-PH levels until day 3. Hepatocyte-specific deletion of HNF4α (HNF4α-KO [knockout]) in mice resulted in 100% mortality post-PH, despite increased proliferative marker expression throughout regeneration. Sustained loss of HNF4α target gene expression throughout regeneration indicated that HNF4α-KO mice were unable to compensate for loss of HNF4α transcriptional activity. Deletion of HNF4α resulted in sustained proliferation accompanied by c-Myc and cyclin D1 overexpression and a complete deficiency of hepatocyte function after PH. Interestingly, overexpression of degradation-resistant HNF4α in hepatocytes delayed, but did not prevent, initiation of regeneration after PH. Finally, adeno-associated virus serotype 8 (AAV8)-mediated reexpression of HNF4α in hepatocytes of HNF4α-KO mice post-PH restored HNF4α protein levels, induced target gene expression, and improved survival of HNF4α-KO mice post-PH. Conclusion: In conclusion, these data indicate that HNF4α reexpression following initial decrease is critical for hepatocytes to exit from cell cycle and resume function during the termination phase of LR. These results indicate the role of HNF4α in LR and have implications for therapy of liver failure.
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Affiliation(s)
- Ian Huck
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS
| | - Sumedha Gunewardena
- Department of Biostatistics University of Kansas Medical Center, Kansas City, KS
| | | | - Holger Willenbring
- Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research,Liver Center, Division of Transplantation, University of California San Francisco, San Francisco, CA,Department of Surgery, Division of Transplantation, University of California San Francisco, San Francisco, CA
| | - Udayan Apte
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS
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45
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Sartor C, Bachelot L, Godard C, Lager F, Renault G, Gonzalez FJ, Perret C, Gougelet A, Colnot S. The concomitant loss of APC and HNF4α in adult hepatocytes does not contribute to hepatocarcinogenesis driven by β-catenin activation. Liver Int 2019; 39:727-739. [PMID: 30721564 PMCID: PMC7387933 DOI: 10.1111/liv.14068] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/05/2018] [Revised: 01/29/2019] [Accepted: 01/30/2019] [Indexed: 02/13/2023]
Abstract
BACKGROUND & AIMS Loss of hepatocyte nuclear factor-4α (HNF4α), a critical factor driving liver development and differentiation, is frequently associated with hepatocellular carcinoma (HCC). Our recent data revealed that HNF4α level was decreased in mouse and human HCCs with activated β-catenin signalling. In addition, increasing HNF4α level by miR-34a inhibition slowed tumour progression of β-catenin-activated HCC in mice. METHODS We generated a Hnf4aflox/flox/ Apcflox/flox /TTR-CreERT2 (Hnf4a/Apc∆Hep ) mouse line and evaluated the impact of Hnf4a disruption on HCC development and liver homoeostasis. RESULTS There was no significant impact of Hnf4a disruption on tumour onset and progression in Apc∆Hep model. However, we observed an unexpected phenotype in 28% of Hnf4a∆Hep mice maintained in a conventional animal facility, which presented disorganized portal triads, characterized by stenosis of the portal vein and increased number and size of hepatic arteries and bile ducts. These abnormal portal structures resemble the human idiopathic non-cirrhotic portal hypertension syndrome. We correlated the presence of portal remodelling with a higher expression of protein and mRNA levels of TGFβ and BMP7, a key regulator of the TGFβ-dependent endothelial-to-mesenchymal transition. CONCLUSION These data demonstrate that HNF4α does not play a major role during β-catenin-driven HCC, thus revealing that the tumour suppressor role of HNF4α is far more complex and dependent probably on its temporal expression and tumour context. However, HNF4α loss in adult hepatocytes could induce abnormal portal structures resembling the human idiopathic non-cirrhotic portal hypertension syndrome, which may result from endothelial- and epithelial-to-mesenchymal transitions.
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Affiliation(s)
- Chiara Sartor
- Inserm, U1016, Institut Cochin, Paris, France
- CNRS, UMR8104, Paris, France
- Université Paris Descartes, Sorbonne Paris Cité, Paris, France
- Equipe labellisée LNCC, Paris, France
| | - Laura Bachelot
- Inserm, U1016, Institut Cochin, Paris, France
- CNRS, UMR8104, Paris, France
- Université Paris Descartes, Sorbonne Paris Cité, Paris, France
- Equipe labellisée LNCC, Paris, France
| | - Cécile Godard
- Inserm, U1016, Institut Cochin, Paris, France
- CNRS, UMR8104, Paris, France
- Université Paris Descartes, Sorbonne Paris Cité, Paris, France
- Equipe labellisée LNCC, Paris, France
| | - Franck Lager
- Inserm, U1016, Institut Cochin, Paris, France
- CNRS, UMR8104, Paris, France
- Université Paris Descartes, Sorbonne Paris Cité, Paris, France
- Plateforme Imageries du Vivant – PIV, Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Gilles Renault
- Inserm, U1016, Institut Cochin, Paris, France
- CNRS, UMR8104, Paris, France
- Université Paris Descartes, Sorbonne Paris Cité, Paris, France
- Plateforme Imageries du Vivant – PIV, Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Frank J. Gonzalez
- Laboratory of Metabolism, National Cancer Institute, Bethesda, Maryland
| | - Christine Perret
- Inserm, U1016, Institut Cochin, Paris, France
- CNRS, UMR8104, Paris, France
- Université Paris Descartes, Sorbonne Paris Cité, Paris, France
- Equipe labellisée LNCC, Paris, France
| | - Angélique Gougelet
- Inserm, U1016, Institut Cochin, Paris, France
- CNRS, UMR8104, Paris, France
- Université Paris Descartes, Sorbonne Paris Cité, Paris, France
- Equipe labellisée LNCC, Paris, France
| | - Sabine Colnot
- Inserm, U1016, Institut Cochin, Paris, France
- CNRS, UMR8104, Paris, France
- Université Paris Descartes, Sorbonne Paris Cité, Paris, France
- Equipe labellisée LNCC, Paris, France
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Bhushan B, Stoops JW, Mars WM, Orr A, Bowen WC, Paranjpe S, Michalopoulos GK. TCPOBOP-Induced Hepatomegaly and Hepatocyte Proliferation are Attenuated by Combined Disruption of MET and EGFR Signaling. Hepatology 2019; 69:1702-1718. [PMID: 29888801 PMCID: PMC6289897 DOI: 10.1002/hep.30109] [Citation(s) in RCA: 36] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/21/2018] [Accepted: 08/10/2018] [Indexed: 12/13/2022]
Abstract
TCPOBOP (1,4-Bis [2-(3,5-Dichloropyridyloxy)] benzene) is a constitutive androstane receptor (CAR) agonist that induces robust hepatocyte proliferation and hepatomegaly without any liver injury or tissue loss. TCPOBOP-induced direct hyperplasia has been considered to be CAR-dependent with no evidence of involvement of cytokines or growth factor signaling. Receptor tyrosine kinases (RTKs), MET and epidermal growth factor receptor (EGFR), are known to play a critical role in liver regeneration after partial hepatectomy, but their role in TCPOBOP-induced direct hyperplasia, not yet explored, is investigated in the current study. Disruption of the RTK-mediated signaling was achieved using MET knockout (KO) mice along with Canertinib treatment for EGFR inhibition. Combined elimination of MET and EGFR signaling [MET KO + EGFR inhibitor (EGFRi)], but not individual disruption, dramatically reduced TCPOBOP-induced hepatomegaly and hepatocyte proliferation. TCPOBOP-driven CAR activation was not altered in [MET KO + EGFRi] mice, as measured by nuclear CAR translocation and analysis of typical CAR target genes. However, TCPOBOP-induced cell cycle activation was impaired in [MET KO + EGFRi] mice due to defective induction of cyclins, which regulate cell cycle initiation and progression. TCPOBOP-driven induction of FOXM1, a key transcriptional regulator of cell cycle progression during TCPOBOP-mediated hepatocyte proliferation, was greatly attenuated in [MET KO + EGFRi] mice. Interestingly, TCPOBOP treatment caused transient decline in hepatocyte nuclear factor 4 alpha expression concomitant to proliferative response; this was not seen in [MET KO + EGFRi] mice. Transcriptomic profiling revealed the vast majority (~40%) of TCPOBOP-dependent genes primarily related to proliferative response, but not to drug metabolism, were differentially expressed in [MET KO + EGFRi] mice. Conclusion: Taken together, combined disruption of EGFR and MET signaling lead to dramatic impairment of TCPOBOP-induced proliferative response without altering CAR activation.
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Affiliation(s)
- Bharat Bhushan
- Department of Pathology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania
| | - John W Stoops
- Department of Pathology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania
| | - Wendy M Mars
- Department of Pathology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania
| | - Anne Orr
- Department of Pathology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania
| | - William C Bowen
- Department of Pathology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania
| | - Shirish Paranjpe
- Department of Pathology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania
| | - George K Michalopoulos
- Department of Pathology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania
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47
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Shi Y, Zhou D, Wang B, Zhou D, Shi B. Roles and mechanisms of action of HNF‑4α in the hepatic differentiation of WB‑F344 cells. Int J Mol Med 2019; 43:1021-1032. [PMID: 30535491 DOI: 10.3892/ijmm.2018.4010] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2018] [Accepted: 11/26/2018] [Indexed: 11/06/2022] Open
Abstract
Hepatocyte nuclear factor 4 α (HNF‑4α) is a nuclear receptor and mediates hepatic genes. WB‑F344 liver epithelial cells can differentiate into hepatocytes. The present study aimed to examine the roles and mechanisms of action of HNF‑4α on the hepatic differentiation of WB‑F344 cells. WB‑F344 cells were divided into a normal cell group (WB‑F344), empty vector group (PLKO), and gene silencing group (PLKO‑SH). The expression levels of HNF‑4α were measured using reverse transcription‑quantitative polymerase chain reaction analysis. Proliferation of the cells was determined using a Cell Counting kit‑8 assay. Based on western blot analysis, the protein levels of α‑fetoprotein (AFP), albumin (ALB) and cytokeratin 19 (CK19) were determined. The positive cell rates of the three groups were assessed using periodic acid‑Schiff (PAS) staining. Following construction of an RNA‑sequencing library, differentially expressed genes (DEGs) between the HNF‑4α‑silenced and normal samples were screened using the limma package and enrichment analysis was conducted using the DAVID tool. Protein‑protein interaction (PPI) and microRNA‑targeted regulatory networks were constructed in Cytoscape software. The PLKO‑SH group exhibited a lower mRNA level of HNF‑4α, higher protein level of AFP, lower protein levels of ALB and CK19, increased cell proliferation, and a lower PAS‑positive cell rate. The HNF‑4α‑silenced and normal samples differed in 499 DEGs. In the PPI network, matrix metallopeptidase 9 (MMP9), early growth response 1 (EGR1), SMAD family member 2 (SMAD2), and RAS‑related C3 botulinum substrate 2 (RAC2) were key nodes. HNF‑4α may promote the differentiation of WB‑F344 cells into hepatocytes by targeting MMP9, EGR1, SMAD2 and RAC2.
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Affiliation(s)
- Yumeng Shi
- Cheeloo College of Medicine, Shandong University, Jinan, Shandong 250012, P.R. China
| | - Dehua Zhou
- Department of General Surgery, Tongji Hospital, Tongji University Medical School, Shanghai 200065, P.R. China, P.R. China
| | - Bingyi Wang
- Department of General Surgery, Tongji Hospital, Tongji University Medical School, Shanghai 200065, P.R. China, P.R. China
| | - Deren Zhou
- Department of General Surgery, Tongji Hospital, Tongji University Medical School, Shanghai 200065, P.R. China, P.R. China
| | - Baomin Shi
- Department of General Surgery, Tongji Hospital, Tongji University Medical School, Shanghai 200065, P.R. China, P.R. China
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Huck I, Beggs K, Apte U. Paradoxical Protective Effect of Perfluorooctanesulfonic Acid Against High-Fat Diet-Induced Hepatic Steatosis in Mice. Int J Toxicol 2018; 37:383-392. [PMID: 30134762 PMCID: PMC6150807 DOI: 10.1177/1091581818790934] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/28/2022]
Abstract
Perfluorooctanesulfonic acid (PFOS) is a persistent organic pollutant with worldwide bioaccumulation due to a very long half-life. Perfluorooctanesulfonic acid exposure results in significant hepatic effects including steatosis, proliferation, hepatomegaly, and in rodents, carcinogenesis. The objective of this study was to determine whether PFOS exposure exacerbates nonalcoholic fatty liver disease and nonalcoholic steatohepatitis pathogenesis. Eight-week-old male C57BL/6 J mice (n = 5 per group) were fed ad libitum normal chow diet (ND) alone, 60% high-fat diet (HFD) alone, ND + PFOS, and HFD + PFOS (0.0001% w/w (1 mg/kg) of PFOS) for 6 weeks. Both HFD alone and the ND + PFOS treatment induced significant adiposity and hepatomegaly, but the HFD + PFOS treatment showed a marked protection. Oil Red O staining and quantitative analysis of hepatic lipid content revealed increased hepatic steatosis in ND + PFOS and in HFD alone fed mice, which was prevented in HFD + PFOS treatment. Further studies revealed that ND + PFOS treatment significantly affected expression of lipid trafficking genes to favor steatosis, but these changes were absent in HFD + PFOS group. Specifically, expression of CD36, the major lipid importer in the cells, and peroxisome proliferator-activated receptor gamma (PPARγ), its major regulator, were induced in HFD + no treatment (NT) and ND + PFOS-fed mice but remained unchanged in HFD + PFOS mice. In conclusion, these data indicate that coadministration of PFOS with HFD mitigates steatosis and hepatomegaly induced by HFD and that by PFOS fed in ND diet via regulation of cellular lipid import machinery. These findings suggest dietary lipid content be considered when performing risk management of PFOS in humans and the elucidation of PFOS-induced hepatotoxicity.
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Affiliation(s)
- Ian Huck
- 1 Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA
| | - Kevin Beggs
- 1 Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA
| | - Udayan Apte
- 1 Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA
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Jiang Q, Sun Y, Guo Z, Chen R, Ma S, Fu M, Zhu H, Ning Q, Lei P, Shen G. IL-23 enhances the malignant properties of hepatoma cells by attenuation of HNF4α. Oncotarget 2018; 9:28309-28321. [PMID: 29983862 PMCID: PMC6033364 DOI: 10.18632/oncotarget.24875] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2017] [Accepted: 03/06/2018] [Indexed: 02/07/2023] Open
Abstract
Chronic infection with hepatitis B virus (HBV) is one of the major risk factors for hepatocellular carcinoma. HBV infection can induce the expression of IL-23. However, the effects of IL-23 on carcinogenesis are rare and contradictory. To investigate the potential role of IL-23 on malignant properties of hepatoma cells, in the present study, first, we confirmed that HBV drove infected hepatoma cells to produce more IL-23. And then we found that at low concentration, human recombinant IL-23 (hrIL-23) enhanced malignant properties of hepatoma cells through increasing the proportion of stem/progenitor cells, promoting proliferation and colony formation, reducing apoptosis and inducing motility and invasivity of them. Hepatocyte nuclear factor 4 alpha (HNF4α), which is essential for liver development and hepatocyte function, was found to be downregulated in HBV integrated or transiently transfected hepatoma cells. Its expression was also decreased in cells treated by hrIL-23 or by HepG2.215 culture supernatant and this decrease could be abolished by supplementation of anti-IL-23p19 antibody. Hence, it is speculated that HBV related IL-23 can enhance malignant properties of hepatoma cells through attenuation of HNF4α. The findings identified a potential target of interventional strategies for treating hepatitis B patients through manipulation of the IL-23.
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Affiliation(s)
- Qing Jiang
- Department of Immunology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.,Department of Allergy, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Yuanli Sun
- Department of Immunology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Zilong Guo
- Department of Immunology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Ru Chen
- Department of Immunology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Simin Ma
- Department of Immunology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Mingpeng Fu
- Department of Immunology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Huifen Zhu
- Department of Immunology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Qin Ning
- Department of Infectious Disease, Institute of Infectious Disease, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Ping Lei
- Department of Immunology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Guanxin Shen
- Department of Immunology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
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Borude P, Bhushan B, Apte U. DNA Damage Response Regulates Initiation of Liver Regeneration Following Acetaminophen Overdose. Gene Expr 2018; 18:115-123. [PMID: 29540258 PMCID: PMC5954624 DOI: 10.3727/105221618x15205260749346] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
Acetaminophen (APAP) overdose is the leading cause of acute liver failure (ALF) with limited treatment options. It is known that liver regeneration following APAP-induced ALF is a deciding factor in the final outcome. Previous studies from our laboratory using an incremental dose model involving a regenerating (300 mg/kg, APAP300) and a nonregenerating (600 mg/kg, APAP600) dose of APAP in mice have revealed several proregenerative pathways that regulate regeneration after APAP overdose. Here we report that DNA damage and repair mechanisms regulate initiation of liver regeneration following APAP overdose. Mice treated with nonregenerating APAP600 dose showed prolonged expression of pH2AX, a marker of the DNA double-strand break (DSB), compared with APAP300. In regenerating APAP300 dose-treated mice, H2AX was rapidly dephosphorylated at Tyr142, indicating timely DNA repair. Expression of several DNA repair proteins was substantially lower with APAP600. Poly(ADP) ribose polymerase (PARP) activation, involved in DNA repair, was significantly higher in the APAP300 group compared to the APAP600 group. Activation of p53, the major cell cycle checkpoint protein, was significantly higher with APAP600 as demonstrated by substantially higher expression of its target genes. Taken together, these data show that massive DNA DSB occurs in high-dose APAP toxicity, and lack of prompt DSB repair after APAP overdose leads to prolonged growth arrest and proliferative senescence, resulting in inhibited liver regeneration.
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Affiliation(s)
- Prachi Borude
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA
| | - Bharat Bhushan
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA
| | - Udayan Apte
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA
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