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Chi KY, Kim G, Son JS, Han J, Kim JH. Recent Advances in Three-Dimensional In Vitro Models for Studies of Liver Fibrosis. Tissue Eng Regen Med 2025:10.1007/s13770-025-00719-8. [PMID: 40358834 DOI: 10.1007/s13770-025-00719-8] [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: 02/02/2025] [Revised: 02/02/2025] [Accepted: 03/11/2025] [Indexed: 05/15/2025] Open
Abstract
BACKGROUND Liver fibrosis is a reversible but complex pathological condition associated with chronic liver diseases, affecting over 1.5 billion people worldwide. It is characterized by excessive extracellular matrix deposition resulting from sustained liver injury, often advancing to cirrhosis and cancer. As its progression involves various cell types and pathogenic factors, understanding the intricate mechanisms is essential for the development of effective therapies. In this context, extensive efforts have been made to establish three-dimensional (3D) in vitro platforms that mimic the progression of liver fibrosis. METHODS This review outlines the pathophysiology of liver fibrosis and highlights recent advancements in 3D in vitro liver models, including spheroids, organoids, assembloids, bioprinted constructs, and microfluidic systems. It further assesses their biological relevance, with particular focus on their capacity to reproduce fibrosis-related characteristics. RESULTS 3D in vitro liver models offer significant advantages over conventional two-dimensional cultures. Although each model exhibits unique strengths, they collectively recapitulate key fibrotic features, such as extracellular matrix remodeling, hepatic stellate cell activation, and collagen deposition, in a physiologically relevant 3D setting. In particular, multilineage liver organoids and assembloids integrate architectural complexity with scalability, enabling deeper mechanistic insights and supporting therapeutic evaluation with improved translational relevance. CONCLUSION 3D in vitro liver models represent a promising strategy to bridge the gap between in vitro studies and in vivo realities by faithfully replicating liver-specific architecture and microenvironments. With enhanced reproducibility through standardized protocols, these models hold great potential for advancing drug discovery and facilitating the development of personalized therapies for liver fibrosis.
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Affiliation(s)
- Kyun Yoo Chi
- Laboratory of Stem Cells and Tissue Regeneration, Department of Biotechnology, College of Life Sciences and Biotechnology, Korea University, Seoul, 02841, South Korea
| | - Gyeongmin Kim
- Laboratory of Stem Cells and Tissue Regeneration, Department of Biotechnology, College of Life Sciences and Biotechnology, Korea University, Seoul, 02841, South Korea
| | - Jeong Sang Son
- User Convenience Technology R&D Department, Korea Institute of Industrial Technology (KITECH), Ansan, 15588, South Korea
| | - Jiyou Han
- Department of Biomedical and Chemical Sciences, Hyupsung University, Hwasung-Si, 18330, South Korea
| | - Jong-Hoon Kim
- Laboratory of Stem Cells and Tissue Regeneration, Department of Biotechnology, College of Life Sciences and Biotechnology, Korea University, Seoul, 02841, South Korea.
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Xiang X, Shao Y, Xiang L, Jiao Q, Zhang W, Qin Y, Chen Y. Suppression of Liver Fibrogenesis with Photothermal Sorafenib Nanovesicles via Selectively Inhibiting Glycolysis and Amplification of Active HSCs. Mol Pharm 2025; 22:1939-1957. [PMID: 40053386 DOI: 10.1021/acs.molpharmaceut.4c01135] [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] [Indexed: 04/08/2025]
Abstract
As the major driving factor of hepatic fibrosis, the activated hepatic stellate cells (aHSCs) rely on active glycolysis to support their aberrant proliferation and secretion of the extracellular matrix. Sorafenib (Sor) can combat liver fibrosis by suppressing HIF-1α and glycolysis, but its poor solubility, rapid metabolism, and low bioavailability restrict such a clinical application. Here, Sor was loaded onto polydopamine nanoparticles and then encapsulated by a retinoid-decorated red blood cell membrane, yielding HSC-targeted Sor nanovesicles (PDA/Sor@RMV-VA) with a high Sor-loading capacity and photothermally controlled drug release for antifibrotic treatment. These Sor RMVs not only exhibited a good particle size, dispersity and biocompatibility, prolonged circulation time, enhanced aHSC targetability, and hepatic accumulation both in vitro and in vivo, but also displayed a mild photothermal activity proper for promoting sorafenib release and accumulation in CCl4-induced fibrotic mouse livers without incurring phototoxicity. Compared with nontargeting Sor formulations, PDA/Sor@RMV-VA more effectively downregulated HIF-1α and glycolytic enzyme in both cultured aHSCs and fibrotic mice and reversed myofibroblast phenotype and amplification of aHSCs and thus more significantly improved liver damage, inflammation, and fibrosis, all of which could be even further advanced with NIR irradiation. These results fully demonstrate the antifibrotic power and therapeutic potential of PDA/Sor@RMV-VA as an antifibrotic nanomedicine, which would support a new clinical treatment for hepatic fibrosis.
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Affiliation(s)
- Xianjing Xiang
- School of Pharmaceutical Sciences, University of South China, Hengyang 410001, China
| | - Yaru Shao
- School of Pharmaceutical Sciences, University of South China, Hengyang 410001, China
| | - Li Xiang
- School of Pharmaceutical Sciences, University of South China, Hengyang 410001, China
- Hengyang Medical School, University of South China, Hengyang, Hunan 410001, China
| | - Qiangqiang Jiao
- School of Pharmaceutical Sciences, University of South China, Hengyang 410001, China
| | - Wenhui Zhang
- School of Pharmaceutical Sciences, University of South China, Hengyang 410001, China
| | - Yuting Qin
- School of Pharmaceutical Sciences, University of South China, Hengyang 410001, China
| | - Yuping Chen
- School of Pharmaceutical Sciences, University of South China, Hengyang 410001, China
- Hengyang Medical School, University of South China, Hengyang, Hunan 410001, China
- MOE Key Laboratory of Rare Pediatric Diseases, Hengyang Medical School, University of South China, Hengyang, Hunan 410001, China
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Wu YT, Li QZ, Wu YQ, Mu M, Wu H, Tian HY, Zhao XK. Nintedanib attenuates NLRP3 inflammasome-driven liver fibrosis by targeting Src signaling. Int Immunopharmacol 2024; 143:113630. [PMID: 39549551 DOI: 10.1016/j.intimp.2024.113630] [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: 07/19/2024] [Revised: 10/16/2024] [Accepted: 11/11/2024] [Indexed: 11/18/2024]
Abstract
Liver injury induces an inflammatory response that activates hepatic stellate cells, which is the initial factor of liver fibrosis. Nintedanib, a multi-targeted tyrosine kinase inhibitor targeting the Src signalling pathway, has been approved for the treatment of idiopathic pulmonary fibrosis. However, it is still not known whether nintedanib ameliorates liver fibrosis by inhibiting inflammasome activation. Here, a carbon tetrachloride (CCl4)-induced liver fibrosis model was used to assess the anti-fibrotic efficacy of nintedanib in vivo. Lipopolysaccharide and ATP were used to activate nucleotide oligomerisation domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasomes in LX-2 cells, and the efficacy of nintedanib on NLRP3 inflammasome activation was evaluated. Moreover, we used Src-overexpressing and Src-downregulating lentiviruses to transfect LX-2 cells to explore the targets of nintedanib. Nintedanib attenuated inflammation and extracellular matrix accumulation in CCl4-induced fibrotic livers and reduced the expression of NLRP3, fibrotic makers, and the phosphorylation of Src, epidermal growth factor receptor (EGFR), AKT, ERK1/2 in LX-2 cells. Furthermore, nintedanib thwarted NLRP3 inflammasome activation by suppressing the phosphorylation of Src and its downstream signalling pathway and reducing reactive oxygen species production. Our study indicates that nintedanib effectively suppresses NLRP3 inflammasome activation and has the potential for the treatment of liver fibrosis.
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Affiliation(s)
- Ye-Ting Wu
- Department of Infectious Diseases, The Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
| | - Qi-Zhe Li
- Department of Sport Medicine, The Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
| | - Yi-Qi Wu
- The Second Clinical College of Hainan Medical University, Hainan, Haikou, China
| | - Mao Mu
- Department of Infectious Diseases, The Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
| | - Huan Wu
- Department of Infectious Diseases, The Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China
| | - Hai-Ying Tian
- Department of Ultrasound Medicine, Guizhou Provincial People's Hospital, Guiyang, Guizhou, China
| | - Xue-Ke Zhao
- Department of Infectious Diseases, The Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China.
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Ezhilarasan D, Najimi M. Quiescent hepatic stellate cell activation in liver fibrosis: Have we found the right trigger yet? Clin Res Hepatol Gastroenterol 2024; 48:102420. [PMID: 39002817 DOI: 10.1016/j.clinre.2024.102420] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/01/2024] [Accepted: 07/11/2024] [Indexed: 07/15/2024]
Affiliation(s)
- Devaraj Ezhilarasan
- Department of Pharmacology, Hepatology and Molecular Medicine Lab, Saveetha Dental College, Saveetha Institute of Medical and Technical Sciences, No.162, PH Rd, Chennai, Tamil Nadu 600 077, India.
| | - Mustapha Najimi
- Laboratory of Pediatric Hepatology and Cell Therapy, Institute of Experimental and Clinical Research (IREC), UCLouvain, Brussels, Belgium
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Wells RG. Liver fibrosis: Our evolving understanding. Clin Liver Dis (Hoboken) 2024; 23:e0243. [PMID: 38961878 PMCID: PMC11221862 DOI: 10.1097/cld.0000000000000243] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/11/2024] [Accepted: 03/29/2024] [Indexed: 07/05/2024] Open
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Crawford JM, Bioulac-Sage P, Hytiroglou P. Structure, Function and Responses to Injury. MACSWEEN'S PATHOLOGY OF THE LIVER 2024:1-95. [DOI: 10.1016/b978-0-7020-8228-3.00001-6] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/03/2025]
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Yoo SH, Nahm JH, Lee WK, Lee HW, Chang HY, Lee JI. Loss of Krüppel-like factor-10 facilitates the development of chemical-induced liver cancer in mice. Mol Med 2023; 29:156. [PMID: 37946098 PMCID: PMC10636809 DOI: 10.1186/s10020-023-00751-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2023] [Accepted: 10/30/2023] [Indexed: 11/12/2023] Open
Abstract
BACKGROUND Krüppel-like factor 10 (KLF10) is involved in a positive feedback loop that regulates transforming growth factor β (TGFβ) signaling, and TGFβ plays an important role in the pathogenesis of liver disease. Here, we investigated whether KLF10 deletion affects the development of liver fibrosis and hepatocellular carcinoma (HCC). METHODS We induced KLF10 deletion in C57BL/6 mice. Liver fibrosis was induced by feeding a diet high in fat and sucrose (high-fat diet [HFD]), whereas HCC was produced by intraperitoneal administration of N-diethylnitrosamine (DEN). An in vitro experiment was performed to evaluate the role of KLF10 in the cancer microenvironment using Hep3B and LX2 cells. An immunohistochemical study of KLF10 expression was performed using human HCC samples from 60 patients who had undergone liver resection. RESULTS KLF10 deletion resulted in an increased DEN-induced HCC burden with significant upregulation of SMAD2, although loss of KLF10 did not alter HFD-induced liver fibrosis. DEN-treated mice with KLF10 deletion exhibited increased levels of mesenchymal markers (N-cadherin and SNAI2) and tumor metastasis markers (matrix metalloproteinases 2 and 9). KLF10 depletion in Hep3B and LX2 cells using siRNA was associated with increased invasiveness. Compared with co-culture of KLF10-preserved Hep3B cells and KLF10-intact LX2 cells, co-culture of KLF10-preserved Hep3B cells and KLF10-depleted LX2 cells resulted in significantly enhanced invasion. Low KLF10 expression in resected human HCC specimens was associated with poor survival. CONCLUSION The results of this study suggest that loss of KLF10 facilitates liver cancer development with alteration in TGFβ signaling.
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Affiliation(s)
- Sung Hwan Yoo
- Department of Internal Medicine, Gangnam Severance Hospital, Yonsei University College of Medicine, 211 Eonju-Ro, Gangnam-Gu, Seoul, 06273, Republic of Korea
| | - Ji Hae Nahm
- Department of Pathology, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, 06273, Republic of Korea
| | - Woon Kyu Lee
- Laboratory of Developmental Genetics, Department of Biomedical Sciences, Inha University College of Medicine, Incheon, 22212, Republic of Korea
| | - Hyun Woong Lee
- Department of Internal Medicine, Gangnam Severance Hospital, Yonsei University College of Medicine, 211 Eonju-Ro, Gangnam-Gu, Seoul, 06273, Republic of Korea
| | - Hye Young Chang
- Medical Research Center, Gangnam Severance Hospital, Seoul, 06230, Republic of Korea
| | - Jung Il Lee
- Department of Internal Medicine, Gangnam Severance Hospital, Yonsei University College of Medicine, 211 Eonju-Ro, Gangnam-Gu, Seoul, 06273, Republic of Korea.
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Identification of Liver Fibrosis-Related MicroRNAs in Human Primary Hepatic Stellate Cells Using High-Throughput Sequencing. Genes (Basel) 2022; 13:genes13122201. [PMID: 36553468 PMCID: PMC9778123 DOI: 10.3390/genes13122201] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2022] [Revised: 11/19/2022] [Accepted: 11/21/2022] [Indexed: 11/25/2022] Open
Abstract
MicroRNAs (miRNAs) participate in hepatic stellate cell (HSC) activation, which drives liver fibrosis initiation and progression. We aimed to identify novel hepatic fibrosis targets using miRNA sequencing (miRNA-seq) of human primary HSCs. Surgically resected liver tissues were used to extract HSCs. Based on next-generation sequencing, miRNA-seq was performed on four pairs of HSCs before and after in vitro culture. Additionally, we compared our data with open access miRNA-seq data derived from fourteen cirrhotic and nine healthy liver tissues. Selected miRNAs associated with fibrosis were verified by quantitative real-time PCR. Target mRNAs of differentially expressed (DE) miRNAs were predicted to construct co-expression networks. We identified 230 DEmiRNAs (118 upregulated and 112 downregulated) upon HSC activation. Of the 17 miRNAs with the most significant differences in expression, liver disease-related miRNAs included miR-758-3p, miR-493-5p, miR-409-3p, miR-31-5p, miR-1268a, and miR-381-3p, which might play roles in hepatic fibrosis. Moreover, let-7g-5p, miR-107, miR-122-5p, miR-127-3p, miR-139-5p, miR-148a-3p, miR-194-5p, miR-215-5p, miR-26a-5p, miR-340-5p, miR-451a, and miR-99a-5p were common between our data and the publicly available sequencing data. A co-expression network comprising 1891 matched miRNA-mRNA pairs representing 138 DEmiRNAs and 1414 DEmRNAs was constructed. MiR-1268a and miR-665, possessing the richest target DEmRNAs, may be vital in HSC activation. The targeted genes were involved in collagen metabolism, extracellular matrix structural constituent, cytoskeletal protein binding, and cell adhesion. The miRNAs we identified may provide a basis and reference for the selection of diagnostic and therapeutic targets for hepatic fibrosis.
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Ilha M, Meira Martins LA, da Silveira Moraes K, Dias CK, Thomé MP, Petry F, Rohden F, Borojevic R, Trindade VMT, Klamt F, Barbé‐Tuana F, Lenz G, Guma FCR. Caveolin-1 influences mitochondrial plasticity and function in hepatic stellate cell activation. Cell Biol Int 2022; 46:1787-1800. [PMID: 35971753 PMCID: PMC9804617 DOI: 10.1002/cbin.11876] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/15/2022] [Revised: 03/21/2022] [Accepted: 05/26/2022] [Indexed: 01/05/2023]
Abstract
Caveolin-1 (Cav-1) is an integral membrane protein present in all organelles, responsible for regulating and integrating multiple signals as a platform. Mitochondria are extremely adaptable to external cues in chronic liver diseases, and expression of Cav-1 may affect mitochondrial flexibility in hepatic stellate cells (HSCs) activation. We previously demonstrated that exogenous expression of Cav-1 was sufficient to increase some classical markers of activation in HSCs. Here, we aimed to evaluate the influence of exogenous expression and knockdown of Cav-1 on regulating the mitochondrial plasticity, metabolism, endoplasmic reticulum (ER)-mitochondria distance, and lysosomal activity in HSCs. To characterize the mitochondrial, lysosomal morphology, and ER-mitochondria distance, we perform transmission electron microscope analysis. We accessed mitochondria and lysosomal networks and functions through a confocal microscope and flow cytometry. The expression of mitochondrial machinery fusion/fission genes was examined by real-time polymerase chain reaction. Total and mitochondrial cholesterol content was measured using Amplex Red. To define energy metabolism, we used the Oroboros system in the cells. We report that GRX cells with exogenous expression or knockdown of Cav-1 changed mitochondrial morphometric parameters, OXPHOS metabolism, ER-mitochondria distance, lysosomal activity, and may change the activation state of HSC. This study highlights that Cav-1 may modulate mitochondrial function and structural reorganization in HSC activation, being a potential candidate marker for chronic liver diseases and a molecular target for therapeutic intervention.
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Affiliation(s)
- Mariana Ilha
- Programa de Pós‐Graduação em Ciências Biológicas‐Bioquímica, Instituto de Ciências Básicas da SaúdeUniversidade Federal do Rio Grande do Sul – UFRGSPorto AlegreRio Grande do SulBrasil,Department of Clinical Nutrition, Institute of Public Health and Clinical NutritionUniversity of Eastern FinlandKuopioFinland
| | - Leo A. Meira Martins
- Programa de Pós‐Graduação em Ciências Biológicas‐Bioquímica, Instituto de Ciências Básicas da SaúdeUniversidade Federal do Rio Grande do Sul – UFRGSPorto AlegreRio Grande do SulBrasil,Departamento de Fisiologia, Instituto de Ciências Básicas da SaúdeUniversidade Federal do Rio Grande do Sul ‐ UFRGSPorto AlegreRio Grande do SulBrasil
| | - Ketlen da Silveira Moraes
- Departamento de Bioquímica, Instituto de Ciências Básicas da SaúdeUniversidade Federal do Rio Grande do Sul ‐ UFRGSPorto AlegreRio Grande do SulBrasil
| | - Camila K. Dias
- Programa de Pós‐Graduação em Ciências Biológicas‐Bioquímica, Instituto de Ciências Básicas da SaúdeUniversidade Federal do Rio Grande do Sul – UFRGSPorto AlegreRio Grande do SulBrasil
| | - Marcos P. Thomé
- Departamento de Biofísica e Centro de BiotecnologiaUniversidade Federal do Rio Grande do Sul ‐ UFRGSPorto AlegreRio Grande do SulBrasil
| | - Fernanda Petry
- Programa de Pós‐Graduação em Ciências Biológicas‐Bioquímica, Instituto de Ciências Básicas da SaúdeUniversidade Federal do Rio Grande do Sul – UFRGSPorto AlegreRio Grande do SulBrasil
| | - Francieli Rohden
- Programa de Pós‐Graduação em Ciências Biológicas‐Bioquímica, Instituto de Ciências Básicas da SaúdeUniversidade Federal do Rio Grande do Sul – UFRGSPorto AlegreRio Grande do SulBrasil
| | - Radovan Borojevic
- Centro de Medicina RegenerativaFaculdade Arthur Sa Earp Neto ‐ Faculdade de Medicina de PetrópolisRio de JaneiroBrasil
| | - Vera M. T. Trindade
- Departamento de Bioquímica, Instituto de Ciências Básicas da SaúdeUniversidade Federal do Rio Grande do Sul ‐ UFRGSPorto AlegreRio Grande do SulBrasil
| | - Fábio Klamt
- Departamento de Bioquímica, Instituto de Ciências Básicas da SaúdeUniversidade Federal do Rio Grande do Sul ‐ UFRGSPorto AlegreRio Grande do SulBrasil
| | - Florência Barbé‐Tuana
- Programa de Pós‐Graduação em Biologia Celular e MolecularEscola de Ciências da Pontifícia Universidade Católica do Rio Grande do Sul‐ PUCRSPorto AlegreRio Grande do SulBrasil
| | - Guido Lenz
- Departamento de Biofísica e Centro de BiotecnologiaUniversidade Federal do Rio Grande do Sul ‐ UFRGSPorto AlegreRio Grande do SulBrasil
| | - Fátima C. R. Guma
- Departamento de Bioquímica, Instituto de Ciências Básicas da SaúdeUniversidade Federal do Rio Grande do Sul ‐ UFRGSPorto AlegreRio Grande do SulBrasil,Centro de Microscopia e MicroanáliseUniversidade Federal do Rio Grande do Sul ‐ UFRGSPorto AlegreRio Grande do SulBrasil
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Liu S, Premont RT, Park KH, Rockey DC. β-PIX cooperates with GIT1 to regulate endothelial nitric oxide synthase in sinusoidal endothelial cells. Am J Physiol Gastrointest Liver Physiol 2022; 323:G511-G522. [PMID: 36044673 PMCID: PMC9639759 DOI: 10.1152/ajpgi.00034.2022] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/11/2022] [Revised: 08/11/2022] [Accepted: 08/24/2022] [Indexed: 01/31/2023]
Abstract
Previous studies have demonstrated that G protein-coupled receptor kinase interacting-1 protein (GIT1) associates with endothelial nitric oxide synthase (eNOS) to regulate nitric oxide production in sinusoidal endothelial cells (SECs). Here, we hypothesized that GIT1's tightly associated binding partner, β-PIX (p21-activated kinase-interacting exchange factor β, ARHGEF7) is specifically important in the regulation of eNOS activity. We examined β-PIX expression in normal rat liver by immunohistochemistry and explored β-PIX protein-protein interactions using immunoprecipitation and immunoblotting. The role of β-PIX in regulating eNOS enzymatic activity was studied in GIT1-deficient SECs. Finally, structural analysis of interaction sites in GIT1 and β-PIX required to regulate eNOS activity were mapped. β-PIX was expressed primarily in SECs in normal liver and was either absent or expressed at extremely low levels in other liver cells (stellate cells, Kupffer cells, and hepatocytes). β-PIX interacted with GIT1 and eNOS to form a trimolecular signaling module in normal SECs and was important in stimulating eNOS activity. Of note, GIT1-β-PIX interaction led to synergistic enhancement of eNOS activity, and β-PIX-driven increase in eNOS activity was GIT1 dependent. Disruption of β-PIX or GIT1 in normal SECs using β-PIX siRNA or GIT1-deficient SECs led to reduced eNOS activity. Finally, specific GIT1 domains [Spa2 homology domain (SHD) and synaptic localization domain (SLD), aa 331-596] and the β-PIX COOH terminal (aa 496-555) appeared to be critical in the regulation eNOS activity. The data indicate that β-PIX regulates eNOS phosphorylation and function in normal SECs and highlight the importance of the GIT1/β-PIX/eNOS trimolecular complex in normal liver SEC function.NEW & NOTEWORTHY β-PIX is a multidomain protein known to be a GIT1 binding partner. We report here that in the normal liver, the distribution and cellular localization of β-PIX are restricted largely to sinusoidal endothelial cells. Furthermore, β-PIX interacts with eNOS and GIT1 promotes eNOS activity and NO production and therefore exerts a novel posttranslational regulatory function on eNOS activity in sinusoidal endothelial cells. We also have identified specific molecular domains important in GIT1 and β-PIX's interaction with eNOS, which may represent novel therapeutic targets in the control of sinusoidal blood flow and intrahepatic resistance.
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Affiliation(s)
- Songling Liu
- Digestive Disease Research Center, Medical University of South Carolina, Charleston, South Carolina
| | - Richard T Premont
- Harrington Discovery Institute, University Hospitals Cleveland Medical Center, Cleveland, Ohio
- Institute for Transformative Molecular Medicine, Case Western Reserve University School of Medicine, Cleveland, Ohio
| | - Ki-Hoon Park
- Digestive Disease Research Center, Medical University of South Carolina, Charleston, South Carolina
| | - Don C Rockey
- Digestive Disease Research Center, Medical University of South Carolina, Charleston, South Carolina
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Pituitary Tumor-Transforming Gene 1/Delta like Non-Canonical Notch Ligand 1 Signaling in Chronic Liver Diseases. Int J Mol Sci 2022; 23:ijms23136897. [PMID: 35805898 PMCID: PMC9267054 DOI: 10.3390/ijms23136897] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2022] [Revised: 06/17/2022] [Accepted: 06/18/2022] [Indexed: 02/06/2023] Open
Abstract
The management of chronic liver diseases (CLDs) remains a challenge, and identifying effective treatments is a major unmet medical need. In the current review we focus on the pituitary tumor transforming gene (PTTG1)/delta like non-canonical notch ligand 1 (DLK1) axis as a potential therapeutic target to attenuate the progression of these pathological conditions. PTTG1 is a proto-oncogene involved in proliferation and metabolism. PTTG1 expression has been related to inflammation, angiogenesis, and fibrogenesis in cancer and experimental fibrosis. On the other hand, DLK1 has been identified as one of the most abundantly expressed PTTG1 targets in adipose tissue and has shown to contribute to hepatic fibrosis by promoting the activation of hepatic stellate cells. Here, we extensively analyze the increasing amount of information pointing to the PTTG1/DLK1 signaling pathway as an important player in the regulation of these disturbances. These data prompted us to hypothesize that activation of the PTTG1/DLK1 axis is a key factor upregulating the tissue remodeling mechanisms characteristic of CLDs. Therefore, disruption of this signaling pathway could be useful in the therapeutic management of CLDs.
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Balaphas A, Meyer J, Gameiro C, Frobert A, Giraud MN, Egger B, Bühler LH, Gonelle-Gispert C. Optimized Isolation and Characterization of C57BL/6 Mouse Hepatic Stellate Cells. Cells 2022; 11:1379. [PMID: 35563686 PMCID: PMC9102395 DOI: 10.3390/cells11091379] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2022] [Revised: 03/17/2022] [Accepted: 03/29/2022] [Indexed: 02/04/2023] Open
Abstract
To obtain meaningful results of hepatic stellate cell (HSC) function, it is crucial to use highly pure HSC populations. Our aim was to optimize HSC isolation from mice livers without exploiting the characteristically transient vitamin A autofluorescence of HSC. HSCs were isolated from C57BL/6 mice using a two-step collagenase digestion and Nycodenz gradient separation followed by CD11b-negative sorting step in order to remove contaminating macrophages and dendritic cells. Isolated cells were analyzed for yield, viability, purity, and potential new markers using immunofluorescence and flow cytometry. We obtained a yield of 350,595 ± 100,773 HSC per mouse liver and a viability of isolated cells of 92.4 ± 3.1%. We observed a low macrophage/dendritic cell contamination of 1.22 ± 0.54%. Using flow cytometry, we demonstrated that CD38 was expressed at the surface of HSC subpopulations and that all expressed intracellular markers specific for HSC in the liver. This isolation method, avoiding fluorescent activated cell sorting (FACS), allowed isolation of HSCs with high purity. Further, flow cytometry analysis suggests that CD38 may be a reliable marker of HSCs and may include subpopulations of HSCs without retinoid droplets.
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Affiliation(s)
- Alexandre Balaphas
- Division of Digestive Surgery, University Hospitals of Geneva, 1205 Geneva, Switzerland; (A.B.); (J.M.)
- Department of Surgery, Clinical Medicine Section, Faculty of Medicine, University of Geneva, 1206 Geneva, Switzerland
| | - Jeremy Meyer
- Division of Digestive Surgery, University Hospitals of Geneva, 1205 Geneva, Switzerland; (A.B.); (J.M.)
| | - Cécile Gameiro
- Flow Cytometry Core Facility, Faculty of Medicine, University of Geneva, 1206 Geneva, Switzerland;
| | - Aurélien Frobert
- Surgical Research Unit, Faculty of Science and Medicine, Section of Medicine, University of Fribourg, 1700 Fribourg, Switzerland; (A.F.); (M.-N.G.); (B.E.); (L.H.B.)
| | - Marie-Noëlle Giraud
- Surgical Research Unit, Faculty of Science and Medicine, Section of Medicine, University of Fribourg, 1700 Fribourg, Switzerland; (A.F.); (M.-N.G.); (B.E.); (L.H.B.)
| | - Bernhard Egger
- Surgical Research Unit, Faculty of Science and Medicine, Section of Medicine, University of Fribourg, 1700 Fribourg, Switzerland; (A.F.); (M.-N.G.); (B.E.); (L.H.B.)
| | - Leo H. Bühler
- Surgical Research Unit, Faculty of Science and Medicine, Section of Medicine, University of Fribourg, 1700 Fribourg, Switzerland; (A.F.); (M.-N.G.); (B.E.); (L.H.B.)
| | - Carmen Gonelle-Gispert
- Surgical Research Unit, Faculty of Science and Medicine, Section of Medicine, University of Fribourg, 1700 Fribourg, Switzerland; (A.F.); (M.-N.G.); (B.E.); (L.H.B.)
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Hasani Jusufi A, Trajkovska M, Popova-Jovanovska R, Calovska-Ivanova V, Ramadani A, Andreevski V. The Role and Significance of Non-invasive Methods, with a Particular Focus on Shear Wave Elastography in Hepatic Fibrosis Staging. Open Access Maced J Med Sci 2022. [DOI: 10.3889/oamjms.2022.9048] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/05/2022] Open
Abstract
Shear Wave Elastography (SWE) represents a new, non-invasive method, used in the diagnosis of diffuse liver diseases. The method has been widely used instead of liver biopsy - an invasive procedure with potential major risk complications. Compared to liver biopsy, SWE provides an examination of larger areas of the liver, thus providing better staging of hepatic fibrosis.
30 patients were included in the study on basis of previous clinical, biochemical, and ultrasound findings indicating a presence of a chronic liver lesion. Patients were divided into three groups: 6 patients with steatosis, 13 patients with viral hepatitis, and 11 patients with liver cirrhosis. Liver damage biochemical markers, serum markers of liver fibrosis, and SWE were determined in all patients. Statistical analysis revealed a positive correlation between SWE results, and the values of biochemical markers of the hepatic lesion, as well as serum markers of liver fibrosis.
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Yuan S, Wei C, Liu G, Zhang L, Li J, Li L, Cai S, Fang L. Sorafenib attenuates liver fibrosis by triggering hepatic stellate cell ferroptosis via HIF-1α/SLC7A11 pathway. Cell Prolif 2022; 55:e13158. [PMID: 34811833 PMCID: PMC8780895 DOI: 10.1111/cpr.13158] [Citation(s) in RCA: 246] [Impact Index Per Article: 82.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2021] [Revised: 08/17/2021] [Accepted: 08/31/2021] [Indexed: 12/27/2022] Open
Abstract
OBJECTIVES Evidences demonstrate that sorafenib alleviates liver fibrosis via inhibiting HSC activation and ECM accumulation. The underlying mechanism remains unclear. Ferroptosis, a novel programmed cell death, regulates diverse physiological/pathological processes. In this study, we aim to investigate the functional role of HSC ferroptosis in the anti-fibrotic effect of sorafenib. MATERIALS AND METHODS The effects of sorafenib on HSC ferroptosis and ECM expression were assessed in mouse model of liver fibrosis induced by CCl4 . In vitro, Fer-1 and DFO were used to block ferroptosis and then explored the anti-fibrotic effect of sorafenib by detecting α-SMA, COL1α1 and fibronectin proteins. Finally, HIF-1α siRNA, plasmid and stabilizers were applied to assess related signalling pathway. RESULTS Sorafenib attenuated liver injury and ECM accumulation in CCl4 -induced fibrotic livers, accompanied by reduction of SLC7A11 and GPX4 proteins. In sorafenib-treated HSC-T6 cells, ferroptotic events (depletion of SLC7A11, GPX4 and GSH; accumulation iron, ROS and MDA) were discovered. Intriguingly, these ferroptotic events were not appeared in hepatocytes or macrophages. Sorafenib-elicited HSC ferroptosis and ECM reduction were abrogated by Fer-1 and DFO. Additionally, both HIF-1α and SLC7A11 proteins were reduced in sorafenib-treated HSC-T6 cells. SLC7A11 was positively regulated by HIF-1α, inactivation of HIF-1α/SLC7A11 pathway was required for sorafenib-induced HSC ferroptosis, and elevation of HIF-1α could inhibit ferroptosis, ultimately limited the anti-fibrotic effect. CONCLUSIONS Sorafenib triggers HSC ferroptosis via HIF-1α/SLC7A11 signalling, which in turn attenuates liver injury and fibrosis.
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Affiliation(s)
- Siyu Yuan
- Department of PharmacyThe First Affiliated Hospital of Anhui Medical UniversityHefeiChina
| | - Can Wei
- Department of UrologyThe Second People's Hospital of HefeiHefeiChina
| | - Guofang Liu
- School of PharmacyAnhui University of Chinese MedicineHefeiChina
| | - Lijun Zhang
- School of PharmacyAnhui Medical UniversityHefeiChina
| | - Jiahao Li
- School of PharmacyAnhui University of Chinese MedicineHefeiChina
| | - Lingling Li
- Department of PharmacyThe First Affiliated Hospital of Anhui Medical UniversityHefeiChina
| | - Shiyi Cai
- School of PharmacyAnhui Medical UniversityHefeiChina
| | - Ling Fang
- Department of PharmacyThe First Affiliated Hospital of Anhui Medical UniversityHefeiChina
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15
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Lele S, Lee SD, Sarkar D, Levy MF. Purification and Isolation of Hepatic Stellate Cells. Methods Mol Biol 2022; 2455:93-101. [PMID: 35212989 PMCID: PMC8930280 DOI: 10.1007/978-1-0716-2128-8_9] [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: 06/14/2023]
Abstract
Quiescent human hepatic stellate cells (HSCs) serve as important reservoirs of fat-soluble vitamins in the body, namely vitamin A. In an activated form, HSCs are the drivers of fibrosis following chronic liver injury. In non-alcoholic steatohepatitis (NASH) specifically, activated HSCs are drivers of induction and progression of fibrogenesis. Isolation and purification of HSCs from donor liver samples provides an avenue to study HSCs and their fibrotic capabilities. Manual and chemical digestion of donor liver via dissection and Pronase, collagenase, and DNAse treatment creates a suspension of non-parenchymal liver cells. Quiescent HSCs can be further isolated from this suspension by density-gradient centrifugation in a 6%, 8%, 12%, and 15% arabinogalactan medium. After collection of HSCs from the low-density layers of the gradient, they can be grown on uncoated plastic. Rodent HSCs can also be isolated via density-gradient centrifugation. To isolate activated HSCs, liver tissue explants or established immortalized HSC lines can be utilized. Here, we described protocols for isolation of human and rodent HSCs.
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Affiliation(s)
- Sonia Lele
- School of Medicine, Virginia Commonwealth University, Richmond, VA, USA
| | - Seung Duk Lee
- Division of Transplant Surgery, Department of Surgery, Virginia Commonwealth University, Richmond, VA, USA
| | - Devanand Sarkar
- Department of Human and Molecular Genetics, School of Medicine, Virginia Commonwealth University, Richmond, VA, USA
- Massey Cancer Center, Virginia Commonwealth University, Richmond, VA, USA
- VCU Institute of Molecular Medicine, Virginia Commonwealth University, Richmond, VA, USA
| | - Marlon F Levy
- Division of Transplant Surgery, Department of Surgery, Virginia Commonwealth University, Richmond, VA, USA.
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16
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Bhattacharya D, Becker C, Readhead B, Goossens N, Novik J, Fiel MI, Cousens LP, Magnusson B, Backmark A, Hicks R, Dudley JT, Friedman SL. Repositioning of a novel GABA-B receptor agonist, AZD3355 (Lesogaberan), for the treatment of non-alcoholic steatohepatitis. Sci Rep 2021; 11:20827. [PMID: 34675338 PMCID: PMC8531016 DOI: 10.1038/s41598-021-99008-2] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/02/2021] [Accepted: 09/14/2021] [Indexed: 01/02/2023] Open
Abstract
Non-alcoholic steatohepatitis (NASH) is a rising health challenge, with no approved drugs. We used a computational drug repositioning strategy to uncover a novel therapy for NASH, identifying a GABA-B receptor agonist, AZD3355 (Lesogaberan) previously evaluated as a therapy for esophageal reflux. AZD3355's potential efficacy in NASH was tested in human stellate cells, human precision cut liver slices (hPCLS), and in vivo in a well-validated murine model of NASH. In human stellate cells AZD3355 significantly downregulated profibrotic gene and protein expression. Transcriptomic analysis of these responses identified key regulatory nodes impacted by AZD3355, including Myc, as well as MAP and ERK kinases. In PCLS, AZD3355 down-regulated collagen1α1, αSMA and TNF-α mRNAs as well as secreted collagen1α1. In vivo, the drug significantly improved histology, profibrogenic gene expression, and tumor development, which was comparable to activity of obeticholic acid in a robust mouse model of NASH, but awaits further testing to determine its relative efficacy in patients. These data identify a well-tolerated clinical stage asset as a novel candidate therapy for human NASH through its hepatoprotective, anti-inflammatory and antifibrotic mechanisms of action. The approach validates computational methods to identify novel therapies in NASH in uncovering new pathways of disease development that can be rapidly translated into clinical trials.
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Affiliation(s)
- Dipankar Bhattacharya
- Division of Liver Diseases, Icahn School of Medicine at Mount Sinai, Box 1123, 1425 Madison Ave. Room 1170, New York, NY, 10029, USA
| | - Christine Becker
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA
- Division of Clinical Immunology, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Benjamin Readhead
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA
- Arizona State University-Banner Neurodegenerative Disease Research Center, Arizona, USA
| | - Nicolas Goossens
- Division of Liver Diseases, Icahn School of Medicine at Mount Sinai, Box 1123, 1425 Madison Ave. Room 1170, New York, NY, 10029, USA
- Division of Gastroenterology, Geneva University Hospital, Geneva, Switzerland
| | - Jacqueline Novik
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Maria Isabel Fiel
- Department of Pathology, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Leslie P Cousens
- Emerging Innovations, Discovery Sciences, R&D, AstraZeneca, Boston, MA, USA
| | - Björn Magnusson
- Discovery Biology, Discovery Sciences, R&D, AstraZeneca, Gothenburg, Sweden
| | - Anna Backmark
- Discovery Biology, Discovery Sciences, R&D, AstraZeneca, Gothenburg, Sweden
| | - Ryan Hicks
- BioPharmaceuticals R&D Cell Therapy, Research and Early Development, Cardiovascular, Renal and Metabolism (CVRM), BioPharmaceuticals R&D, AstraZeneca, Gothenburg, Sweden
| | - Joel T Dudley
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Scott L Friedman
- Division of Liver Diseases, Icahn School of Medicine at Mount Sinai, Box 1123, 1425 Madison Ave. Room 1170, New York, NY, 10029, USA.
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Stefanovic B, Stefanovic L, Manojlovic Z. Imaging of type I procollagen biosynthesis in cells reveals biogenesis in highly organized bodies; Collagenosomes. Matrix Biol Plus 2021; 12:100076. [PMID: 34278289 PMCID: PMC8261018 DOI: 10.1016/j.mbplus.2021.100076] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2021] [Revised: 06/18/2021] [Accepted: 06/18/2021] [Indexed: 10/26/2022] Open
Abstract
Mechanistic aspects of type I procollagen biosynthesis in cells are poorly understood. To provide more insight into this process we designed a system to directly image type I procollagen biogenesis by co-expression of fluorescently labeled full size procollagen α1(I) and one α2(I) polypeptides. High resolution images show that collagen α1(I) and α2(I) polypeptides are produced in coordination in discrete structures on the ER membrane, which we termed the collagenosomes. Collagenosomes are disk shaped bodies, 0.5-1 μM in diameter and 200-400 nm thick, in the core of which folding of procollagen takes place. Collagenosomes are intimately associated with the ER membrane and their formation requires intact translational machinery, suggesting that they are the sites of nascent procollagen biogenesis. Collagenosomes show little co-localization with the COPII transport vesicles, which export type I procollagen from the ER, suggesting that these two structures are distinct. LARP6 is the protein which regulates translation of type I collagen mRNAs. The characteristic organization of collagenosomes depends on binding of LARP6 to collagen mRNAs. Without LARP6 regulation, collagenosomes are poorly organized and the folding of α1(I) and α2(I) polypeptides into procollagen in their cores is diminished. This indicates that formation of collagenosomes is dependent on regulated translation of collagen mRNAs. In live cells the size, number and shape of collagenosomes show little change within several hours, suggesting that they are stable structures of type I procollagen biogenesis. This is the first report of structural organization of type I collagen biogenesis in collagenosomes, while the fluorescent reporter system based on simultaneous imaging of both type I collagen polypeptides will enable the detailed elucidation of their structure and function.
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Affiliation(s)
- Branko Stefanovic
- Department of Biomedical Sciences and Translational Science Laboratory, College of Medicine, Florida State University, 1115 West Call Street, Tallahassee, FL 32306, USA
| | - Lela Stefanovic
- Department of Biomedical Sciences and Translational Science Laboratory, College of Medicine, Florida State University, 1115 West Call Street, Tallahassee, FL 32306, USA
| | - Zarko Manojlovic
- Keck School of Medicine of University of Southern California, 1450 Biggy Street, NRT 4510, Los Angeles, CA 90033, USA
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Sufleţel RT, Melincovici CS, Gheban BA, Toader Z, Mihu CM. Hepatic stellate cells - from past till present: morphology, human markers, human cell lines, behavior in normal and liver pathology. ROMANIAN JOURNAL OF MORPHOLOGY AND EMBRYOLOGY 2021; 61:615-642. [PMID: 33817704 PMCID: PMC8112759 DOI: 10.47162/rjme.61.3.01] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 01/18/2023]
Abstract
Hepatic stellate cell (HSC), initially analyzed by von Kupffer, in 1876, revealed to be an extraordinary mesenchymal cell, essential for both hepatocellular function and lesions, being the hallmark of hepatic fibrogenesis and carcinogenesis. Apart from their implications in hepatic injury, HSCs play a vital role in liver development and regeneration, xenobiotic response, intermediate metabolism, and regulation of immune response. In this review, we discuss the current state of knowledge regarding HSCs morphology, human HSCs markers and human HSC cell lines. We also summarize the latest findings concerning their roles in normal and liver pathology, focusing on their impact in fibrogenesis, chronic viral hepatitis and liver tumors.
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Affiliation(s)
- Rada Teodora Sufleţel
- Discipline of Histology, Department of Morphological Sciences, Iuliu Haţieganu University of Medicine and Pharmacy, Cluj-Napoca, Romania;
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19
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Payen VL, Lavergne A, Alevra Sarika N, Colonval M, Karim L, Deckers M, Najimi M, Coppieters W, Charloteaux B, Sokal EM, El Taghdouini A. Single-cell RNA sequencing of human liver reveals hepatic stellate cell heterogeneity. JHEP Rep 2021; 3:100278. [PMID: 34027339 PMCID: PMC8121977 DOI: 10.1016/j.jhepr.2021.100278] [Citation(s) in RCA: 69] [Impact Index Per Article: 17.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/30/2020] [Revised: 02/11/2021] [Accepted: 02/28/2021] [Indexed: 02/07/2023] Open
Abstract
Background & Aims The multiple vital functions of the human liver are performed by highly specialised parenchymal and non-parenchymal cells organised in complex collaborative sinusoidal units. Although crucial for homeostasis, the cellular make-up of the human liver remains to be fully elucidated. Here, single-cell RNA-sequencing was used to unravel the heterogeneity of human liver cells, in particular of hepatocytes (HEPs) and hepatic stellate cells (HSCs). Method The transcriptome of ~25,000 freshly isolated human liver cells was profiled using droplet-based RNA-sequencing. Recently published data sets and RNA in situ hybridisation were integrated to validate and locate newly identified cell populations. Results In total, 22 cell populations were annotated that reflected the heterogeneity of human parenchymal and non-parenchymal liver cells. More than 20,000 HEPs were ordered along the portocentral axis to confirm known, and reveal previously undescribed, zonated liver functions. The existence of 2 subpopulations of human HSCs with unique gene expression signatures and distinct intralobular localisation was revealed (i.e. portal and central vein-concentrated GPC3+ HSCs and perisinusoidally located DBH+ HSCs). In particular, these data suggest that, although both subpopulations collaborate in the production and organisation of extracellular matrix, GPC3+ HSCs specifically express genes involved in the metabolism of glycosaminoglycans, whereas DBH+ HSCs display a gene signature that is reminiscent of antigen-presenting cells. Conclusions This study highlights metabolic zonation as a key determinant of HEP transcriptomic heterogeneity and, for the first time, outlines the existence of heterogeneous HSC subpopulations in the human liver. These findings call for further research on the functional implications of liver cell heterogeneity in health and disease. Lay summary This study resolves the cellular landscape of the human liver in an unbiased manner and at high resolution to provide new insights into human liver cell biology. The results highlight the physiological heterogeneity of human hepatic stellate cells.
A cell atlas from the near-native transcriptome of >25,000 human liver cells is presented. Hepatocytes were ordered along the portocentral axis to reveal previously undescribed gene expression patterns and zonated liver functions. Two subpopulations of human hepatic stellate cells (HSCs) are reported, characterised by different spatial distribution in the native tissue. Characteristic gene signatures of HSC subpopulations are suggestive of far-reaching functional differences.
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Key Words
- BSA, bovine serum albumin
- CC, cholangiocyte
- CV, central vein
- DEG, differentially expressed gene
- EC, endothelial cell
- ECM, extracellular matrix
- Extracellular matrix
- FFPE, formaldehyde-fixed paraffin embedded
- GAG, glycosaminoglycan
- GEO, Gene Expression Omnibus
- GO, gene ontology
- HEP, hepatocyte
- HLA, human leukocyte antigen
- HRP, horseradish peroxidase
- HSC, hepatic stellate cell
- Hepatocyte
- ISH, in situ hybridisation
- KLR, killer lectin-like receptor
- LP, lymphoid cell
- Liver cell atlas
- MP, macrophage
- MZ, midzonal
- PC, pericentral
- PP, periportal
- PV, portal vein
- TBS, Tris buffered saline
- TSA, tyramide signal amplification
- UMAP, uniform manifold approximation and projection
- UMI, unique molecular identifier
- VIM, vimentin
- Zonation
- scRNA-seq, single-cell RNA-sequencing
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Affiliation(s)
- Valéry L. Payen
- Laboratory of Pediatric Hepatology and Cell Therapy (PEDI), IREC Institute, Université catholique de Louvain, Brussels, Belgium
- Laboratory of Advanced Drug Delivery and Biomaterials (ADDB), LDRI Institute, Université catholique de Louvain, Brussels, Belgium
| | - Arnaud Lavergne
- Genomics Platform, GIGA Institute, Université de Liège, Liège, Belgium
| | - Niki Alevra Sarika
- Laboratory of Pediatric Hepatology and Cell Therapy (PEDI), IREC Institute, Université catholique de Louvain, Brussels, Belgium
- Laboratory of Advanced Drug Delivery and Biomaterials (ADDB), LDRI Institute, Université catholique de Louvain, Brussels, Belgium
| | - Megan Colonval
- Genomics Platform, GIGA Institute, Université de Liège, Liège, Belgium
| | - Latifa Karim
- Genomics Platform, GIGA Institute, Université de Liège, Liège, Belgium
| | - Manon Deckers
- Genomics Platform, GIGA Institute, Université de Liège, Liège, Belgium
| | - Mustapha Najimi
- Laboratory of Pediatric Hepatology and Cell Therapy (PEDI), IREC Institute, Université catholique de Louvain, Brussels, Belgium
| | - Wouter Coppieters
- Genomics Platform, GIGA Institute, Université de Liège, Liège, Belgium
| | | | - Etienne M. Sokal
- Laboratory of Pediatric Hepatology and Cell Therapy (PEDI), IREC Institute, Université catholique de Louvain, Brussels, Belgium
- Corresponding authors. Address: Laboratory of Pediatric Hepatology and Cell Therapy (PEDI), IREC Institute, Université catholique de Louvain, Avenue Mounier 52 Box B1.52.03, 1200 Brussels, Belgium.
| | - Adil El Taghdouini
- Laboratory of Pediatric Hepatology and Cell Therapy (PEDI), IREC Institute, Université catholique de Louvain, Brussels, Belgium
- Corresponding authors. Address: Laboratory of Pediatric Hepatology and Cell Therapy (PEDI), IREC Institute, Université catholique de Louvain, Avenue Mounier 52 Box B1.52.03, 1200 Brussels, Belgium.
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Abstract
Hepatic stellate cells (HSCs) are resident non-parenchymal liver pericytes whose plasticity enables them to regulate a remarkable range of physiologic and pathologic responses. To support their functions in health and disease, HSCs engage pathways regulating carbohydrate, mitochondrial, lipid, and retinoid homeostasis. In chronic liver injury, HSCs drive hepatic fibrosis and are implicated in inflammation and cancer. To do so, the cells activate, or transdifferentiate, from a quiescent state into proliferative, motile myofibroblasts that secrete extracellular matrix, which demands rapid adaptation to meet a heightened energy need. Adaptations include reprogramming of central carbon metabolism, enhanced mitochondrial number and activity, endoplasmic reticulum stress, and liberation of free fatty acids through autophagy-dependent hydrolysis of retinyl esters that are stored in cytoplasmic droplets. As an archetype for pericytes in other tissues, recognition of the HSC's metabolic drivers and vulnerabilities offer the potential to target these pathways therapeutically to enhance parenchymal growth and modulate repair.
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Affiliation(s)
- Parth Trivedi
- Division of Liver Diseases, The Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA
| | - Shuang Wang
- Division of Liver Diseases, The Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA
| | - Scott L Friedman
- Division of Liver Diseases, The Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA.
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21
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Aramchol downregulates stearoyl CoA-desaturase 1 in hepatic stellate cells to attenuate cellular fibrogenesis. JHEP Rep 2021; 3:100237. [PMID: 34151243 PMCID: PMC8189934 DOI: 10.1016/j.jhepr.2021.100237] [Citation(s) in RCA: 34] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/24/2020] [Revised: 01/13/2021] [Accepted: 01/17/2021] [Indexed: 02/07/2023] Open
Abstract
Background & Aims Aramchol is a fatty acid-bile acid conjugate that reduces liver fat content and is being evaluated in a phase III clinical trial for non-alcoholic steatohepatitis (NASH). Aramchol attenuates NASH in mouse models and decreases steatosis by downregulating the fatty acid synthetic enzyme stearoyl CoA desaturase 1 (SCD1) in hepatocytes. Although hepatic stellate cells (HSCs) also store lipids as retinyl esters, the impact of Aramchol in this cell type is unknown. Methods We investigated the effects of Aramchol on a human HSC line (LX-2), primary human HSCs (phHSCs), and primary human hepatocytes (phHeps). Results In LX-2 and phHSCs, 10 μM Aramchol significantly reduced SCD1 mRNA while inducing PPARG (PPARγ) mRNA, with parallel changes in the 2 proteins; ACTA2, COL1A1, β-PDGFR (bPDGFR) mRNAs were also significantly reduced in LX-2. Secretion of collagen 1 (Col1α1) was inhibited by 10 μM Aramchol. SCD1 knockdown in LX-2 cells phenocopied the effect of Aramchol by reducing fibrogenesis, and addition of Aramchol to these cells did not rescue fibrogenic gene expression. Conversely, in LX-2 overexpressing SCD1, Aramchol no longer suppressed fibrogenic gene expression. The drug also induced genes in LX-2 that promote cholesterol efflux and inhibited ACAT2, which catalyses cholesterol synthesis. In phHeps, Aramchol also reduced SCD1 and increased PPARG mRNA expression. Conclusions Aramchol downregulates SCD1 and elevates PPARG in HSCs, reducing COL1A1 and ACTA2 mRNAs and COL1A1 secretion. These data suggest a direct inhibitory effect of Aramchol in HSCs through SCD1 inhibition, as part of a broader impact on both fibrogenic genes as well as mediators of cholesterol homeostasis. These findings illustrate novel mechanisms of Aramchol activity, including potential antifibrotic activity in patients with NASH and fibrosis. Lay summary In this study, we have explored the potential activity of Aramchol, a drug currently in clinical trials for fatty liver disease, in blocking fibrosis, or scarring, by hepatic stellate cells, the principal collagen-producing (i.e. fibrogenic) cell type in liver injury. In both isolated human hepatic stellate cells and in a human hepatic stellate cell line, the drug suppresses the key fat-producing enzyme, stearoyl CoA desaturase 1 (SCD1), which leads to reduced expression of genes and proteins associated with hepatic fibrosis, while inducing the protective gene, PPARγ. The drug loses activity when SCD1 is already reduced by gene knockdown, reinforcing the idea that inhibition of SCD1 is a main mode of activity for Aramchol. These findings strengthen the rationale for testing Aramchol in patients with NASH.
The antifibrotic activity of Aramchol was assessed in human hepatic stellate cells (HSCs). Aramchol reduces fibrogenic gene expression by inhibiting SCD1 and inducing PPARγ. Aramchol inhibits pathways that increase HSC cholesterol content. The antifibrotic activity of Aramchol reinforces its potential efficacy in human NASH.
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Key Words
- ABCA1, ATP-binding cassette transporter 1
- EMT, epithelial-mesenchymal-transition
- Fatty liver disease
- Fibrosis
- GSEA, gene set enrichment analysis
- GSH, glutathione
- GSSG, glutathione disulfide
- HRP, horse radish peroxidase
- HSC, hepatic stellate cell
- Hepatic fibrosis
- Hh, Hedgehog
- MCD, methionine-choline depleted diet
- MMP-2, matrix metalloproteinase 2
- MUFAs, monounsaturated fatty acids
- NAFLD, non-alcoholic fatty liver disease
- NASH, non-alcoholic steatohepatitis
- Non-alcoholic steatohepatitis
- PDMS, polydimethylsiloxane
- PPAR, peroxisome proliferator-activated receptor
- SCD1, stearoyl CoA-desaturase 1
- SMA, smooth muscle actin
- TAA, thioacetamide
- phHSCs, primary human hepatic stellate cells
- phHeps, primary human hepatocytes
- siRNA, small inhibitory RNA
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Chang SN, Kim SH, Dey DK, Park SM, Nasif O, Bajpai VK, Kang SC, Lee J, Park JG. 5-O-Demethylnobiletin Alleviates CCl 4-Induced Acute Liver Injury by Equilibrating ROS-Mediated Apoptosis and Autophagy Induction. Int J Mol Sci 2021; 22:1083. [PMID: 33499185 PMCID: PMC7865239 DOI: 10.3390/ijms22031083] [Citation(s) in RCA: 35] [Impact Index Per Article: 8.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/04/2020] [Revised: 01/18/2021] [Accepted: 01/19/2021] [Indexed: 01/08/2023] Open
Abstract
Polymethoxyflavanoids (PMFs) have exhibited a vast array of therapeutic biological properties. 5-O-Demethylnobiletin (5-DN) is one such PMF having anti-inflammatory activity, yet its role in hepatoprotection has not been studied before. Results from in vitro study revealed that 5-DN did not exert a high level of cytotoxicity on HepG2 cells at 40 μM, and it was able to rescue HepG2 cell death induced by carbon tetrachloride (CCl4). Subsequently, we investigated acute liver injury on BALB/c mice induced by CCl4 through the intraperitoneal injection of 1 mL/kg CCl4 and co-administration of 5-DN at (1 and 2 mg/kg) by oral gavage for 15 days. The results illustrated that treatment with 5-DN attenuated CCl4-induced elevated serum aminotransferase (AST)/alanine aminotransferase (ALT) ratio and significantly ameliorated severe hepatic damage such as inflammation and fibrosis evidenced through lesser aberrations in the liver histology of 5-DN dose groups. Additionally, 5-DN efficiently counteracted and equilibrated the production of ROS accelerated by CCl4 and dramatically downregulated the expression of CYP2E1 vitally involved in converting CCl4 to toxic free radicals and also enhanced the antioxidant enzymes. 5-DN treatment also inhibited cell proliferation and inflammatory pathway abnormally regulated by CCl4 treatment. Furthermore, the apoptotic response induced by CCl4 treatment was remarkably reduced by enhanced Bcl-2 expression and noticeable reduction in Bax, Bid, cleaved caspase 3, caspase 9, and apaf-1 expression. 5-DN treatment also induced the conversion of LC3 and promoted the autophagic flux. Conclusively, 5-DN exhibited hepatoprotective effects in vitro and in vivo and prevented liver fibrosis induced by CCl4.
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Affiliation(s)
- Sukkum Ngullie Chang
- Department of Biotechnology, Daegu University, Gyeongsan 38453, Korea; (S.N.C.); (D.K.D.); (S.C.K.)
- Advanced Bio Convergence Center (ABCC), Pohang Technopark Foundation, Pohang 37668, Korea; (S.H.K.); (S.M.P.)
| | - Se Ho Kim
- Advanced Bio Convergence Center (ABCC), Pohang Technopark Foundation, Pohang 37668, Korea; (S.H.K.); (S.M.P.)
- School of Chemical Engineering, Yeungnam University, Gyeongsan 38541, Korea
| | - Debasish Kumar Dey
- Department of Biotechnology, Daegu University, Gyeongsan 38453, Korea; (S.N.C.); (D.K.D.); (S.C.K.)
| | - Seon Min Park
- Advanced Bio Convergence Center (ABCC), Pohang Technopark Foundation, Pohang 37668, Korea; (S.H.K.); (S.M.P.)
| | - Omaima Nasif
- Department of Physiology, College of Medicine, King Saud University (Medical City), King Khalid University Hospital, P.O. Box 2925, Riyadh 11461, Saudi Arabia;
| | - Vivek K. Bajpai
- Department of Energy and Materials Engineering, Dongguk University-Seoul, 30 Pildong-ro 1-gil, Seoul 04620, Korea
| | - Sun Chul Kang
- Department of Biotechnology, Daegu University, Gyeongsan 38453, Korea; (S.N.C.); (D.K.D.); (S.C.K.)
| | - Jintae Lee
- School of Chemical Engineering, Yeungnam University, Gyeongsan 38541, Korea
| | - Jae Gyu Park
- Advanced Bio Convergence Center (ABCC), Pohang Technopark Foundation, Pohang 37668, Korea; (S.H.K.); (S.M.P.)
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23
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Su S, Cao S, Liu J, Xu X. Mitochondria-specific delivery system for targeted regulation of mitochondrial gene expression. STAR Protoc 2021; 2:100275. [PMID: 33511358 PMCID: PMC7817499 DOI: 10.1016/j.xpro.2020.100275] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022] Open
Abstract
Targeted regulation of mitochondrial gene expression is challenging due to the lack of a mitochondria-specific delivery system. We have previously developed various stimuli-responsive nanoparticle (NP)-based delivery systems to transport nucleic acids for regulation of target gene expression. This protocol describes the design and preparation of an NP platform for mitochondria-specific gene delivery (mito-NP). We use mito-NP in primary liver fibroblasts that are transplanted into mice. Mito-NP can be used to deliver various nucleic acid therapeutics and to treat mitochondria-regulated diseases. For complete details on the use and execution of this protocol, please refer to Zhao et al. (2020).
A protocol for mitochondria-specific gene delivery system is described The delivery system achieves targeted regulation of mitochondrial gene expression This protocol is applicable to mitochondria-specific delivery of various nucleic acids
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Affiliation(s)
- Shicheng Su
- Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, China.,Medical Research Center, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, China
| | - Shuwen Cao
- Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, China.,Medical Research Center, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, China
| | - Jiayu Liu
- Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, China.,Medical Research Center, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, China
| | - Xiaoding Xu
- Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, China.,Medical Research Center, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, China
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24
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Wang Z, Cheng ZX, Abrams ST, Lin ZQ, Yates E, Yu Q, Yu WP, Chen PS, Toh CH, Wang GZ. Extracellular histones stimulate collagen expression in vitro and promote liver fibrogenesis in a mouse model via the TLR4-MyD88 signaling pathway. World J Gastroenterol 2020; 26:7513-7527. [PMID: 33384551 PMCID: PMC7754552 DOI: 10.3748/wjg.v26.i47.7513] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/27/2020] [Revised: 11/08/2020] [Accepted: 12/06/2020] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND Liver fibrosis progressing to liver cirrhosis and hepatic carcinoma is very common and causes more than one million deaths annually. Fibrosis develops from recurrent liver injury but the molecular mechanisms are not fully understood. Recently, the TLR4-MyD88 signaling pathway has been reported to contribute to fibrosis. Extracellular histones are ligands of TLR4 but their roles in liver fibrosis have not been investigated. AIM To investigate the roles and potential mechanisms of extracellular histones in liver fibrosis. METHODS In vitro, LX2 human hepatic stellate cells (HSCs) were treated with histones in the presence or absence of non-anticoagulant heparin (NAHP) for neutralizing histones or TLR4-blocking antibody. The resultant cellular expression of collagen I was detected using western blotting and immunofluorescent staining. In vivo, the CCl4-induced liver fibrosis model was generated in male 6-week-old ICR mice and in TLR4 or MyD88 knockout and parental mice. Circulating histones were detected and the effect of NAHP was evaluated. RESULTS Extracellular histones strongly stimulated LX2 cells to produce collagen I. Histone-enhanced collagen expression was significantly reduced by NAHP and TLR4-blocking antibody. In CCl4-treated wild type mice, circulating histones were dramatically increased and maintained high levels during the duration of fibrosis-induction. Injection of NAHP not only reduced alanine aminotransferase and liver injury scores, but also significantly reduced fibrogenesis. Since the TLR4-blocking antibody reduced histone-enhanced collagen I production in HSC, the CCl4 model with TLR4 and MyD88 knockout mice was used to demonstrate the roles of the TLR4-MyD88 signaling pathway in CCl4-induced liver fibrosis. The levels of liver fibrosis were indeed significantly reduced in knockout mice compared to wild type parental mice. CONCLUSION Extracellular histones potentially enhance fibrogenesis via the TLR4-MyD88 signaling pathway and NAHP has therapeutic potential by detoxifying extracellular histones.
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Affiliation(s)
- Zhi Wang
- Department of Pathology and Pathophysiology, Medical School, Southeast University, Nanjing 210009, Jiangsu Province, China
- Department of Gastroenterology, Zhongda Hospital, Nanjing 210009, Jiangsu Province, China
| | - Zhen-Xing Cheng
- Department of Pathology and Pathophysiology, Medical School, Southeast University, Nanjing 210009, Jiangsu Province, China
- Department of Clinical Infection, Microbiology and Immunology, University of Liverpool, Liverpool L69 7BE, United Kingdom
- Department of Gastroenterology, The First Affiliated Hospital, Anhui Medical University, Hefei 230032, Anhui Province, China
| | - Simon T Abrams
- Department of Clinical Infection, Microbiology and Immunology, University of Liverpool, Liverpool L69 7BE, United Kingdom
| | - Zi-Qi Lin
- Department of Integrated Traditional Chinese and Western Medicine, Sichuan Provincial Pancreatitis Centre and West China-Liverpool Biomedical Research Centre, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
| | - Edwin Yates
- Department of Integrative Biology, University of Liverpool, Liverpool L69 7ZB, United Kingdom
| | - Qian Yu
- Department of Gastroenterology, Zhongda Hospital, Nanjing 210009, Jiangsu Province, China
| | - Wei-Ping Yu
- Department of Pathology and Pathophysiology, Medical School, Southeast University, Nanjing 210009, Jiangsu Province, China
| | - Ping-Sheng Chen
- Department of Pathology and Pathophysiology, Medical School, Southeast University, Nanjing 210009, Jiangsu Province, China
| | - Cheng-Hock Toh
- Department of Clinical Infection, Microbiology and Immunology, University of Liverpool, Liverpool L69 7BE, United Kingdom
- Roald Dahl Haemostasis & Thrombosis Ctr, Royal Liverpool University Hospital, Liverpool L69 7BE, United Kingdom
| | - Guo-Zheng Wang
- Department of Clinical Infection, Microbiology and Immunology, University of Liverpool, Liverpool L69 7BE, United Kingdom
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25
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Li S, Zhao W, Zhao Z, Cheng B, Li S, Liu C. Levistilide A reverses rat hepatic fibrosis by suppressing angiotensin II‑induced hepatic stellate cells activation. Mol Med Rep 2020; 22:2191-2198. [PMID: 32705207 PMCID: PMC7411401 DOI: 10.3892/mmr.2020.11326] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2018] [Accepted: 03/26/2020] [Indexed: 12/14/2022] Open
Abstract
The renin angiotensin system (RAS) serves an important role in the development of hepatic fibrosis. Therefore, the present study investigated the effect of levistilide A (Lev A) on hepatic fibrosis via regulation of RAS. The effects of Lev A on the proliferation and activation of hepatic stellate cells (HSCs) were measured using a 5-ethynyl-2′-deoxyuridine assay, western blot analysis and immunofluorescence. The in vivo anti-hepatic fibrosis effect of Lev A was examined using a CCL4-induced rat fibrosis model. Lev A significantly prohibited angiotensin (Ang) II-induced proliferation of HSCs, and overexpression of smooth muscle α-actin (α-SMA) and F-actin in HSCs. Lev A partly reversed Ang II-induced angiotensin type 1 receptor (AT1R) upregulation and ERK and c-Jun phosphorylation. In CCL4-induced hepatic fibrosis rats, Lev A treatment significantly decreased the expression of collagen, α-SMA and hydroxyproline in rat liver, and improved liver functions. Lev A treatment also significantly inhibited the CCL4-induced increase in plasma Ang II, and upregulation of AT1R and phosphorylated ERK in rat liver. In conclusion, Lev A is a potential agent for the treatment of hepatic fibrosis by suppressing Ang II/AT1R/ERK/c-Jun activation in HSCs.
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Affiliation(s)
- Shu Li
- Department of Gastroenterology, Baoshan Branch, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201900, P.R. China
| | - Wei Zhao
- Institute of Liver Diseases, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, P.R. China
| | - Zhimin Zhao
- Institute of Liver Diseases, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, P.R. China
| | - Binbin Cheng
- Department of Tradition Chinese Medicine, Changhai Hospital, Second Military Medical University, Shanghai 200433, P.R. China
| | - Shuang Li
- Department of Gastroenterology, Baoshan Branch, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201900, P.R. China
| | - Chenghai Liu
- Institute of Liver Diseases, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, P.R. China
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26
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Ishay Y, Nachman D, Khoury T, Ilan Y. The role of the sphingolipid pathway in liver fibrosis: an emerging new potential target for novel therapies. Am J Physiol Cell Physiol 2020; 318:C1055-C1064. [PMID: 32130072 DOI: 10.1152/ajpcell.00003.2020] [Citation(s) in RCA: 22] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Sphingolipids (SL) are a family of bioactive lipids and a major cellular membrane structural component. SLs include three main compounds: ceramide (Cer), sphingosine (Sp), and sphingosine-1-phosphate (S-1P), all of which have emerging roles in biological functions in cells, especially in the liver. They are under investigation in various liver diseases, including cirrhosis and end-stage liver disease. In this review, we provide an overview on the role of SLs in liver pathobiology and focus on their potential role in the development of hepatic fibrosis. We describe recent evidence and suggest SLs are a promising potential therapeutic target for the treatment of liver disease and fibrosis.
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Affiliation(s)
- Yuval Ishay
- Department of Internal Medicine A, Hadassah-Hebrew University Medical Center, Jerusalem, Israel
| | - Dean Nachman
- Department of Internal Medicine A, Hadassah-Hebrew University Medical Center, Jerusalem, Israel
| | - Tawfik Khoury
- Gastroenterology and Liver Units, Department of Medicine, Hadassah-Hebrew University Medical Center, Jerusalem, Israel
| | - Yaron Ilan
- Gastroenterology and Liver Units, Department of Medicine, Hadassah-Hebrew University Medical Center, Jerusalem, Israel
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27
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Lee DY, Yun SM, Song MY, Ji SD, Son JG, Kim EH. Administration of Steamed and Freeze-Dried Mature Silkworm Larval Powder Prevents Hepatic Fibrosis and Hepatocellular Carcinogenesis by Blocking TGF-β/STAT3 Signaling Cascades in Rats. Cells 2020; 9:E568. [PMID: 32121064 PMCID: PMC7140417 DOI: 10.3390/cells9030568] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2020] [Revised: 02/22/2020] [Accepted: 02/26/2020] [Indexed: 02/07/2023] Open
Abstract
Hepatocellular carcinoma (HCC) is the leading cause of cancer-related deaths worldwide and the majority of HCC patients occur with a background of hepatic fibrosis and cirrhosis. We have previously reported the hepatoprotective effects of steamed and freeze-dried mature silkworm larval powder (SMSP) in a chronic ethanol-treated rat model. Here, we assessed the anti-fibrotic and anti-carcinogenic effects of SMSP on diethylnitrosamine (DEN)-treated rats. Wistar rats were intraperitoneally injected with DEN once a week for 12 or 16 weeks with or without SMSP administration (0.1 and 1 g/kg). SMSP administration significantly attenuated tumor foci formation and proliferation in the livers of the rats treated with DEN for 16 weeks. SMSP administration also inhibited hepatic fibrosis by decreasing the levels of collagen fiber and the expression of pro-collagen I and alpha-smooth muscle actin (α-SMA). Moreover, SMSP supplementation improved the major parameters of fibrosis such as transforming growth factor-β (TGF-β), connective tissue growth factor (CTGF), tumor necrosis factor-alpha (TNF-α), plasminogen activator inhibitor-1 (PAI-1), and collagen type I (Col1A1) in the livers from the rats treated with DEN for 16 weeks. As s possible mechanisms, we investigated the effects of SMSP on the TGF-β and signal transducer and activator of transcription 3 (STAT3)-mediated signaling cascades, which are known to promote hepatic fibrosis. We found that SMSP treatment inhibited the activation of TGF-β and the phosphorylation of STAT3 pathway in DEN-treated rats. Moreover, SMSP administration suppressed the expressions of the target genes of TGF-β and STAT3 induced by DEN treatment. Our findings provide experimental evidences that SMSP administration has inhibitory effects of hepatic fibrosis and HCC induced by DEN in vivo and could be a promising strategy for the prevention or treatment of hepatic fibrosis and hepatocellular carcinogenesis.
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Affiliation(s)
- Da-Young Lee
- College of Pharmacy and Institute of Pharmaceutical Sciences, CHA University, Seongnam 13488, Korea; (D.-Y.L.); (S.-M.Y.); (M.-Y.S.)
| | - Sun-Mi Yun
- College of Pharmacy and Institute of Pharmaceutical Sciences, CHA University, Seongnam 13488, Korea; (D.-Y.L.); (S.-M.Y.); (M.-Y.S.)
| | - Moon-Young Song
- College of Pharmacy and Institute of Pharmaceutical Sciences, CHA University, Seongnam 13488, Korea; (D.-Y.L.); (S.-M.Y.); (M.-Y.S.)
| | - Sang-Deok Ji
- Department of Agricultural Biology, National Institute of Agricultural Science, Rural Development Administration, Wanju 55365, Korea; (S.-D.J.); (J.-G.S.)
| | - Jong-Gon Son
- Department of Agricultural Biology, National Institute of Agricultural Science, Rural Development Administration, Wanju 55365, Korea; (S.-D.J.); (J.-G.S.)
| | - Eun-Hee Kim
- College of Pharmacy and Institute of Pharmaceutical Sciences, CHA University, Seongnam 13488, Korea; (D.-Y.L.); (S.-M.Y.); (M.-Y.S.)
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28
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Ahn SY, Maeng YS, Kim YR, Choe YH, Hwang HS, Hyun YM. In vivo monitoring of dynamic interaction between neutrophil and human umbilical cord blood-derived mesenchymal stem cell in mouse liver during sepsis. Stem Cell Res Ther 2020; 11:44. [PMID: 32014040 PMCID: PMC6998265 DOI: 10.1186/s13287-020-1559-4] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2019] [Revised: 11/30/2019] [Accepted: 01/09/2020] [Indexed: 12/24/2022] Open
Abstract
Background Sepsis is a global inflammatory disease that causes death. It has been reported that mesenchymal stem cell (MSC) treatment can attenuate inflammatory and septic symptoms. In this study, we investigated how interactions between neutrophils and human umbilical cord blood (hUCB)-MSCs in the liver of septic mice are involved in mitigating sepsis that is mediated by MSCs. Accordingly, we aimed to determine whether hUCB-MSC application could be an appropriate treatment for sepsis. Methods To induce septic condition, lipopolysaccharide (LPS) was intraperitoneally (i.p.) injected into mice 24 h after the intravenous (i.v.) injection of saline or hUCB-MSCs. To determine the effect of hUCB-MSCs on the immune response during sepsis, histologic analysis, immunoassays, and two-photon intravital imaging were performed 6 h post-LPS injection. For the survival study, mice were monitored for 6 days after LPS injection. Results The injection (i.v.) of hUCB-MSCs alleviated the severity of LPS-induced sepsis by increasing IL-10 levels (p < 0.001) and decreasing mortality (p < 0.05) in septic mice. In addition, this significantly reduced the recruitment of neutrophils (p < 0.001) to the liver. In hUCB-MSC-treated condition, we also observed several distinct patterns of dynamic interactions between neutrophils and hUCB-MSCs in the inflamed mouse liver, as well as vigorous interactions between hepatic stellate cells (HSCs or ito cells) and hUCB-MSCs. Interestingly, hUCB-MSCs that originated from humans were not recognized as foreign in the mouse body and consequently did not cause graft rejection. Conclusions These distinct interaction patterns between innate immune cells and hUCB-MSCs demonstrated that hUCB-MSCs have beneficial effects against LPS-induced sepsis through associations with neutrophils. In addition, the immunomodulatory properties of hUCB-MSCs might enable immune evasion in the host. Taken together, our results suggest the prospects of hUCB-MSCs as a therapeutic tool to inhibit inflammation and alleviate pathological immune responses such as sepsis.
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Affiliation(s)
- Sung Yong Ahn
- Department of Anatomy, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Yong-Sun Maeng
- Department of Obstetrics and Gynecology, Yonsei University College of Medicine, Seoul, Republic of Korea.,Institute of Women's Life Medical Science, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Yu Rim Kim
- Department of Anatomy, Yonsei University College of Medicine, Seoul, Republic of Korea.,BK21 PLUS Project for Medical Science, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Young Ho Choe
- Department of Anatomy, Yonsei University College of Medicine, Seoul, Republic of Korea.,BK21 PLUS Project for Medical Science, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Han Sung Hwang
- Department of Obstetrics and Gynecology, Research Institute of Medical Science, Konkuk University School of Medicine, Seoul, Republic of Korea.
| | - Young-Min Hyun
- Department of Anatomy, Yonsei University College of Medicine, Seoul, Republic of Korea. .,BK21 PLUS Project for Medical Science, Yonsei University College of Medicine, Seoul, Republic of Korea.
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29
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Han XQ, Xu SQ, Lin JG. Curcumin Recovers Intracellular Lipid Droplet Formation Through Increasing Perilipin 5 Gene Expression in Activated Hepatic Stellate Cells In Vitro. Curr Med Sci 2019; 39:766-777. [DOI: 10.1007/s11596-019-2104-5] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2019] [Revised: 09/02/2019] [Indexed: 02/06/2023]
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30
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Kitade M, Kaji K, Nishimura N, Seki K, Nakanishi K, Tsuji Y, Sato S, Saikawa S, Takaya H, Kawaratani H, Namisaki T, Moriya K, Mitoro A, Yoshiji H. Blocking development of liver fibrosis augments hepatic progenitor cell-derived liver regeneration in a mouse chronic liver injury model. Hepatol Res 2019; 49:1034-1045. [PMID: 30989766 DOI: 10.1111/hepr.13351] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/12/2018] [Revised: 04/08/2019] [Accepted: 04/10/2019] [Indexed: 02/08/2023]
Abstract
AIM The roles of hepatic progenitor cells (HPCs) in regeneration of a diseased liver are unclear. Hepatic stellate cells (HSCs) contribute to liver fibrosis but are also a component of the HPC niche. Hepatic progenitor cells expand along with HSC activation and liver fibrosis. However, little is known about the interplay of liver fibrosis and HPC-mediated liver regeneration. This study aimed to investigate HSCs and HPCs in liver regeneration. METHODS Liver injury in mice was induced with 3,5-diethoxycarbonyl-1,4-dihydrocollidine, and HPC expansion and fibrosis were assessed. An angiotensin II type 1 receptor blocker (ARB) was administered to assess its effect on fibrosis and regeneration. RESULTS Treatment with ARB attenuated fibrosis and expansion of α-smooth muscle actin-positive activated HSCs as indicated by increased liver weight and Ki-67-positive hepatocytes. Immunohistochemical staining suggested that HPC differentiation was shifted toward hepatocytes (HCs) when ARB treatment decreased HPC encapsulation by HSCs and extracellular matrix. Conditioned medium produced by culturing the human HSC LX-2 line strongly augmented differentiation to biliary epithelial cells (BECs) but inhibited that to HCs. Activated HSCs expressed Jagged1, a NOTCH ligand, which plays a central role in differentiation of HPCs toward BECs. CONCLUSIONS Hepatic stellate cells, the HPC niche cells, control differentiation of HPCs, directing them toward BECs rather than HCs in a diseased liver model. Antifibrosis treatment with an ARB preferentially redirects HPC differentiation toward HCs by blocking the NOTCH pathway in the HPC niche, resulting in more efficient HPC-mediated liver regeneration.
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Affiliation(s)
- Mitsuteru Kitade
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Kosuke Kaji
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Norihisa Nishimura
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Kenichiro Seki
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Keisuke Nakanishi
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Yuki Tsuji
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Shinya Sato
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Soichiro Saikawa
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Hiroaki Takaya
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Hideto Kawaratani
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Tadashi Namisaki
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Kei Moriya
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Akira Mitoro
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
| | - Hitoshi Yoshiji
- Third Department of Internal Medicine, Nara Medical University, Nara, Japan
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31
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Stefanovic B, Manojlovic Z, Vied C, Badger CD, Stefanovic L. Discovery and evaluation of inhibitor of LARP6 as specific antifibrotic compound. Sci Rep 2019; 9:326. [PMID: 30674965 PMCID: PMC6344531 DOI: 10.1038/s41598-018-36841-y] [Citation(s) in RCA: 35] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2018] [Accepted: 11/27/2018] [Indexed: 01/17/2023] Open
Abstract
Fibrosis is characterized by excessive production of type I collagen. Biosynthesis of type I collagen in fibrosis is augmented by binding of protein LARP6 to the 5' stem-loop structure (5'SL), which is found exclusively in type I collagen mRNAs. A high throughput screen was performed to discover inhibitors of LARP6 binding to 5'SL, as potential antifibrotic drugs. The screen yielded one compound (C9) which was able to dissociate LARP6 from 5' SL RNA in vitro and to inactivate the binding of endogenous LARP6 in cells. Treatment of hepatic stellate cells (liver cells responsible for fibrosis) with nM concentrations of C9 reduced secretion of type I collagen. In precision cut liver slices, as an ex vivo model of hepatic fibrosis, C9 attenuated the profibrotic response at 1 μM. In prophylactic and therapeutic animal models of hepatic fibrosis C9 prevented development of fibrosis or hindered the progression of ongoing fibrosis when administered at 1 mg/kg. Toxicogenetics analysis revealed that only 42 liver genes changed expression after administration of C9 for 4 weeks, suggesting minimal off target effects. Based on these results, C9 represents the first LARP6 inhibitor with significant antifibrotic activity.
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Affiliation(s)
- Branko Stefanovic
- Department of Biomedical Sciences, College of Medicine, Florida State University, 1115 West Call Street, Tallahassee, FL, 32306, USA.
| | - Zarko Manojlovic
- Keck School of Medicine of University of Southern California, 1450 Biggy Street, NRT 4510, Los Angeles, CA, 90033, USA
| | - Cynthia Vied
- Translational Science Laboratory, College of Medicine, Florida State University, 1115 West Call Street, Tallahassee, FL, 32306, USA
| | - Crystal-Dawn Badger
- Translational Science Laboratory, College of Medicine, Florida State University, 1115 West Call Street, Tallahassee, FL, 32306, USA
- Proteomics and Metabolomics Facility, Colorado State University, 401 West Pitkin Street, Fort Collins, CO, 80521, USA
| | - Lela Stefanovic
- Department of Biomedical Sciences, College of Medicine, Florida State University, 1115 West Call Street, Tallahassee, FL, 32306, USA
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Lim BJ, Lee WK, Lee HW, Lee KS, Kim JK, Chang HY, Lee JI. Selective deletion of hepatocyte platelet-derived growth factor receptor α and development of liver fibrosis in mice. Cell Commun Signal 2018; 16:93. [PMID: 30509307 PMCID: PMC6276164 DOI: 10.1186/s12964-018-0306-2] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/03/2018] [Accepted: 11/21/2018] [Indexed: 01/10/2023] Open
Abstract
BACKGROUND Platelet-derived growth factor receptor α (PDGFRα) expression is increased in activated hepatic stellate cells (HSCs) in cirrhotic liver, while normal hepatocytes express PDGFRα at a negligible level. However, cancerous hepatocytes may show upregulation of PDGFRα, and hepatocellular carcinoma is preceded by chronic liver injury. The role of PDGFRα in non-cancerous hepatocytes and liver fibrosis is unclear. We hypothesized that upon liver injury, PDGFRα in insulted hepatocytes contributes to liver fibrosis by facilitating intercellular crosstalk between hepatocytes and HSCs. METHODS Hepatocytes were isolated from normal and thioacetamide (TAA)-induced cirrhotic livers for assessment of PDGFRα expression. Conditional knock-out (KO) C57BL/6 mice, in which PDGFRα was selectively deleted in hepatocytes, were generated. Liver fibrosis was induced by injecting TAA for 8 weeks. Hep3B cells were transfected with a small interfering RNA (siRNA) (PDGFRα or control) and co-cultured with LX2 cells. RESULTS PDGFRα expression was increased in hepatocytes from fibrotic livers compared to normal livers. Conditional PDGFRα KO mice had attenuated TAA-induced liver fibrosis with decreased HSC activation and proliferation. Immunoblot analyses revealed decreased expression of phospho-p44/42 MAPK in TAA-treated KO mice; these mice also showed almost complete suppression of the upregulation of mouse double minute 2. Although KO mice exhibited increased expression of transforming growth factor (TGF)-β and Smad2/3, this was compensated for by increased expression of inhibitory Smad7. LX2 cells co-cultured with PDGFRα siRNA-infected Hep3B cells showed decreased PDGFRα, α smooth muscle actin, collagen α1(I), TGFβ, and Smad2/3 expression. LX2/PDGFRα-deleted hepatocyte co-culture medium showed decreased PDGF-BB and PDGF-CC levels. CONCLUSIONS Deletion of PDGFRα in hepatocytes attenuated the upregulation of PDGFRα in HSCs after TAA treatment, resulting in decreased liver fibrosis and HSC activation. This suggests that in the event of chronic liver injury, PDGFRα in hepatocytes plays an important role in liver fibrosis by affecting PDGFRα expression in HSCs.
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Affiliation(s)
- Beom Jin Lim
- Department of Pathology, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Woon-Kyu Lee
- Laboratory of Developmental Genetics, Department of Biomedical Sciences, Inha University College of Medicine, Incheon, Republic of Korea
| | - Hyun Woong Lee
- Department of Internal Medicine, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Kwan Sik Lee
- Department of Internal Medicine, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Ja Kyung Kim
- Department of Internal Medicine, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Hye Young Chang
- Medical Research Center, Gangnam Severance Hospital, Seoul, South Korea
| | - Jung Il Lee
- Department of Internal Medicine, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, Republic of Korea
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Jophlin LL, Koutalos Y, Chen C, Shah V, Rockey DC. Hepatic stellate cells retain retinoid-laden lipid droplets after cellular transdifferentiation into activated myofibroblasts. Am J Physiol Gastrointest Liver Physiol 2018; 315:G713-G721. [PMID: 30024770 PMCID: PMC6293250 DOI: 10.1152/ajpgi.00251.2017] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
Loss of retinyl ester (RE)-rich lipid droplets (LDs) from hepatic stellate cells (HSCs) is cited as a key event in their cellular transdifferentiation to activated, pro-fibrotic myofibroblasts; however, it remains unclear if changes in LD morphology or RE content are causal for transdifferentiation. To better understand LD dynamics in vitro within a common model of HSC activation, we used novel approaches preserving LD morphology and allowing for quantitation of RE. The size and quantity of LDs within in vitro and in vivo bile duct ligation (BDL)-activated HSCs were quantitated using adipocyte differentiation-related protein (ADRP) labeling and oil red o (ORO) staining (gold standard), and RE content was determined using fluorescence microscopy. We found during HSC activation in vitro that LD number differed significantly when measured by ADRP and ORO, respectively ( day 1: 56 vs. 5, P = 0.03; day 4: 101 vs. 39, P = 0.03; day 14: 241 vs. 12, P = 0.02). Ex vivo HSCs activated in vivo contained the same number of LDs as day 4 in vitro activated HSCs (118 vs. 101, P = 0.54). Decline in LD RE occurred beyond day 4 in vitro and day 1 ex vivo , after HSC transdifferentiation was underway. Lastly, in situ HSCs examined using electron microscopy show LDs tend to be smaller but are ultimately retained in BDL injured livers. Therefore, we conclude that during HSC transdifferentiation, LDs are not lost but are retained, decreasing in size. Additionally, RE content declines after transdifferentiation is underway. These data suggest that these LD changes are not causal for HSC transdifferentiation. NEW & NOTEWORTHY Loss of retinoid-laden lipid droplets from hepatic stellate cells has long been held as a hallmark of their transdifferentiation into activated myofibroblasts, the dominant cells that drive hepatic fibrosis. This study demonstrates that stellate cells activated in culture and after liver injury in vivo retain their lipid droplets and that these droplets become smaller and more numerous, with decreases in droplet retinoid concentration occurring only after cellular transdifferentiation is underway.
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Affiliation(s)
- Loretta L. Jophlin
- 1Department of Medicine, Medical University of South Carolina, Charleston, South Carolina,3Department of Gastroenterology and Hepatology, Mayo Clinic, Rochester, Minnesota
| | - Yiannis Koutalos
- 2Department of Ophthalmology, Medical University of South Carolina, Charleston, South Carolina
| | - Chunhe Chen
- 2Department of Ophthalmology, Medical University of South Carolina, Charleston, South Carolina
| | - Vijay Shah
- 3Department of Gastroenterology and Hepatology, Mayo Clinic, Rochester, Minnesota
| | - Don C. Rockey
- 1Department of Medicine, Medical University of South Carolina, Charleston, South Carolina
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Zhang Y, Liu J, Ma Y, Wang J, Zhu J, Liu J, Zhang J. Integrated profiling of long non-coding RNAs and mRNAs identifies novel regulators associated with liver fibrosis. Pathol Res Pract 2018; 214:1794-1803. [PMID: 30193772 DOI: 10.1016/j.prp.2018.08.021] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/28/2018] [Revised: 08/12/2018] [Accepted: 08/23/2018] [Indexed: 12/21/2022]
Abstract
Liver fibrosis is the underlying cause of cirrhosis and liver failure in any type of chronic liver injury. However, the function of lncRNAs in liver fibrosis is largely unknown. In this study, we performed transcriptome sequencing of CCl4 induced mouse fibrotic liver tissues and normal liver tissues, and found 118 lncRNAs and 1921 mRNAs were significantly up-regulated, whereas 59 lncRNAs and 1504 mRNAs were down-regulated in fibrotic livers. Gene ontology analysis revealed that the differentially expressed lncRNAs are implicated in cell junction, cell differentiation and cell proliferation, pathways closely associated with EMT. The co-expression network of highly conserved lncRNAs and highly co-expressed mRNAs were constructed, whose differential expression were further confirmed by quantitative real-time PCR. Two highly conserved lncRNAs, Gm11149 and Gm20471, were identified to be significantly up-regulated in liver fibrosis. Their target genes, Ncam1 and Prrx1 respectively, are important modulators of EMT. Thus, the relative dynamic levels of Gm11149 and Ncam1, Gm20471 and Prrx1 were further monitored during the progress of liver fibrosis and their co-expression pattern was proved. Collectively, our results uncovered a crucial role of lncRNAs in the regulation of liver fibrosis and the lncRNA -mRNA network might provide new therapeutic strategies.
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Affiliation(s)
- Yitong Zhang
- Department of Digestive Diseases, Huashan Hospital, Fudan University, Shanghai, China
| | - Jing Liu
- Ministry of Education Key Laboratory of Contemporary Anthropology and State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai, China
| | - Yanyun Ma
- Ministry of Education Key Laboratory of Contemporary Anthropology and State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai, China
| | - Jingjie Wang
- Department of Digestive Diseases, Huashan Hospital, Fudan University, Shanghai, China
| | - Jie Zhu
- Department of Digestive Diseases, Huashan Hospital, Fudan University, Shanghai, China
| | - Jie Liu
- Department of Digestive Diseases, Huashan Hospital, Fudan University, Shanghai, China
| | - Jun Zhang
- Department of Digestive Diseases, Huashan Hospital, Fudan University, Shanghai, China.
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Abstract
Stellate cells are resident lipid-storing cells of the pancreas and liver that transdifferentiate to a myofibroblastic state in the context of tissue injury. Beyond having roles in tissue homeostasis, stellate cells are increasingly implicated in pathological fibrogenic and inflammatory programs that contribute to tissue fibrosis and that constitute a growth-permissive tumor microenvironment. Although the capacity of stellate cells for extracellular matrix production and remodeling has long been appreciated, recent research efforts have demonstrated diverse roles for stellate cells in regulation of epithelial cell fate, immune modulation, and tissue health. Our present understanding of stellate cell biology in health and disease is discussed here, as are emerging means to target these multifaceted cells for therapeutic benefit.
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Affiliation(s)
- Mara H Sherman
- Department of Cell, Developmental & Cancer Biology, Oregon Health & Science University, Portland, Oregon 97201, USA;
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36
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Effect of Curcumin and Gliotoxin on Rat Liver Myofibroblast Culture. BIONANOSCIENCE 2018. [DOI: 10.1007/s12668-017-0494-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/18/2022]
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37
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Sui G, Cheng G, Yuan J, Hou X, Kong X, Niu H. Interleukin (IL)-13, Prostaglandin E2 (PGE2), and Prostacyclin 2 (PGI2) Activate Hepatic Stellate Cells via Protein kinase C (PKC) Pathway in Hepatic Fibrosis. Med Sci Monit 2018; 24:2134-2141. [PMID: 29633755 PMCID: PMC5909417 DOI: 10.12659/msm.906442] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
BACKGROUND Protein kinase C (PKC), interleukin (IL)-13, prostaglandin E2 (PGE2), and prostacyclin 2 (PGI2) can all play crucial roles in pulmonary fibrosis. However, their functions remain unclear in hepatic fibrosis mediated by hepatic stellate cells (HSCs), which has been demonstrated to be related to transforming growth factor-β (TGF-β) and platelet-derived growth factor (PDGF). MATERIAL AND METHODS All the experiments were based on LX-2 Hepatic stellate cells. The expression of TGF-β1 and PDGF were assessed by ELISA, RT-PCR, and Western blotting in human HSCs treated by IL-13, PGE2, and PGI2, respectively. At the same time, bridge assay and CCK8 assay were used to detect the cell proliferation and activity, PKC activity assay was used to test the activity of PKC, and PKC agonist and antagonist were used to verify the results obtained previously. RESULTS We found that IL-13, PGE2, and PGI2 significantly enhanced the expression of TGF-β1 and PDGF in human HSCs, which also clearly improved the proliferation and cell activity of HSCs. Moreover, PKC activity was significantly increased following IL-13, PGE2, and PGI2 treatments. We also found that the expression of TGF-β1 and PDGF, as well as the proliferation and cell activity of HSCs, were significantly enhanced by the PKC agonist phorbol 12-myristate 13-acetate (PMA), but suppressed by the PKC antagonist calphostin C. CONCLUSIONS We found that IL-13, PGE2, and PGI2 stimulated HSCs proliferation and secretion of TGF-β1 and PDGF by activating PKC, which predicted their potential roles in hepatic fibrosis.
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Affiliation(s)
- Guode Sui
- Department of Emergency General Surgery, Affiliated Hospital of Qingdao University, Qingdao, Shandong, China (mainland)
| | - Guang Cheng
- Department of Emergency General Surgery, Affiliated Hospital of Qingdao University, Qingdao, Shandong, China (mainland)
| | - Junjun Yuan
- Department of Emergency General Surgery, Affiliated Hospital of Qingdao University, Shandong, China (mainland)
| | - Xuena Hou
- Department of Emergency General Surgery, Affiliated Hospital of Qingdao University, Shandong, China (mainland)
| | - Xiaochen Kong
- Department of Emergency General Surgery, Affiliated Hospital of Qingdao University, Shandong, China (mainland)
| | - Haitao Niu
- Department of Urology, Affiliated Hospital of Qingdao University, Qingdao, Shandong, China (mainland)
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38
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Ding D, Chen LL, Zhai YZ, Hou CJ, Tao LL, Lu SH, Wu J, Liu XP. Trichostatin A inhibits the activation of Hepatic stellate cells by Increasing C/EBP-α Acetylation in vivo and in vitro. Sci Rep 2018. [PMID: 29535398 PMCID: PMC5849734 DOI: 10.1038/s41598-018-22662-6] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
Reversal of activated hepatic stellate cells (HSCs) to a quiescent state and apoptosis of activated HSCs are key elements in the reversion of hepatic fibrosis. CCAAT/enhancer binding protein α (C/EBP-α) has been shown to inhibit HSC activation and promote its apoptosis. This study aims to investigate how C/EBP-α acetylation affects the fate of activated HSCs. Effects of a histone deacetylation inhibitor trichostatin A (TSA) on HSC activation were evaluated in a mouse model of liver fibrosis caused by carbon tetrachloride (CCl4) intoxication. TSA was found to ameliorate CCl4-induced hepatic fibrosis and improve liver function through increasing the protein level and enhancing C/EBP-α acetylation in the mouse liver. C/EBP-α acetylation was determined in HSC lines in the presence or absence of TSA, and the lysine residue K276 was identified as a main acetylation site in C/EBP-α protein. C/EBP-α acetylation increased its stability and protein level, and inhibited HSC activation. The present study demonstrated that C/EBP-α acetylation increases the protein level by inhibiting its ubiquitination-mediated degradation, and may be involved in the fate of activated HSCs. Use of TSA may confer an option in minimizing hepatic fibrosis by suppressing HSC activation, a key process in the initiation and progression of hepatic fibrosis.
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Affiliation(s)
- Di Ding
- Department of Pathology, School of Basic Medical Sciences, Fudan University, Shanghai, 200032, China
| | - Lin-Lin Chen
- Department of Pathology, The Fifth People's Hospital, Fudan University, Shanghai, 200040, China
| | - Ying-Zhen Zhai
- Department of Pathology, School of Basic Medical Sciences, Fudan University, Shanghai, 200032, China
| | - Chen-Jian Hou
- Department of Pathology, School of Basic Medical Sciences, Fudan University, Shanghai, 200032, China
| | - Li-Li Tao
- Department of Pathology, Peking University, Shenzhen Hospital, Shenzhen, 518036, China
| | - Shu-Han Lu
- Department of Nutrition, University of California at Davis, Davis, California, USA
| | - Jian Wu
- Department of Medical Microbiology, Key Laboratory of Molecular Virology, School of Basic Medical Sciences, Fudan University, Shanghai, 200032, China. .,Shanghai Institute of Liver Disease, Fudan University, Shanghai, 200032, China.
| | - Xiu-Ping Liu
- Department of Pathology, School of Basic Medical Sciences, Fudan University, Shanghai, 200032, China. .,Department of Pathology, The Fifth People's Hospital, Fudan University, Shanghai, 200040, China.
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Zhang Y, Luo G, Zhang Y, Zhang M, Zhou J, Gao W, Xuan X, Yang X, Yang D, Tian Z, Ni B, Tang J. Critical effects of long non-coding RNA on fibrosis diseases. Exp Mol Med 2018; 50:e428. [PMID: 29350677 PMCID: PMC5799794 DOI: 10.1038/emm.2017.223] [Citation(s) in RCA: 37] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2017] [Revised: 07/03/2017] [Accepted: 07/05/2017] [Indexed: 02/07/2023] Open
Abstract
The expression or dysfunction of long non-coding RNAs (lncRNAs) is closely related to various hereditary diseases, autoimmune diseases, metabolic diseases and tumors. LncRNAs were also recently recognized as functional regulators of fibrosis, which is a secondary process in many of these diseases and a primary pathology in fibrosis diseases. We review the latest findings on lncRNAs in fibrosis diseases of the liver, myocardium, kidney, lung and peritoneum. We also discuss the potential of disease-related lncRNAs as therapeutic targets for the clinical treatment of human fibrosis diseases.
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Affiliation(s)
- Yue Zhang
- Department of Dermatology, 105th Hospital of PLA, Hefei, China.,Department of Pathophysiology and High Altitude Pathology, Third Military Medical University, Chongqing, China.,Graduate School, Bengbu Medical College, Bengbu, China
| | - Gang Luo
- Department of Pathophysiology and High Altitude Pathology, Third Military Medical University, Chongqing, China
| | - Yi Zhang
- Department of Clinical Laboratory, 150th Hospital of PLA, Luoyang, China
| | - Mengjie Zhang
- Department of Pathophysiology and High Altitude Pathology, Third Military Medical University, Chongqing, China
| | - Jian Zhou
- Department of Pathophysiology and High Altitude Pathology, Third Military Medical University, Chongqing, China
| | - Weiwu Gao
- Department of Pathophysiology and High Altitude Pathology, Third Military Medical University, Chongqing, China
| | - Xiuyun Xuan
- Department of Microbiology and Immunology, Shanxi Medical University, Taiyuan, China
| | - Xia Yang
- Institute of Immunology, PLA, Third Military Medical University, Chongqing, China
| | - Di Yang
- Institute of Immunology, PLA, Third Military Medical University, Chongqing, China
| | - Zhiqiang Tian
- Institute of Immunology, PLA, Third Military Medical University, Chongqing, China
| | - Bing Ni
- Department of Pathophysiology and High Altitude Pathology, Third Military Medical University, Chongqing, China
| | - Jun Tang
- Department of Dermatology, 105th Hospital of PLA, Hefei, China.,Graduate School, Bengbu Medical College, Bengbu, China
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40
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Crawford JM, Bioulac-Sage P, Hytiroglou P. Structure, Function, and Responses to Injury. MACSWEEN'S PATHOLOGY OF THE LIVER 2018:1-87. [DOI: 10.1016/b978-0-7020-6697-9.00001-7] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/03/2025]
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41
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Shang L, Hosseini M, Liu X, Kisseleva T, Brenner DA. Human hepatic stellate cell isolation and characterization. J Gastroenterol 2018; 53:6-17. [PMID: 29094206 DOI: 10.1007/s00535-017-1404-4] [Citation(s) in RCA: 91] [Impact Index Per Article: 13.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/15/2017] [Accepted: 09/22/2017] [Indexed: 02/04/2023]
Abstract
The hepatic stellate cells (HSCs) localize at the space of Disse in the liver and have multiple functions. They are identified as the major contributor to hepatic fibrosis. Significant understanding of HSCs has been achieved using rodent models and isolated murine HSCs; as well as investigating human liver tissues and human HSCs. There is growing interest and need of translating rodent study findings to human HSCs and human liver diseases. However, species-related differences impose challenges on the translational research. In this review, we focus on the current information on human HSCs isolation methods, human HSCs markers, and established human HSC cell lines.
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Affiliation(s)
- Linshan Shang
- Department of Medicine, University of California, San Diego, La Jolla, USA
| | - Mojgan Hosseini
- Department of Pathology, University of California, San Diego, La Jolla, USA
| | - Xiao Liu
- Department of Surgery, University of California, San Diego, La Jolla, USA
| | - Tatiana Kisseleva
- Department of Surgery, University of California, San Diego, La Jolla, USA
| | - David Allen Brenner
- Department of Medicine, University of California, San Diego, La Jolla, USA.
- School of Medicine, UC San Diego, 9500 Gilman Drive, La Jolla, CA, 92093-0602, USA.
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42
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Hajavi J, Momtazi AA, Johnston TP, Banach M, Majeed M, Sahebkar A. Curcumin: A Naturally Occurring Modulator of Adipokines in Diabetes. J Cell Biochem 2017; 118:4170-4182. [DOI: 10.1002/jcb.26121] [Citation(s) in RCA: 30] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2017] [Accepted: 05/08/2017] [Indexed: 01/30/2023]
Affiliation(s)
- Jafar Hajavi
- Immunology Research CenterMashhad University of Medical SciencesMashhadIran
| | - Amir Abbas Momtazi
- Student Research Committee, Nanotechnology Research CenterDepartment of Medical Biotechnology, School of Medicine, Mashhad University of Medical SciencesMashhadIran
| | - Thomas P. Johnston
- Division of Pharmaceutical Sciences, School of PharmacyUniversity of Missouri‐Kansas CityKansas CityMissouri
| | - Maciej Banach
- Department of Hypertension, WAM University Hospital in LodzMedical University of LodzZeromskiego 113LodzPoland
| | | | - Amirhossein Sahebkar
- Biotechnology Research CenterMashhad University of Medical SciencesMashhad9177948564Iran
- Neurogenic Inflammation Research CenterMashhad University of Medical SciencesMashhadIran
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43
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Ferdek PE, Jakubowska MA. Biology of pancreatic stellate cells-more than just pancreatic cancer. Pflugers Arch 2017; 469:1039-1050. [PMID: 28382480 PMCID: PMC5554282 DOI: 10.1007/s00424-017-1968-0] [Citation(s) in RCA: 81] [Impact Index Per Article: 10.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2017] [Revised: 03/13/2017] [Accepted: 03/16/2017] [Indexed: 01/18/2023]
Abstract
Pancreatic stellate cells, normally quiescent, are capable of remarkable transition into their activated myofibroblast-like phenotype. It is now commonly accepted that these cells play a pivotal role in the desmoplastic reaction present in severe pancreatic disorders. In recent years, enormous scientific effort has been devoted to understanding their roles in pancreatic cancer, which continues to remain one of the most deadly diseases. Therefore, it is not surprising that considerably less attention has been given to studying physiological functions of pancreatic stellate cells. Here, we review recent advances not only in the field of pancreatic stellate cell pathophysiology but also emphasise their roles in physiological processes.
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Affiliation(s)
- Pawel E Ferdek
- Medical Research Council Group, Cardiff School of Biosciences, Cardiff University, Cardiff, Wales, CF10 3AX, UK.
| | - Monika A Jakubowska
- Medical Research Council Group, Cardiff School of Biosciences, Cardiff University, Cardiff, Wales, CF10 3AX, UK
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44
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Chen JY, Newcomb B, Zhou C, Pondick JV, Ghoshal S, York SR, Motola DL, Coant N, Yi JK, Mao C, Tanabe KK, Bronova I, Berdyshev EV, Fuchs BC, Hannun Y, Chung RT, Mullen AC. Tricyclic Antidepressants Promote Ceramide Accumulation to Regulate Collagen Production in Human Hepatic Stellate Cells. Sci Rep 2017; 7:44867. [PMID: 28322247 PMCID: PMC5359599 DOI: 10.1038/srep44867] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2016] [Accepted: 02/15/2017] [Indexed: 12/21/2022] Open
Abstract
Activation of hepatic stellate cells (HSCs) in response to injury is a key step in hepatic fibrosis, and is characterized by trans-differentiation of quiescent HSCs to HSC myofibroblasts, which secrete extracellular matrix proteins responsible for the fibrotic scar. There are currently no therapies to directly inhibit hepatic fibrosis. We developed a small molecule screen to identify compounds that inactivate human HSC myofibroblasts through the quantification of lipid droplets. We screened 1600 compounds and identified 21 small molecules that induce HSC inactivation. Four hits were tricyclic antidepressants (TCAs), and they repressed expression of pro-fibrotic factors Alpha-Actin-2 (ACTA2) and Alpha-1 Type I Collagen (COL1A1) in HSCs. RNA sequencing implicated the sphingolipid pathway as a target of the TCAs. Indeed, TCA treatment of HSCs promoted accumulation of ceramide through inhibition of acid ceramidase (aCDase). Depletion of aCDase also promoted accumulation of ceramide and was associated with reduced COL1A1 expression. Treatment with B13, an inhibitor of aCDase, reproduced the antifibrotic phenotype as did the addition of exogenous ceramide. Our results show that detection of lipid droplets provides a robust readout to screen for regulators of hepatic fibrosis and have identified a novel antifibrotic role for ceramide.
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Affiliation(s)
- Jennifer Y Chen
- Gastrointestinal Unit, Massachusetts General Hospital, Harvard Medical School, Boston, MA USA
| | - Benjamin Newcomb
- Health Science Center, Stony Brook University, Stony Brook, NY USA
| | - Chan Zhou
- Gastrointestinal Unit, Massachusetts General Hospital, Harvard Medical School, Boston, MA USA
| | - Joshua V Pondick
- Gastrointestinal Unit, Massachusetts General Hospital, Harvard Medical School, Boston, MA USA
| | - Sarani Ghoshal
- Division of Surgical Oncology, Massachusetts General Hospital Cancer Center, Harvard Medical School, Boston, MA USA
| | - Samuel R York
- Gastrointestinal Unit, Massachusetts General Hospital, Harvard Medical School, Boston, MA USA
| | - Daniel L Motola
- Gastrointestinal Unit, Massachusetts General Hospital, Harvard Medical School, Boston, MA USA
| | - Nicolas Coant
- Health Science Center, Stony Brook University, Stony Brook, NY USA
| | - Jae Kyo Yi
- Health Science Center, Stony Brook University, Stony Brook, NY USA
| | - Cungui Mao
- Health Science Center, Stony Brook University, Stony Brook, NY USA
| | - Kenneth K Tanabe
- Division of Surgical Oncology, Massachusetts General Hospital Cancer Center, Harvard Medical School, Boston, MA USA
| | | | | | - Bryan C Fuchs
- Division of Surgical Oncology, Massachusetts General Hospital Cancer Center, Harvard Medical School, Boston, MA USA
| | - Yusuf Hannun
- Health Science Center, Stony Brook University, Stony Brook, NY USA
| | - Raymond T Chung
- Gastrointestinal Unit, Massachusetts General Hospital, Harvard Medical School, Boston, MA USA
| | - Alan C Mullen
- Gastrointestinal Unit, Massachusetts General Hospital, Harvard Medical School, Boston, MA USA.,Harvard Stem Cell Institute, Cambridge, MA 02138 USA
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45
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The stellate cell system (vitamin A-storing cell system). Anat Sci Int 2017; 92:387-455. [PMID: 28299597 DOI: 10.1007/s12565-017-0395-9] [Citation(s) in RCA: 73] [Impact Index Per Article: 9.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2016] [Accepted: 02/15/2017] [Indexed: 01/18/2023]
Abstract
Past, present, and future research into hepatic stellate cells (HSCs, also called vitamin A-storing cells, lipocytes, interstitial cells, fat-storing cells, or Ito cells) are summarized and discussed in this review. Kupffer discovered black-stained cells in the liver using the gold chloride method and named them stellate cells (Sternzellen in German) in 1876. Wake rediscovered the cells in 1971 using the same gold chloride method and various modern histological techniques including electron microscopy. Between their discovery and rediscovery, HSCs disappeared from the research history. Their identification, the establishment of cell isolation and culture methods, and the development of cellular and molecular biological techniques promoted HSC research after their rediscovery. In mammals, HSCs exist in the space between liver parenchymal cells (PCs) or hepatocytes and liver sinusoidal endothelial cells (LSECs) of the hepatic lobule, and store 50-80% of all vitamin A in the body as retinyl ester in lipid droplets in the cytoplasm. SCs also exist in extrahepatic organs such as pancreas, lung, and kidney. Hepatic (HSCs) and extrahepatic stellate cells (EHSCs) form the stellate cell (SC) system or SC family; the main storage site of vitamin A in the body is HSCs in the liver. In pathological conditions such as liver fibrosis, HSCs lose vitamin A, and synthesize a large amount of extracellular matrix (ECM) components including collagen, proteoglycan, glycosaminoglycan, and adhesive glycoproteins. The morphology of these cells also changes from the star-shaped HSCs to that of fibroblasts or myofibroblasts.
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Korenblat K. Management of ascites in cirrhosis and portal hypertension. BLUMGART'S SURGERY OF THE LIVER, BILIARY TRACT AND PANCREAS, 2-VOLUME SET 2017:1189-1195.e2. [DOI: 10.1016/b978-0-323-34062-5.00081-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/05/2025]
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Henrique Silva A, Lima Jr E, Vasquez Mansilla M, Zysler RD, Mojica Pisciotti ML, Locatelli C, Kumar Reddy Rajoli R, Owen A, Creczynski-Pasa TB, Siccardi M. A physiologically based pharmacokinetic model to predict the superparamagnetic iron oxide nanoparticles (SPIONs) accumulation in vivo. EUROPEAN JOURNAL OF NANOMEDICINE 2017. [DOI: 10.1515/ejnm-2017-0001] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
Abstract
AbstractSuperparamagnetic iron oxide nanoparticles (SPIONs) have been identified as a promising material for biomedical applications. These include as contrast agents for medical imaging, drug delivery and/or cancer cell treatment. The nanotoxicological profile of SPIONs has been investigated in different studies and the distribution of SPIONs in the human body has not been fully characterized. The aim of this study was to develop a physiologically-based pharmacokinetic (PBPK) model to predict the pharmacokinetics of SPIONs. The distribution and accumulation of SPIONs in organs were simulated taking into consideration their penetration through capillary walls and their active uptake by specialized macrophages in the liver, spleen and lungs. To estimate the kinetics of SPION uptake, a novel experimental approach using primary macrophages was developed. The murine PBPK model was validated against in vivo pharmacokinetic data, and accurately described accumulation in liver, spleen and lungs. After validation of the murine model, a similar PBPK approach was developed to simulate the distribution of SPIONs in humans. These data demonstrate the utility of PBPK modeling for estimating biodistribution of inorganic nanoparticles and represents an initial platform to provide computational prediction of nanoparticle pharmacokinetics.
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Kim RS, Hasegawa D, Goossens N, Tsuchida T, Athwal V, Sun X, Robinson CL, Bhattacharya D, Chou HI, Zhang DY, Fuchs BC, Lee Y, Hoshida Y, Friedman SL. The XBP1 Arm of the Unfolded Protein Response Induces Fibrogenic Activity in Hepatic Stellate Cells Through Autophagy. Sci Rep 2016; 6:39342. [PMID: 27996033 PMCID: PMC5172197 DOI: 10.1038/srep39342] [Citation(s) in RCA: 74] [Impact Index Per Article: 8.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2016] [Accepted: 11/18/2016] [Indexed: 02/07/2023] Open
Abstract
Autophagy and the unfolded protein response (UPR) both promote activation of hepatic stellate cells (HSC), however the link between the two stimuli remains unclear. Here we have explored the role of X-box binding protein 1 (XBP1), one of three UPR effector pathways and sought to establish the interdependence between autophagy and the UPR during HSC activation. XBP1 induction accompanied both culture-based HSC activation and ER stress induced by tunicamycin. Ectopic overexpression of XBP1 induced collagen 1-alpha expression in HSCs, which was inhibited by knockdown of ATG7, a critical autophagy mediator. Genome-wide transcriptomic profiling indicated an upregulation of collagen synthesis pathways, but not of the transforming growth factor (TGF)-b pathway, a canonical fibrogenic driver, suggesting that XBP1 activates a specific subset of fibrogenesis pathways independent of TGF-β1. XBP1 target gene signatures were significantly induced in rodent liver fibrosis models (n = 3-5) and in human samples of non-alcoholic fatty liver disease (NAFLD) (n = 72-135). Thus, XBP1-mediated UPR contributes to fibrogenic HSC activation and is functionally linked to cellular autophagy.
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Affiliation(s)
- Rosa S. Kim
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
| | - Daisuke Hasegawa
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
- Divisions of Gastroenterology and Hepatology, St. Marianna University School of Medicine, Kawasaki, Japan
| | - Nicolas Goossens
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
- Divisions of Gastroenterology and Hepatology, Geneva University Hospital, Geneva, Switzerland
| | - Takuma Tsuchida
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
- Research Division, Mitsubishi Tanabe Pharma Corporation, Saitama, Japan
| | - Varinder Athwal
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
- Institute of Human Development, University of Manchester, United Kingdom
| | - Xiaochen Sun
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
| | | | - Dipankar Bhattacharya
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
| | - Hsin-I Chou
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
| | - David Y. Zhang
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
| | - Bryan C. Fuchs
- Division of Surgical Oncology, Massachusetts General Hospital Cancer Center, Harvard Medical School, Boston, Massachusetts, US
| | - Youngmin Lee
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
| | - Yujin Hoshida
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
| | - Scott L. Friedman
- Division of Liver Diseases, Department of Medicine, Liver Cancer Program, Tisch Cancer Institute, Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, US
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Öner-İyidoğan Y, Seyithanoğlu M, Tanrıkulu-Küçük S, Koçak H, Beyhan-Özdaş Ş, Koçak-Toker N. The effect of dietary curcumin on hepatic chymase activity and serum fetuin-A levels in rats fed on a high-fat diet. J Food Biochem 2016. [DOI: 10.1111/jfbc.12347] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/11/2023]
Affiliation(s)
- Yıldız Öner-İyidoğan
- Department of Biochemistry, Istanbul Faculty of Medicine; Istanbul University, Çapa; Istanbul Turkey
| | - Muhammed Seyithanoğlu
- Department of Biochemistry, Istanbul Faculty of Medicine; Istanbul University, Çapa; Istanbul Turkey
| | - Sevda Tanrıkulu-Küçük
- Department of Biochemistry, Faculty of Medicine; Istanbul Bilim University; Esentepe Istanbul Turkey
| | - Hikmet Koçak
- Department of Biochemistry, Faculty of Medicine; Istanbul Bilim University; Esentepe Istanbul Turkey
| | - Şule Beyhan-Özdaş
- Department of Medical Biology and Genetic, Faculty of Medicine; Istanbul Bilim University; Esentepe Istanbul Turkey
| | - Necla Koçak-Toker
- Department of Biochemistry, Istanbul Faculty of Medicine; Istanbul University, Çapa; Istanbul Turkey
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Li S, Wang Q, Tao Y, Liu C. Swertiamarin Attenuates Experimental Rat Hepatic Fibrosis by Suppressing Angiotensin II-Angiotensin Type 1 Receptor-Extracellular Signal-Regulated Kinase Signaling. J Pharmacol Exp Ther 2016; 359:247-255. [PMID: 27543328 DOI: 10.1124/jpet.116.234179] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2016] [Accepted: 08/17/2016] [Indexed: 03/08/2025] Open
Abstract
The rennin-angiotensin system (RAS) is crucial in hepatic fibrosis development, and therapies targeting this system may be a promising treatment for hepatic fibrosis. In this study, we investigated the effects of swertiamarin (Swe), an ethanol extract of Gentiana manshurica Kitag, on hepatic fibrosis and its underlying mechanisms through regulating RAS. Primary rat hepatic stellate cells (HSCs) were isolated and treated with angiotensin II (Ang II) with or without Swe and losartan. The proliferation and activation of HSCs were measured. Rat hepatic fibrosis was induced by intraperitoneal dimethylnitrosamine (DMN) injection for 4 weeks. Rats were treated with Swe or losartan from the third week until the end of the experiment. Hydroxyproline content in liver tissue was assayed with Jamall's method, and liver collagen deposition was visualized using Sirius red staining. RAS components were analyzed by Western blot, immunofluorescent staining, and real-time reverse-transcription polymerase chain reaction. The results showed that Swe significantly inhibited Ang II-induced HSC proliferation and activation. Swe also significantly suppressed DMN-induced α-smooth muscle actin production in rat livers and improved liver function. Swe partially inhibited Ang II-induced angiotensin type 1 receptor (AT1R) up-regulation and suppressed Ang II-induced extracellular signal-regulated kinase (ERK) and c-jun phosphorylation in HSCs. In the DMN-treated rats, Swe treatment significantly inhibited the plasma Ang II levels. DMN-induced AT1R up-regulation, and phosphorylation of ERK and c-jun in rat liver were also inhibited by Swe. In conclusion, Swe may attenuate hepatic fibrosis through inhibiting HSC activation by regulating the RAS.
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Affiliation(s)
- Shu Li
- Institute of Liver Diseases, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine (S.L., Q.W., Y.T., C.L.); Baoshan Branch, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine (S.L.); Shanghai Clinical Key Laboratory of Traditional Chinese Medicine (C.L.); E-Institute of TCM Internal Medicine, Shanghai Municipal Education Commission (C.L.), Shanghai, People's Republic of China
| | - Qinglan Wang
- Institute of Liver Diseases, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine (S.L., Q.W., Y.T., C.L.); Baoshan Branch, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine (S.L.); Shanghai Clinical Key Laboratory of Traditional Chinese Medicine (C.L.); E-Institute of TCM Internal Medicine, Shanghai Municipal Education Commission (C.L.), Shanghai, People's Republic of China
| | - Yanyan Tao
- Institute of Liver Diseases, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine (S.L., Q.W., Y.T., C.L.); Baoshan Branch, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine (S.L.); Shanghai Clinical Key Laboratory of Traditional Chinese Medicine (C.L.); E-Institute of TCM Internal Medicine, Shanghai Municipal Education Commission (C.L.), Shanghai, People's Republic of China
| | - Chenghai Liu
- Institute of Liver Diseases, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine (S.L., Q.W., Y.T., C.L.); Baoshan Branch, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine (S.L.); Shanghai Clinical Key Laboratory of Traditional Chinese Medicine (C.L.); E-Institute of TCM Internal Medicine, Shanghai Municipal Education Commission (C.L.), Shanghai, People's Republic of China
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