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Chaudhary JK, Danga AK, Kumari A, Bhardwaj A, Rath PC. Role of stem cells in ageing and age-related diseases. Mech Ageing Dev 2025; 225:112069. [PMID: 40324541 DOI: 10.1016/j.mad.2025.112069] [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: 01/17/2025] [Revised: 04/30/2025] [Accepted: 05/01/2025] [Indexed: 05/07/2025]
Abstract
Stem cell functions and ageing are deeply interconnected, continually influencing each other in multiple ways. Stem cells play a vital role in organ maintenance, regeneration, and homeostasis, all of which decline over time due to gradual reduction in their self-renewal, differentiation, and growth factor secretion potential. The functional decline is attributed to damaging extrinsic environmental factors and progressively worsening intrinsic genetic and biochemical processes. These ageing-associated deteriorative changes have been extensively documented, paving the way for the discovery of novel biomarkers of ageing for detection, diagnosis, and treatment of age-related diseases. Age-dependent changes in adult stem cells include numerical decline, loss of heterogeneity, and reduced self-renewal and differentiation, leading to a drastic reduction in regenerative potential and thereby driving the ageing process. Conversely, ageing also adversely alters the stem cell niche, disrupting the molecular pathways underlying stem cell homing, self-renewal, differentiation, and growth factor secretion, all of which are critical for tissue repair and regeneration. A holistic understanding of these molecular mechanisms, through empirical research and clinical trials, is essential for designing targeted therapies to modulate ageing and improve health parameters in older individuals.
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Affiliation(s)
- Jitendra Kumar Chaudhary
- Molecular Biology Laboratory, School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India; Department of Zoology, Shivaji College, University of Delhi, New Delhi 110027, India.
| | - Ajay Kumar Danga
- Molecular Biology Laboratory, School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India; National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, India.
| | - Anita Kumari
- Molecular Biology Laboratory, School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
| | - Akshay Bhardwaj
- Global Research Alliances, Ashoka University, Rajiv Gandhi Education City, Sonepat, Haryana 131029, India.
| | - Pramod C Rath
- Molecular Biology Laboratory, School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
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2
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Liu S, Cheng C, Zhu L, Zhao T, Wang Z, Yi X, Yan F, Wang X, Li C, Cui T, Yang B. Liver organoids: updates on generation strategies and biomedical applications. Stem Cell Res Ther 2024; 15:244. [PMID: 39113154 PMCID: PMC11304926 DOI: 10.1186/s13287-024-03865-3] [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: 04/25/2024] [Accepted: 07/27/2024] [Indexed: 08/10/2024] Open
Abstract
The liver is the most important metabolic organ in the body. While mouse models and cell lines have further deepened our understanding of liver biology and related diseases, they are flawed in replicating key aspects of human liver tissue, particularly its complex structure and metabolic functions. The organoid model represents a major breakthrough in cell biology that revolutionized biomedical research. Organoids are in vitro three-dimensional (3D) physiological structures that recapitulate the morphological and functional characteristics of tissues in vivo, and have significant advantages over traditional cell culture methods. In this review, we discuss the generation strategies and current advances in the field focusing on their application in regenerative medicine, drug discovery and modeling diseases.
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Affiliation(s)
- Sen Liu
- Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang, 110016, China
- State Key Laboratory of Druggability Evaluation and Systematic Translational Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin, 300301, China
| | | | - Liuyang Zhu
- First Central Clinical College of Tianjin Medical University, Tianjin, 300192, China
| | - Tianyu Zhao
- State Key Laboratory of Druggability Evaluation and Systematic Translational Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin, 300301, China
| | - Ze Wang
- State Key Laboratory of Druggability Evaluation and Systematic Translational Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin, 300301, China
- Research Unit for Drug Metabolism, Chinese Academy of Medical Sciences, Beijing, 100730, China
| | - Xiulin Yi
- State Key Laboratory of Druggability Evaluation and Systematic Translational Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin, 300301, China
- Research Unit for Drug Metabolism, Chinese Academy of Medical Sciences, Beijing, 100730, China
| | - Fengying Yan
- State Key Laboratory of Druggability Evaluation and Systematic Translational Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin, 300301, China
- Research Unit for Drug Metabolism, Chinese Academy of Medical Sciences, Beijing, 100730, China
| | - Xiaoliang Wang
- State Key Laboratory of Druggability Evaluation and Systematic Translational Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin, 300301, China
| | - Chunli Li
- Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang, 110016, China.
| | - Tao Cui
- State Key Laboratory of Druggability Evaluation and Systematic Translational Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin, 300301, China.
- Research Unit for Drug Metabolism, Chinese Academy of Medical Sciences, Beijing, 100730, China.
| | - Baofeng Yang
- Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang, 110016, China.
- School of Pharmacy, Harbin Medical University, Harbin, 150081, China.
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3
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Hu Y, Bao X, Zhang Z, Chen L, Liang Y, Qu Y, Zhou Q, Zhou X, Fang J, Xiao Z, Fu Y, Yang H, Liu W, Lv Y, Cao H, Chen G, Ping J, Zhang H, Mu Y, Liu C, Lin CP, Wu J, Liu P, Chen J. Hepatic progenitor cell-originated ductular reaction facilitates liver fibrosis through activation of hedgehog signaling. Theranostics 2024; 14:2379-2395. [PMID: 38646644 PMCID: PMC11024850 DOI: 10.7150/thno.91572] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2023] [Accepted: 03/17/2024] [Indexed: 04/23/2024] Open
Abstract
Background: It is poorly understood what cellular types participate in ductular reaction (DR) and whether DR facilitates recovery from injury or accelerates hepatic fibrosis. The aim of this study is to gain insights into the role of hepatic progenitor cell (HPC)-originated DR during fibrotic progression. Methods: DR in liver specimens of PBC, chronic HBV infection (CHB) or NAFLD, and four rodent fibrotic models by different pathogenic processes was evaluated. Gli1 expression was inhibited in rodent models or cell culture and organoid models by AAV-shGli1 or treating with GANT61. Results: Severity of liver fibrosis was positively correlated with DR extent in patients with PBC, CHB or NAFLD. HPCs were activated, expanded, differentiated into reactive cholangiocytes and constituted "HPC-originated DR", accompanying with exacerbated fibrosis in rodent models of HPC activation & proliferation (CCl4/2-AAF-treated), Μdr2-/- spontaneous PSC, BDL-cholestatic fibrosis or WD-fed/CCl4-treated NASH-fibrosis. Gli1 expression was significantly increased in enriched pathways in vivo and in vitro. Enhanced Gli1 expression was identified in KRT19+-reactive cholangiocytes. Suppressing Gli1 expression by administration of AAV-shGli1 or GANT61 ameliorated HPC-originated DR and fibrotic extent. KRT19 expression was reduced after GANT61 treatment in sodium butyrate-stimulated WB-F344 cells or organoids or in cells transduced with Gli1 knockdown lentiviral vectors. In contrast, KRT19 expression was elevated after transducing Gli1 overexpression lentiviral vectors in these cells. Conclusions: During various modes of chronic injury, Gli1 acted as an important mediator of HPC activation, expansion, differentiation into reactive cholangiocytes that formed DR, and subsequently provoked hepatic fibrogenesis.
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Affiliation(s)
- Yonghong Hu
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
- Institute of Surgery of Integrated Traditional Chinese and Western Medicine, Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
| | - Xinyu Bao
- School of Life Science and Technology, ShanghaiTech University, Shanghai, 201210, China
- Shanghai Clinical Research and Trial Center, Shanghai 201210, China
| | - Zheng Zhang
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Long Chen
- Institute of Interdisciplinary Integrative Medicine Research, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
| | - Yue Liang
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Yan Qu
- Department of Hepatobiliary Surgery, Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
| | - Qun Zhou
- Institute of Interdisciplinary Integrative Medicine Research, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
| | - Xiaoxi Zhou
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Jing Fang
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Zhun Xiao
- Institute of Interdisciplinary Integrative Medicine Research, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
| | - Yadong Fu
- Institute of Interdisciplinary Integrative Medicine Research, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
| | - Hailin Yang
- Institute of Interdisciplinary Integrative Medicine Research, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
| | - Wei Liu
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Ying Lv
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Hongyan Cao
- Department of Gastroenterology, Shanghai University of Traditional Chinese Medicine Shanghai TCM - Integrated hospital, Shanghai 201203, China
| | - Gaofeng Chen
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Jian Ping
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Hua Zhang
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Yongping Mu
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Chenghai Liu
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
| | - Chao-Po Lin
- School of Life Science and Technology, ShanghaiTech University, Shanghai, 201210, China
- Shanghai Clinical Research and Trial Center, Shanghai 201210, China
| | - Jian Wu
- Department of Medical Microbiology & Parasitology, MOE/NHC/CAMS Key Laboratory of Medical Molecular Virology, School of Basic Medical Sciences, Fudan University Shanghai Medical College, Shanghai 200032, China
- Department of Gastroenterology & Hepatology, Zhongshan Hospital of Fudan University, Shanghai 200032, China
- Shanghai Institute of Liver Diseases, Fudan University Shanghai Medical College, Shanghai 200032, China
| | - Ping Liu
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
- Institute of Interdisciplinary Integrative Medicine Research, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
| | - Jiamei Chen
- Institute of Liver diseases, Key Laboratory of Liver and Kidney Diseases (Ministry of Education), Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
- Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
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4
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Lee S, Memon A, Chae SC, Shin D, Choi TY. Epcam regulates intrahepatic bile duct reconstruction in zebrafish, providing a potential model for primary cholangitis model. Biochem Biophys Res Commun 2024; 696:149512. [PMID: 38224664 DOI: 10.1016/j.bbrc.2024.149512] [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: 11/13/2023] [Revised: 12/29/2023] [Accepted: 01/09/2024] [Indexed: 01/17/2024]
Abstract
Epithelial cell adhesion molecules (EpCAMs) have been identified as surface markers of proliferating ductal cells, which are referred to as liver progenitor cells (LPCs), during liver regeneration and correspond to malignancies. These cells can differentiate into hepatocytes and biliary epithelial cells (BECs) in vitro. EpCAM-positive LPCs are involved in liver regeneration following severe liver injury; however, the in vivo function of EpCAMs in the regenerating liver remains unclear. In the present study, we used a zebrafish model of LPC-driven liver regeneration to elucidate the function of EpCAMs in the regenerating liver in vivo. Proliferating ductal cells were observed after severe hepatocyte loss in the zebrafish model. Analyses of the liver size as well as hepatocyte and BEC markers revealed successful conversion of LPCs to hepatocytes and BECs in epcam mutants. Notably, epcam mutants exhibited severe defects in intrahepatic duct maturation and bile acid secretion in regenerating hepatocytes, suggesting that epcam plays a critical role in intrahepatic duct reconstruction during LPC-driven liver regeneration. Our findings provide insights into human diseases involving non-parenchymal cells, such as primary biliary cholangitis, by highlighting the regulatory effect of epcam on intrahepatic duct reconstruction.
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Affiliation(s)
- Siyeo Lee
- Department of Pathology, Digestive Disease Research Institute, Wonkwang University School of Medicine, Iksan, 54538, Republic of Korea; Department of Biomedical Science, Graduate School Wonkwang University, Iksan, Jeonbuk, 54538, Republic of Korea
| | - Azra Memon
- Department of Pathology, Digestive Disease Research Institute, Wonkwang University School of Medicine, Iksan, 54538, Republic of Korea
| | - Soo-Cheon Chae
- Department of Pathology, Digestive Disease Research Institute, Wonkwang University School of Medicine, Iksan, 54538, Republic of Korea
| | - Donghun Shin
- Department of Developmental Biology, McGowan Institute for Regenerative Medicine, University of Pittsburgh, Pittsburgh, PA, 15260, USA
| | - Tae-Young Choi
- Department of Pathology, Digestive Disease Research Institute, Wonkwang University School of Medicine, Iksan, 54538, Republic of Korea; Department of Biomedical Science, Graduate School Wonkwang University, Iksan, Jeonbuk, 54538, Republic of Korea.
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5
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Gabriel V, Zdyrski C, Sahoo DK, Ralston A, Wickham H, Bourgois-Mochel A, Ahmed B, Merodio MM, Paukner K, Piñeyro P, Kopper J, Rowe EW, Smith JD, Meyerholz D, Kol A, Viall A, Elbadawy M, Mochel JP, Allenspach K. Adult Animal Stem Cell-Derived Organoids in Biomedical Research and the One Health Paradigm. Int J Mol Sci 2024; 25:701. [PMID: 38255775 PMCID: PMC10815683 DOI: 10.3390/ijms25020701] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/05/2023] [Revised: 12/14/2023] [Accepted: 12/21/2023] [Indexed: 01/24/2024] Open
Abstract
Preclinical biomedical research is limited by the predictiveness of in vivo and in vitro models. While in vivo models offer the most complex system for experimentation, they are also limited by ethical, financial, and experimental constraints. In vitro models are simplified models that do not offer the same complexity as living animals but do offer financial affordability and more experimental freedom; therefore, they are commonly used. Traditional 2D cell lines cannot fully simulate the complexity of the epithelium of healthy organs and limit scientific progress. The One Health Initiative was established to consolidate human, animal, and environmental health while also tackling complex and multifactorial medical problems. Reverse translational research allows for the sharing of knowledge between clinical research in veterinary and human medicine. Recently, organoid technology has been developed to mimic the original organ's epithelial microstructure and function more reliably. While human and murine organoids are available, numerous other organoids have been derived from traditional veterinary animals and exotic species in the last decade. With these additional organoid models, species previously excluded from in vitro research are becoming accessible, therefore unlocking potential translational and reverse translational applications of animals with unique adaptations that overcome common problems in veterinary and human medicine.
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Affiliation(s)
- Vojtech Gabriel
- Department of Biomedical Sciences, College of Veterinary Medicine, Iowa State University, Ames, IA 50011, USA; (H.W.); (B.A.); (J.P.M.)
| | | | - Dipak K. Sahoo
- Department of Veterinary Clinical Sciences, Iowa State University, Ames, IA 50011, USA; (D.K.S.); (A.B.-M.); (J.K.)
| | - Abigail Ralston
- 3D Health Solutions Inc., Ames, IA 50010, USA; (C.Z.); (A.R.); (M.M.M.)
| | - Hannah Wickham
- Department of Biomedical Sciences, College of Veterinary Medicine, Iowa State University, Ames, IA 50011, USA; (H.W.); (B.A.); (J.P.M.)
| | - Agnes Bourgois-Mochel
- Department of Veterinary Clinical Sciences, Iowa State University, Ames, IA 50011, USA; (D.K.S.); (A.B.-M.); (J.K.)
| | - Basant Ahmed
- Department of Biomedical Sciences, College of Veterinary Medicine, Iowa State University, Ames, IA 50011, USA; (H.W.); (B.A.); (J.P.M.)
| | - Maria M. Merodio
- 3D Health Solutions Inc., Ames, IA 50010, USA; (C.Z.); (A.R.); (M.M.M.)
| | - Karel Paukner
- Atherosclerosis Research Laboratory, Centre for Experimental Medicine, Institute for Clinical and Experimental Medicine, 14021 Prague, Czech Republic;
| | - Pablo Piñeyro
- Department of Veterinary Diagnostic and Production Animal Medicine, College of Veterinary Medicine, Iowa State University, Ames, IA 50011, USA; (P.P.); (J.D.S.)
| | - Jamie Kopper
- Department of Veterinary Clinical Sciences, Iowa State University, Ames, IA 50011, USA; (D.K.S.); (A.B.-M.); (J.K.)
| | - Eric W. Rowe
- Department of Biomedical Sciences, College of Veterinary Medicine, Iowa State University, Ames, IA 50011, USA; (H.W.); (B.A.); (J.P.M.)
| | - Jodi D. Smith
- Department of Veterinary Diagnostic and Production Animal Medicine, College of Veterinary Medicine, Iowa State University, Ames, IA 50011, USA; (P.P.); (J.D.S.)
| | - David Meyerholz
- Department of Pathology, University of Iowa, Iowa City, IA 52242, USA;
| | - Amir Kol
- Department of Pathology, University of California, Davis, CA 94143, USA; (A.K.); (A.V.)
| | - Austin Viall
- Department of Pathology, University of California, Davis, CA 94143, USA; (A.K.); (A.V.)
| | - Mohamed Elbadawy
- Department of Pathology, College of Veterinary Medicine, University of Georgia, Athens, GA 30530, USA;
- Department of Pharmacology, Faculty of Veterinary Medicine, Benha University, Toukh 13736, Egypt
| | - Jonathan P. Mochel
- Department of Biomedical Sciences, College of Veterinary Medicine, Iowa State University, Ames, IA 50011, USA; (H.W.); (B.A.); (J.P.M.)
- 3D Health Solutions Inc., Ames, IA 50010, USA; (C.Z.); (A.R.); (M.M.M.)
- Department of Pathology, College of Veterinary Medicine, University of Georgia, Athens, GA 30530, USA;
| | - Karin Allenspach
- 3D Health Solutions Inc., Ames, IA 50010, USA; (C.Z.); (A.R.); (M.M.M.)
- Department of Veterinary Clinical Sciences, Iowa State University, Ames, IA 50011, USA; (D.K.S.); (A.B.-M.); (J.K.)
- Department of Pathology, College of Veterinary Medicine, University of Georgia, Athens, GA 30530, USA;
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6
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Beumer J, Clevers H. Hallmarks of stemness in mammalian tissues. Cell Stem Cell 2024; 31:7-24. [PMID: 38181752 PMCID: PMC10769195 DOI: 10.1016/j.stem.2023.12.006] [Citation(s) in RCA: 28] [Impact Index Per Article: 28.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2023] [Revised: 12/03/2023] [Accepted: 12/08/2023] [Indexed: 01/07/2024]
Abstract
All adult tissues experience wear and tear. Most tissues can compensate for cell loss through the activity of resident stem cells. Although the cellular maintenance strategies vary greatly between different adult (read: postnatal) tissues, the function of stem cells is best defined by their capacity to replace lost tissue through division. We discuss a set of six complementary hallmarks that are key enabling features of this basic function. These include longevity and self-renewal, multipotency, transplantability, plasticity, dependence on niche signals, and maintenance of genome integrity. We discuss these hallmarks in the context of some of the best-understood adult stem cell niches.
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Affiliation(s)
- Joep Beumer
- Institute of Human Biology (IHB), Roche Pharma Research and Early Development, Basel, Switzerland.
| | - Hans Clevers
- Institute of Human Biology (IHB), Roche Pharma Research and Early Development, Basel, Switzerland.
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7
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Amin MN, El-Far YM, El-Mowafy M, Elgaml A. Tazemetostat decreases β-catenin and CD13 protein expression in HEPG-2 and Hepatitis B virus-transfected HEPG-2 with decreased cell viability. Clin Epigenetics 2023; 15:180. [PMID: 37941056 PMCID: PMC10634085 DOI: 10.1186/s13148-023-01593-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/16/2023] [Accepted: 10/30/2023] [Indexed: 11/10/2023] Open
Abstract
Hepatocellular carcinoma (HCC) is one of the global health concerns. Hepatitis B virus (HBV) is one of the major causes of HCC. Poor clinical outcome of HCC patients is attributed to a small population of cancer cells known as cancer stem cells (CSCs). In this work, we studied the effect of inhibiting the enhancer of zeste homologue 2 (EZH2), a histone methyltransferase known to be overexpressed in CSCs, using tazemetostat (Taz). The effect of Taz was assessed in the HCC cell line (HEPG2) and Hepatitis B virus-transfected HEPG2 (HBV/HEPG2) cells. MTT assay showed a significant decrease in HEPG2 cells viability after 48 h treatment with either 0.5, 1, 4 or 6 μM Taz. HEPG2 and HBV/HEPG2 cells were incubated with either 0.5 or 1 μM Taz for 48 h, and then, the cells and supernatants were collected for protein expression analysis of EZH2, CD13, epithelial cell adhesion molecule (EpCAM) and β-catenin using enzyme-linked immunosorbent assay (ELISA). Taz showed a significant dose-dependent inhibition of EZH2, CD13 and β-catenin in HEPG2 and HBV/HEPG2 cells. Also, EpCAM protein levels were significantly decreased in HBV/HEPG2 but not in HEPG2 cell line alone. Our results indicate that Taz inhibition of EZH2 leads to downregulation of β-catenin signaling and eventually decreased expression of CD13 and EpCAM, which are characteristic for CSCs. The present study suggests that Taz could be a promising treatment for HCC including HBV-induced HCC that might be used in combination with radio/chemotherapy to target CSCs and prevent tumor relapse.
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Affiliation(s)
- Mohamed N Amin
- Biochemistry Department, Faculty of Pharmacy, Mansoura University, Mansoura, 35516, Egypt
| | - Yousra M El-Far
- Biochemistry Department, Faculty of Pharmacy, Mansoura University, Mansoura, 35516, Egypt
| | - Mohammed El-Mowafy
- Microbiology and Immunology Department, Faculty of Pharmacy, Mansoura University, Mansoura, 35516, Egypt
| | - Abdelaziz Elgaml
- Microbiology and Immunology Department, Faculty of Pharmacy, Mansoura University, Mansoura, 35516, Egypt.
- Microbiology and Immunology Department, Faculty of Pharmacy, Horus University, New Damietta, 34518, Egypt.
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8
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Izadpanah A, Mohammadkhani N, Masoudnia M, Ghasemzad M, Saeedian A, Mehdizadeh H, Poorebrahim M, Ebrahimi M. Update on immune-based therapy strategies targeting cancer stem cells. Cancer Med 2023; 12:18960-18980. [PMID: 37698048 PMCID: PMC10557910 DOI: 10.1002/cam4.6520] [Citation(s) in RCA: 10] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2023] [Revised: 08/16/2023] [Accepted: 08/30/2023] [Indexed: 09/13/2023] Open
Abstract
Accumulating data reveals that tumors possess a specialized subset of cancer cells named cancer stem cells (CSCs), responsible for metastasis and recurrence of malignancies, with various properties such as self-renewal, heterogenicity, and capacity for drug resistance. Some signaling pathways or processes like Notch, epithelial to mesenchymal transition (EMT), Hedgehog (Hh), and Wnt, as well as CSCs' surface markers such as CD44, CD123, CD133, and epithelial cell adhesion molecule (EpCAM) have pivotal roles in acquiring CSCs properties. Therefore, targeting CSC-related signaling pathways and surface markers might effectively eradicate tumors and pave the way for cancer survival. Since current treatments such as chemotherapy and radiation therapy cannot eradicate all of the CSCs and tumor relapse may happen following temporary recovery, improving novel and more efficient therapeutic options to combine with current treatments is required. Immunotherapy strategies are the new therapeutic modalities with promising results in targeting CSCs. Here, we review the targeting of CSCs by immunotherapy strategies such as dendritic cell (DC) vaccines, chimeric antigen receptors (CAR)-engineered immune cells, natural killer-cell (NK-cell) therapy, monoclonal antibodies (mAbs), checkpoint inhibitors, and the use of oncolytic viruses (OVs) in pre-clinical and clinical studies. This review will mainly focus on blood malignancies but also describe solid cancers.
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Affiliation(s)
- Amirhossein Izadpanah
- Department of Stem Cells and Developmental Biology, Cell Science Research CenterRoyan Institute for Stem Cell Biology and Technology, ACECRTehranIran
| | - Niloufar Mohammadkhani
- Department of Clinical BiochemistrySchool of Medicine, Shahid Beheshti University of Medical SciencesTehranIran
| | - Mina Masoudnia
- Department of ImmunologySchool of Medicine, Shahid Beheshti University of Medical SciencesTehranIran
| | - Mahsa Ghasemzad
- Department of Stem Cells and Developmental Biology, Cell Science Research CenterRoyan Institute for Stem Cell Biology and Technology, ACECRTehranIran
- Department of Molecular Cell Biology‐Genetics, Faculty of Basic Sciences and Advanced Technologies in BiologyUniversity of Science and CultureTehranIran
| | - Arefeh Saeedian
- Radiation Oncology Research CenterCancer Research Institute, Tehran University of Medical SciencesTehranIran
- Department of Radiation OncologyCancer Institute, Imam Khomeini Hospital Complex, Tehran University of Medical SciencesTehranIran
| | - Hamid Mehdizadeh
- Department of Stem Cells and Developmental Biology, Cell Science Research CenterRoyan Institute for Stem Cell Biology and Technology, ACECRTehranIran
| | - Mansour Poorebrahim
- Arnie Charbonneau Cancer Research Institute, University of CalgaryAlbertaCalgaryCanada
| | - Marzieh Ebrahimi
- Department of Stem Cells and Developmental Biology, Cell Science Research CenterRoyan Institute for Stem Cell Biology and Technology, ACECRTehranIran
- Department of regenerative medicineCell Science research Center, Royan Institute for stem cell biology and technology, ACECRTehranIran
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Pantelidou P, Sinakos E, Germanidis G, Pagkalidou E, Haidich AB, Akriviadis E, Hytiroglou P. Assessment of histologic risk factors for hepatocellular carcinoma in patients with chronic hepatitis B of advanced stage. Pathol Res Pract 2023; 249:154741. [PMID: 37586217 DOI: 10.1016/j.prp.2023.154741] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/11/2023] [Revised: 08/03/2023] [Accepted: 08/03/2023] [Indexed: 08/18/2023]
Abstract
Histologic markers of increased risk for hepatocellular carcinoma can provide useful information for the management of patients with chronic hepatitis B. The expression of epithelial cell adhesion molecule (EpCAM, a marker of hepatic progenitor cells), p21 (a marker of hepatocyte senescence), glutamine synthetase (a marker of perivenular hepatocytes) and CD34 (a marker of sinusoidal capillarization) were assessed by immunohistochemistry in 52 liver biopsy specimens from patients with advanced stage chronic hepatitis B. Nineteen patients developed hepatocellular carcinoma during a follow-up period of 133 months. The findings were compared with those of 18 liver biopsy specimens from patients with early-stage chronic hepatitis B and 6 liver biopsy specimens without significant pathologic findings. EpCAM expression in hepatocytes was significantly increased in specimens with advanced stage, as compared with all other specimens. EpCAM positivity in over 30 % of hepatocytes was only seen in 3 specimens from patients who subsequently developed hepatocellular carcinoma. The expression of p21, glutamine synthetase and CD34 was not associated with hepatocellular carcinoma development. Nevertheless, glutamine synthetase immunostains highlighted zonality abnormalities that were useful in chronic hepatitis B staging. In conclusion, extensive immunopositivity of hepatocytes for EpCAM in chronic hepatitis B may represent a marker of increased hepatocellular carcinoma risk. Glutamine synthetase immunostaining represents a useful adjunct in determining the stage of chronic hepatitis B in diagnostic practice.
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Affiliation(s)
- Pavlina Pantelidou
- Department of Pathology, School of Medicine, Aristotle University of Thessaloniki, Greece
| | - Emmanouil Sinakos
- Fourth Department of Internal Medicine, School of Medicine, Aristotle University of Thessaloniki, Greece
| | - Georgios Germanidis
- First Department of Internal Medicine, School of Medicine, Aristotle University of Thessaloniki, Greece
| | - Eirini Pagkalidou
- Department of Hygiene, Social-Preventive Medicine and Medical Statistics, School of Medicine, Aristotle University of Thessaloniki, Greece
| | - Anna Bettina Haidich
- Department of Hygiene, Social-Preventive Medicine and Medical Statistics, School of Medicine, Aristotle University of Thessaloniki, Greece
| | - Evangelos Akriviadis
- Fourth Department of Internal Medicine, School of Medicine, Aristotle University of Thessaloniki, Greece
| | - Prodromos Hytiroglou
- Department of Pathology, School of Medicine, Aristotle University of Thessaloniki, Greece.
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10
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Ding P, Chen P, Ouyang J, Li Q, Li S. Clinicopathological and prognostic value of epithelial cell adhesion molecule in solid tumours: a meta-analysis. Front Oncol 2023; 13:1242231. [PMID: 37664060 PMCID: PMC10468606 DOI: 10.3389/fonc.2023.1242231] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2023] [Accepted: 07/27/2023] [Indexed: 09/05/2023] Open
Abstract
Background Malignant tumors, mainly solid tumors, are a significant obstacle to the improvement of life expectancy at present. Epithelial cell adhesion molecule (EpCAM), a cancer stem cell biomarker, showed widespread expression in most normal epithelial cells and most cancers. Although the clinical significance of EpCAM in various malignant solid tumors has been studied extensively, the latent relationships between EpCAM and pathological and clinical characteristics in solid tumors and differences in the roles of EpCAM among tumors have not been clearly determined. The destination point of this study was to analyze the value of EpCAM in solid tumors in clinicopathological and prognostic dimension using a meta-analysis approach. Method and materials A comprehensive and systematic search of the researches published up to March 7th, 2022, in PubMed, EMBASE, Web of Science, Cochrane library and PMC databases was performed. The relationships between EpCAM overexpression, clinicopathological characteristics, and survival outcomes were analyzed. Pooled hazard ratios (HRs) with 95% confidence intervals (CIs) and odds ratios (ORs) were estimated as indicators of the degree of correlation. This research was registered on PROSPERO (International prospective register of systematic reviews), ID: CRD42022315070. Results In total, 57 articles and 14184 cases were included in this study. High EpCAM expression had a significant coherence with a poorer overall survival (OS) (HR: 1.30, 95% CI: 1.08-1.58, P < 0.01) and a worse disease-free survival (DFS) (HR: 1.58, 95% CI: 1.28-1.95, P < 0.01), especially of gastrointestinal tumors' OS (HR: 1.50, 95% CI: 1.15-1.95, P < 0.01), and DFS (HR: 1.84, 95% CI: 1.52-2.33, P < 0.01). The DFS of head and neck tumors (HR: 2.33, 95% CI: 1.51-3.61, P < 0.01) was also associated with the overexpression of EpCAM. There were no positive relationships between the overexpression of EpCAM and sex (RR: 1.03, 95% CI: 0.99-1.07, P = 0.141), T classification (RR: 0.93, 95% CI: 0.82-1.06, P = 0.293), lymph node metastasis (RR: 0.85, 95% CI: 0.54-1.32, P = 0.461), distant metastasis (RR: 0.97, 95% CI: 0.84-1.10, P = 0.606), vascular infiltration (RR: 1.05, 95% CI: 0.85-1.29, P = 0.611), and TNM stage (RR: 0.93, 95% CI: 0.83-1.04, P = 0.187). However, the overexpression of EpCAM exhibited a significant association with the histological grades (RR: 0.88, 95% CI: 0.80-0.97, P < 0.01). Conclusion Based on pooled HRs, the positive expression of EpCAM was totally correlated to a worse OS and DFS in solid tumors. The expression of EpCAM was related to a worse OS in gastrointestinal tumors and a worse DFS in gastrointestinal tumors and head and neck tumors. Moreover, EpCAM expression was correlated with the histological grade. The results presented pointed out that EpCAM could serve as a prognostic biomarker for gastrointestinal and head and neck tumors. Systematic review registration https://www.crd.york.ac.uk/prospero, identifier CRD42022315070.
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Affiliation(s)
- Peiwen Ding
- Department of Oncology, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, China
- Clinical School, Chengdu University of Traditional Chinese Medicine, Chengdu, China
| | - Panyu Chen
- Operating Room, Sichuan University West China Hospital School of Nursing, Chengdu, China
| | - Jiqi Ouyang
- Department of Gastroenterology, China Academy of Chinese Medical Sciences Guang’anmen Hospital, Beijing, China
| | - Qiang Li
- Department of Oncology, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, China
- Clinical School, Chengdu University of Traditional Chinese Medicine, Chengdu, China
| | - Shijie Li
- Department of Oncology, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, China
- Clinical School, Chengdu University of Traditional Chinese Medicine, Chengdu, China
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11
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Smith-Cortinez N, Tan AK, Stokroos RJ, Versnel H, Straatman LV. Regeneration of Hair Cells from Endogenous Otic Progenitors in the Adult Mammalian Cochlea: Understanding Its Origins and Future Directions. Int J Mol Sci 2023; 24:ijms24097840. [PMID: 37175547 PMCID: PMC10177935 DOI: 10.3390/ijms24097840] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/13/2023] [Revised: 04/20/2023] [Accepted: 04/21/2023] [Indexed: 05/15/2023] Open
Abstract
Sensorineural hearing loss is caused by damage to sensory hair cells and/or spiral ganglion neurons. In non-mammalian species, hair cell regeneration after damage is observed, even in adulthood. Although the neonatal mammalian cochlea carries regenerative potential, the adult cochlea cannot regenerate lost hair cells. The survival of supporting cells with regenerative potential after cochlear trauma in adults is promising for promoting hair cell regeneration through therapeutic approaches. Targeting these cells by manipulating key signaling pathways that control mammalian cochlear development and non-mammalian hair cell regeneration could lead to regeneration of hair cells in the mammalian cochlea. This review discusses the pathways involved in the development of the cochlea and the impact that trauma has on the regenerative capacity of the endogenous progenitor cells. Furthermore, it discusses the effects of manipulating key signaling pathways targeting supporting cells with progenitor potential to promote hair cell regeneration and translates these findings to the human situation. To improve hearing recovery after hearing loss in adults, we propose a combined approach targeting (1) the endogenous progenitor cells by manipulating signaling pathways (Wnt, Notch, Shh, FGF and BMP/TGFβ signaling pathways), (2) by manipulating epigenetic control, and (3) by applying neurotrophic treatments to promote reinnervation.
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Affiliation(s)
- Natalia Smith-Cortinez
- Department of Otorhinolaryngology and Head & Neck Surgery, University Medical Center Utrecht, Utrecht University, Heidelberglaan 100, 3584 CX Utrecht, The Netherlands
- UMC Utrecht Brain Center, Utrecht University, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands
| | - A Katherine Tan
- Department of Otorhinolaryngology and Head & Neck Surgery, University Medical Center Utrecht, Utrecht University, Heidelberglaan 100, 3584 CX Utrecht, The Netherlands
- UMC Utrecht Brain Center, Utrecht University, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands
| | - Robert J Stokroos
- Department of Otorhinolaryngology and Head & Neck Surgery, University Medical Center Utrecht, Utrecht University, Heidelberglaan 100, 3584 CX Utrecht, The Netherlands
- UMC Utrecht Brain Center, Utrecht University, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands
| | - Huib Versnel
- Department of Otorhinolaryngology and Head & Neck Surgery, University Medical Center Utrecht, Utrecht University, Heidelberglaan 100, 3584 CX Utrecht, The Netherlands
- UMC Utrecht Brain Center, Utrecht University, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands
| | - Louise V Straatman
- Department of Otorhinolaryngology and Head & Neck Surgery, University Medical Center Utrecht, Utrecht University, Heidelberglaan 100, 3584 CX Utrecht, The Netherlands
- UMC Utrecht Brain Center, Utrecht University, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands
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12
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Hou YT, Wu CC, Wang WT, Yang WT, Liao YH, Chen CY. Monitoring Cultured Rat Hepatocytes Using RNA-Seq In Vitro. Int J Mol Sci 2023; 24:ijms24087534. [PMID: 37108701 PMCID: PMC10139060 DOI: 10.3390/ijms24087534] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2023] [Revised: 04/13/2023] [Accepted: 04/14/2023] [Indexed: 04/29/2023] Open
Abstract
Compared to other techniques, RNA sequencing (RNA-Seq) has the advantage of having details of the expression abundance of all transcripts in a single run. In this study, we used RNA-Seq to monitor the maturity and dynamic characteristics of in vitro hepatocyte cultures. Hepatocytes, including mature hepatocytes and small hepatocytes, were analyzed in vitro using RNA-Seq and quantitative polymerase chain reaction (qPCR). The results demonstrated that the gene expression profiles measured by RNA-Seq showed a similar trend to the expression profiles measured by qPCR, and can be used to infer the success of in vitro hepatocyte cultures. The results of the differential analysis, which compared mature hepatocytes against small hepatocytes, revealed 836 downregulated and 137 upregulated genes. In addition, the success of the hepatocyte cultures could be explained by the gene list screened from the adopted gene enrichment test. In summary, we demonstrated that RNA-Seq could become an effective method for monitoring the whole transcriptome of hepatocyte cultures and provide a more comprehensive list of factors related to the differentiation of small hepatocytes into mature hepatocytes. This monitoring system not only shows high potential in medical applications but may also be a novel method for the clinical diagnosis of liver-related diseases.
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Affiliation(s)
- Yung-Te Hou
- Department of Biomechatronics Engineering, National Taiwan University, Taipei 106, Taiwan
| | - Chia-Chun Wu
- Department of Biomechatronics Engineering, National Taiwan University, Taipei 106, Taiwan
| | - Wen-Ting Wang
- Department of Biomechatronics Engineering, National Taiwan University, Taipei 106, Taiwan
| | - Wen-Tse Yang
- Department of Biomechatronics Engineering, National Taiwan University, Taipei 106, Taiwan
| | - Ying-Hsiu Liao
- Department of Biomechatronics Engineering, National Taiwan University, Taipei 106, Taiwan
| | - Chien-Yu Chen
- Department of Biomechatronics Engineering, National Taiwan University, Taipei 106, Taiwan
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13
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Yan ZJ, Chen L, Wang HY. To be or not to be: The double-edged sword roles of liver progenitor cells. Biochim Biophys Acta Rev Cancer 2023; 1878:188870. [PMID: 36842766 DOI: 10.1016/j.bbcan.2023.188870] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2022] [Revised: 01/11/2023] [Accepted: 01/28/2023] [Indexed: 02/28/2023]
Abstract
Given the liver's remarkable and unique regenerative capacity, researchers have long focused on liver progenitor cells (LPCs) and liver cancer stem cells (LCSCs). LPCs can differentiate into both hepatocytes and cholangiocytes. However, the mechanism underlying cell conversion and its distinct contribution to liver homeostasis and tumorigenesis remain unclear. In this review, we discuss the complicated conversions involving LPCs and LCSCs. As the critical intermediate state in malignant transformation, LPCs play double-edged sword roles. LPCs are not only involved in hepatic wound-healing responses by supplementing liver cells and bile duct cells in the damaged liver but may transform into LCSCs under dysregulation of key signaling pathways, resulting in refractory malignant liver tumors. Because LPC lineages are temporally and spatially dynamic, we discuss crucial LPC subgroups and summarize regulatory factors correlating with the trajectories of LPCs and LCSCs in the liver tumor microenvironment. This review elaborates on the double-edged sword roles of LPCs to help understand the liver's regenerative potential and tumor heterogeneity. Understanding the sources and transformations of LPCs is essential in determining how to exploit their regenerative capacity in the future.
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Affiliation(s)
- Zi-Jun Yan
- International Cooperation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Hospital/National Center for Liver Cancer, Shanghai 200438, PR China; Key Laboratory of Signaling Regulation and Targeting Therapy of Liver Cancer (SMMU), Ministry of Education, Shanghai 200438, PR China; Shanghai Key Laboratory of Hepatobiliary Tumor Biology (EHBH), Shanghai 200438, PR China
| | - Lei Chen
- International Cooperation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Hospital/National Center for Liver Cancer, Shanghai 200438, PR China; Key Laboratory of Signaling Regulation and Targeting Therapy of Liver Cancer (SMMU), Ministry of Education, Shanghai 200438, PR China; Shanghai Key Laboratory of Hepatobiliary Tumor Biology (EHBH), Shanghai 200438, PR China.
| | - Hong-Yang Wang
- International Cooperation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Hospital/National Center for Liver Cancer, Shanghai 200438, PR China; Key Laboratory of Signaling Regulation and Targeting Therapy of Liver Cancer (SMMU), Ministry of Education, Shanghai 200438, PR China; Shanghai Key Laboratory of Hepatobiliary Tumor Biology (EHBH), Shanghai 200438, PR China.
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14
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Liver Organoids, Novel and Promising Modalities for Exploring and Repairing Liver Injury. Stem Cell Rev Rep 2023; 19:345-357. [PMID: 36199007 PMCID: PMC9534590 DOI: 10.1007/s12015-022-10456-3] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 09/24/2022] [Indexed: 02/07/2023]
Abstract
The past decades have witnessed great advances in organoid technology. Liver is the biggest solid organ, performing multifaceted physiological functions. Nowadays, liver organoids have been applied in many fields including pharmaceutical research, precision medicine and disease models. Compared to traditional 2-dimensional cell line cultures and animal models, liver organoids showed the unique advantages. More importantly, liver organoids can well model the features of the liver and tend to be novel and promising modalities for exploring liver injury, thus finding potential treatment targets and repairing liver injury. In this review, we reviewed the history of the development of liver organoids and summarized the application of liver organoids and recent studies using organoids to explore and further repair the liver injury. These novel modalities could provide new insights about the process of liver injury.
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15
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SNAI2 Attenuated the Stem-like Phenotype by Reducing the Expansion of EPCAM high Cells in Cervical Cancer Cells. Int J Mol Sci 2023; 24:ijms24021062. [PMID: 36674577 PMCID: PMC9864029 DOI: 10.3390/ijms24021062] [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: 11/12/2022] [Revised: 12/22/2022] [Accepted: 12/27/2022] [Indexed: 01/09/2023] Open
Abstract
SNAI2 (Snai2) is a zinc-finger transcriptional repressor that belongs to the Snail family. The accumulated evidence suggests that SNAI2 exhibits biphasic effects on regulating a stem-like phenotype in various types of cells, both normal and malignant. In this study, by exogenously expressing SNAI2 in SiHa cells, SNAI2 exhibited the capacity to inhibit a stem-like phenotype in cervical cancer cells. The SNAI2-overexpressing cells inhibited cell growth, tumorsphere formation, tumor growth, enhanced sensitivity to cisplatin, reduced stem cell-related factors' expression, and lowered tumor initiating frequency. In addition, the EPCAMhigh cells sorted from SiHa cells exhibited an enhanced capacity to maintain a stem-like phenotype. Further study demonstrated that the trans-suppression of EPCAM expression by SNAI2 led to blockage of the nuclear translocation of β-catenin, as well as reduction in SOX2 and c-Myc expression in SiHa and HeLa cells, but induction in SNAI2 knockdown cells (CaSki), which would be responsible for the attenuation of the stem-like phenotype in cervical cancer cells mediated by SNAI2. All of these results demonstrated that SNAI2 could attenuate the stem-like phenotype in cervical cancer cells through the EPCAM/β-catenin axis.
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16
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Tripura C, Gunda S, Vishwakarma SK, Thatipalli AR, Jose J, Jerald MK, Khan AA, Pande G. Long-term and non-invasive in vivo tracking of DiD dye-labeled human hepatic progenitors in chronic liver disease models. World J Hepatol 2022; 14:1884-1898. [PMID: 36340748 PMCID: PMC9627437 DOI: 10.4254/wjh.v14.i10.1884] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/26/2022] [Revised: 09/02/2022] [Accepted: 10/02/2022] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND Chronic liver diseases (CLD) are the major public health burden due to the continuous increasing rate of global morbidity and mortality. The inherent limitations of organ transplantation have led to the development of stem cell-based therapy as a supportive and promising therapeutic option. However, identifying the fate of transplanted cells in vivo represents a crucial obstacle. AIM To evaluate the potential applicability of DiD dye as a cell labeling agent for long-term, and non-invasive in vivo tracking of transplanted cells in the liver. METHODS Magnetically sorted, epithelial cell adhesion molecule positive (1 × 106 cells/mL) fetal hepatic progenitor cells were labeled with DiD dye and transplanted into the livers of CLD-severe combined immunodeficiency (SCID) mice. Near-infrared (NIR) imaging was performed for in vivo tracking of the DiD-labeled transplanted cells along with colocalization of hepatic markers for up to 80 d. The existence of human cells within mouse livers was identified using Alu polymerase chain reaction and sequencing. RESULTS NIR fluorescence imaging of CLD-SCID mice showed a positive fluorescence signal of DiD at days 7, 15, 30, 45, 60, and 80 post-transplantation. Furthermore, positive staining of cytokeratin, c-Met, and albumin colocalizing with DiD fluorescence clearly demonstrated that the fluorescent signal of hepatic markers emerged from the DiD-labeled transplanted cells. Recovery of liver function was also observed with serum levels of glutamic-oxaloacetic transaminase, glutamate-pyruvate transaminase, and bilirubin. The detection of human-specific Alu sequence from the transplanted mouse livers provided evidence for the survival of transplanted cells at day 80. CONCLUSION DiD-labeling is promising for long-term and non-invasive in vivo cell tracking, and understanding the regenerative mechanisms incurred by the transplanted cells.
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Affiliation(s)
- Chaturvedula Tripura
- Cell and Stem Cell Biology, CSIR-Centre for Cellular and Molecular Biology, Hyderabad 500007, Telangana, India.
| | - Srinivas Gunda
- Cell and Stem Cell Biology, CSIR-Centre for Cellular and Molecular Biology, Hyderabad 500007, Telangana, India
| | - Sandeep Kumar Vishwakarma
- Central Laboratory for Stem Cell Research and Translational Medicine, Centre for Liver Research and Diagnostics, Deccan College of Medical Sciences, Hyderabad 500058, Telangana, India
| | - Avinash Raj Thatipalli
- Cell and Stem Cell Biology, CSIR-Centre for Cellular and Molecular Biology, Hyderabad 500007, Telangana, India
| | - Jedy Jose
- Cell and Stem Cell Biology, CSIR-Centre for Cellular and Molecular Biology, Hyderabad 500007, Telangana, India
| | - Mahesh Kumar Jerald
- Cell and Stem Cell Biology, CSIR-Centre for Cellular and Molecular Biology, Hyderabad 500007, Telangana, India
| | - Aleem Ahmed Khan
- Central Laboratory for Stem Cell Research and Translational Medicine, Centre for Liver Research and Diagnostics, Deccan College of Medical Sciences, Hyderabad 500058, Telangana, India
| | - Gopal Pande
- Cell and Stem Cell Biology, CSIR-Centre for Cellular and Molecular Biology, Hyderabad 500007, Telangana, India
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17
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Palao N, Sequera C, Cuesta ÁM, Baquero C, Bragado P, Gutierrez-Uzquiza A, Sánchez A, Guerrero C, Porras A. C3G down-regulation enhances pro-migratory and stemness properties of oval cells by promoting an epithelial-mesenchymal-like process. Int J Biol Sci 2022; 18:5873-5884. [PMID: 36263169 PMCID: PMC9576514 DOI: 10.7150/ijbs.73192] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/24/2022] [Accepted: 08/11/2022] [Indexed: 01/12/2023] Open
Abstract
Previous data indicate that C3G (RapGEF1) main isoform is highly expressed in liver progenitor cells (or oval cells) compared to adult mature hepatocytes, suggesting it may play an important role in oval cell biology. Hence, we have explored C3G function in the regulation of oval cell properties by permanent gene silencing using shRNAs. We found that C3G knock-down enhanced migratory and invasive ability of oval cells by promoting a partial epithelial to mesenchymal transition (EMT). This is likely mediated by upregulation of mRNA expression of the EMT-inducing transcription factors, Snail1, Zeb1 and Zeb2, induced in C3G-silenced oval cells. This EMT is associated to a higher expression of the stemness markers, CD133 and CD44. Moreover, C3G down-regulation increased oval cells clonogenic capacity by enhancing cell scattering. However, C3G knock-down did not impair oval cell differentiation into hepatocyte lineage. Mechanistic studies revealed that HGF/MET signaling and its pro-invasive activity was impaired in oval cells with low levels of C3G, while TGF-β signaling was increased. Altogether, these data suggest that C3G might be tightly regulated to ensure liver repair in chronic liver diseases such as non-alcoholic steatohepatitis. Hence, reduced C3G levels could facilitate oval cell expansion, after the proliferation peak, by enhancing migration.
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Affiliation(s)
- Nerea Palao
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad Complutense de Madrid; 28040 Madrid, Spain.,Instituto de Investigación Sanitaria del Hospital Clínico San Carlos (IdISSC), 28040 Madrid, Spain
| | - Celia Sequera
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad Complutense de Madrid; 28040 Madrid, Spain.,Instituto de Investigación Sanitaria del Hospital Clínico San Carlos (IdISSC), 28040 Madrid, Spain.,Aix-Marseille Univ, CNRS, Developmental Biology Institute of Marseille (IBDM), Turing Center for Living Systems, Parc Scientifique de Luminy, 13009 Marseille, France
| | - Ángel M Cuesta
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad Complutense de Madrid; 28040 Madrid, Spain.,Instituto de Investigación Sanitaria del Hospital Clínico San Carlos (IdISSC), 28040 Madrid, Spain
| | - Cristina Baquero
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad Complutense de Madrid; 28040 Madrid, Spain.,Instituto de Investigación Sanitaria del Hospital Clínico San Carlos (IdISSC), 28040 Madrid, Spain
| | - Paloma Bragado
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad Complutense de Madrid; 28040 Madrid, Spain.,Instituto de Investigación Sanitaria del Hospital Clínico San Carlos (IdISSC), 28040 Madrid, Spain
| | - Alvaro Gutierrez-Uzquiza
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad Complutense de Madrid; 28040 Madrid, Spain.,Instituto de Investigación Sanitaria del Hospital Clínico San Carlos (IdISSC), 28040 Madrid, Spain
| | - Aránzazu Sánchez
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad Complutense de Madrid; 28040 Madrid, Spain.,Instituto de Investigación Sanitaria del Hospital Clínico San Carlos (IdISSC), 28040 Madrid, Spain
| | - Carmen Guerrero
- Instituto de Biología Molecular y Celular del Cáncer (IBMCC), Universidad de Salamanca-CSIC, 37007 Salamanca, Spain.,Instituto de Investigación Biomédica de Salamanca (IBSAL), 37007 Salamanca, Spain.,Departamento de Medicina, Universidad de Salamanca, 37007 Salamanca, Spain.,✉ Corresponding authors: A. Porras, Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, UCM, Ciudad Universitaria, Madrid, Spain. Tel.: +34 913941627; E-mail: . Co-correspondence: C. Guerrero, Centro de Investigación del Cáncer, Campus Unamuno s/n, Salamanca, Spain. Tel.: +34 923294801; Fax.: +34 923294795; e-mail:
| | - Almudena Porras
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad Complutense de Madrid; 28040 Madrid, Spain.,Instituto de Investigación Sanitaria del Hospital Clínico San Carlos (IdISSC), 28040 Madrid, Spain.,✉ Corresponding authors: A. Porras, Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, UCM, Ciudad Universitaria, Madrid, Spain. Tel.: +34 913941627; E-mail: . Co-correspondence: C. Guerrero, Centro de Investigación del Cáncer, Campus Unamuno s/n, Salamanca, Spain. Tel.: +34 923294801; Fax.: +34 923294795; e-mail:
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18
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Cessna H, Baritaki S, Zaravinos A, Bonavida B. The Role of RKIP in the Regulation of EMT in the Tumor Microenvironment. Cancers (Basel) 2022; 14:cancers14194596. [PMID: 36230521 PMCID: PMC9559516 DOI: 10.3390/cancers14194596] [Citation(s) in RCA: 18] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2022] [Revised: 09/21/2022] [Accepted: 09/21/2022] [Indexed: 12/03/2022] Open
Abstract
Simple Summary Raf kinase inhibitor protein (RKIP) expression in cancer cells is significantly reduced and promoting cancer cells growth and invasiveness. Overexpresssion of RKIP has been reported to mediate pleiotropic anti-cancer activities including the inhibition of survival signaling pathways, sensitization to cell death by cytotoxic drugs, inhibition of invasion, EMT and metastasis. The molecular mechanism by which RKIP inhibits EMT is not clear. In this review, we have examined how RKIP inhibits the selected EMT gene products (Snail, vimentin, N-cadherin, laminin alpha) and found that it involves signaling cross-talks between RKIP and each of the EMT gene products. These findings were validated by bioinformatic analyses demonstrating in various human cancers a negative correlation between the expression of RKIP and the expression of the EMT gene products. These findings suggest that targeting RKIP induction in cancer cells will result in multiple hits by inhibiting tumor growth, metastasis and reversal of chemo-immuno resistance. Abstract The Raf Kinase Inhibitor Protein (RKIP) is a unique gene product that directly inhibits the Raf/Mek/Erk and NF-kB pathways in cancer cells and resulting in the inhibition of cell proliferation, viability, EMT, and metastasis. Additionally, RKIP is involved in the regulation of cancer cell resistance to both chemotherapy and immunotherapy. The low expression of RKIP expression in many cancer types is responsible, in part, for the pathogenesis of cancer and its multiple properties. The inhibition of EMT and metastasis by RKIP led to its classification as a tumor suppressor. However, the mechanism by which RKIP mediates its inhibitory effects on EMT and metastases was not clear. We have proposed that one mechanism involves the negative regulation by RKIP of the expression of various gene products that mediate the mesenchymal phenotype as well as the positive regulation of gene products that mediate the epithelial phenotype via signaling cross talks between RKIP and each gene product. We examined several EMT mesenchymal gene products such as Snail, vimentin, N-cadherin, laminin and EPCAM and epithelial gene products such as E-cadherin and laminin. We have found that indeed these negative and positive correlations were detected in the signaling cross-talks. In addition, we have also examined bioinformatic data sets on different human cancers and the findings corroborated, in large part, the findings observed in the signaling cross-talks with few exceptions in some cancer types. The overall findings support the underlying mechanism by which the tumor suppressor RKIP regulates the expression of gene products involved in EMT and metastasis. Hence, the development of agent that can selectively induce RKIP expression in cancers with low expressions should result in the activation of the pleiotropic anti-cancer activities of RKIP and resulting in multiple effects including inhibition of tumor cell proliferation, EMT, metastasis and sensitization of resistant tumor cells to respond to both chemotherapeutics and immunotherapeutics.
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Affiliation(s)
- Hannah Cessna
- Department of Microbiology, Immunology & Molecular Genetics, David Geffen School of Medicine, Jonsson Comprehensive Cancer Center, University of California at Los Angeles, Los Angeles, CA 90095, USA
| | - Stavroula Baritaki
- Laboratory of Experimental Oncology, Division of Surgery, School of Medicine, University of Crete, 71003 Heraklion, Greece
| | - Apostolos Zaravinos
- Department of Life Sciences, School of Sciences, European University Cyprus, Nicosia 2404, Cyprus
- Basic and Translational Cancer Research Center (BTCRC), Cancer Genetics, Genomics and Systems Biology Laboratory, Nicosia 1516, Cyprus
| | - Benjamin Bonavida
- Department of Microbiology, Immunology & Molecular Genetics, David Geffen School of Medicine, Jonsson Comprehensive Cancer Center, University of California at Los Angeles, Los Angeles, CA 90095, USA
- Correspondence:
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19
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Kim H, Im I, Jeon JS, Kang EH, Lee HA, Jo S, Kim JW, Woo DH, Choi YJ, Kim HJ, Han JS, Lee BS, Kim JH, Kim SK, Park HJ. Development of human pluripotent stem cell-derived hepatic organoids as an alternative model for drug safety assessment. Biomaterials 2022; 286:121575. [DOI: 10.1016/j.biomaterials.2022.121575] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2021] [Revised: 04/15/2022] [Accepted: 05/09/2022] [Indexed: 12/12/2022]
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20
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Novoselova M, Chernyshev VS, Schulga A, Konovalova EV, Chuprov-Netochin RN, Abakumova TO, German S, Shipunova VO, Mokrousov MD, Prikhozhdenko E, Bratashov DN, Nozdriukhin DV, Bogorodskiy A, Grishin O, Kosolobov SS, Khlebtsov BN, Inozemtseva O, Zatsepin TS, Deyev SM, Gorin DA. Effect of Surface Modification of Multifunctional Nanocomposite Drug Delivery Carriers with DARPin on Their Biodistribution In Vitro and In Vivo. ACS APPLIED BIO MATERIALS 2022; 5:2976-2989. [PMID: 35616387 DOI: 10.1021/acsabm.2c00289] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
We present a targeted drug delivery system for therapy and diagnostics that is based on a combination of contrasting, cytotoxic, and cancer-cell-targeting properties of multifunctional carriers. The system uses multilayered polymer microcapsules loaded with magnetite and doxorubicin. Loading of magnetite nanoparticles into the polymer shell by freezing-induced loading (FIL) allowed the loading efficiency to be increased 5-fold, compared with the widely used layer-by-layer (LBL) assembly. FIL also improved the photoacoustic signal and particle mobility in a magnetic field gradient, a result unachievable by the LBL alone. For targeted delivery of the carriers to cancer cells, the carrier surface was modified with a designed ankyrin repeat protein (DARPin) directed toward the epithelial cell adhesion molecule (EpCAM). Flow cytometry measurements showed that the DARPin-coated capsules specifically interacted with the surface of EpCAM-overexpressing human cancer cells such as MCF7. In vivo and ex vivo biodistribution studies in FvB mice showed that the carrier surface modification with DARPin changed the biodistribution of the capsules toward epithelial cells. In particular, the capsules accumulated substantially in the lungs─a result that can be effectively used in targeted lung cancer therapy. The results of this work may aid in the further development of the "magic bullet" concept and may bring the quality of personalized medicine to another level.
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Affiliation(s)
- Marina Novoselova
- Skolkovo Institute of Science and Technology, Bolshoy Boulevard 30, Building 1, Moscow 121205, Russia
| | - Vasiliy S Chernyshev
- Skolkovo Institute of Science and Technology, Bolshoy Boulevard 30, Building 1, Moscow 121205, Russia.,School of Biological and Medical Physics, Moscow Institute of Physics & Technology, Dolgoprudnyi, Moscow Region 141700, Russia
| | - Alexey Schulga
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, Ul. Miklukho-Maklaya 16/10, Moscow 117997, Russia
| | - Elena V Konovalova
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, Ul. Miklukho-Maklaya 16/10, Moscow 117997, Russia
| | - Roman N Chuprov-Netochin
- School of Biological and Medical Physics, Moscow Institute of Physics & Technology, Dolgoprudnyi, Moscow Region 141700, Russia
| | - Tatiana O Abakumova
- Skolkovo Institute of Science and Technology, Bolshoy Boulevard 30, Building 1, Moscow 121205, Russia
| | - Sergei German
- Skolkovo Institute of Science and Technology, Bolshoy Boulevard 30, Building 1, Moscow 121205, Russia.,Institute of Spectroscopy of the Russian Academy of Sciences, Moscow 108840, Russia
| | - Victoria O Shipunova
- School of Biological and Medical Physics, Moscow Institute of Physics & Technology, Dolgoprudnyi, Moscow Region 141700, Russia.,Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, Ul. Miklukho-Maklaya 16/10, Moscow 117997, Russia
| | - Maksim D Mokrousov
- Skolkovo Institute of Science and Technology, Bolshoy Boulevard 30, Building 1, Moscow 121205, Russia
| | | | - Daniil N Bratashov
- Saratov State University, 83 Astrakhanskaya Street, Saratov 410012, Russia
| | - Daniil V Nozdriukhin
- Skolkovo Institute of Science and Technology, Bolshoy Boulevard 30, Building 1, Moscow 121205, Russia
| | - Andrey Bogorodskiy
- School of Biological and Medical Physics, Moscow Institute of Physics & Technology, Dolgoprudnyi, Moscow Region 141700, Russia
| | - Oleg Grishin
- Saratov State University, 83 Astrakhanskaya Street, Saratov 410012, Russia
| | - Sergey S Kosolobov
- Skolkovo Institute of Science and Technology, Bolshoy Boulevard 30, Building 1, Moscow 121205, Russia
| | - Boris N Khlebtsov
- Institute of Biochemistry and Physiology of Plants and Microorganisms, Russian Academy of Sciences, Saratov 410049, Russia
| | - Olga Inozemtseva
- Saratov State University, 83 Astrakhanskaya Street, Saratov 410012, Russia
| | - Timofei S Zatsepin
- Skolkovo Institute of Science and Technology, Bolshoy Boulevard 30, Building 1, Moscow 121205, Russia.,Lomonosov Moscow State University, Moscow 119991, Russia
| | - Sergey M Deyev
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, Ul. Miklukho-Maklaya 16/10, Moscow 117997, Russia
| | - Dmitry A Gorin
- Skolkovo Institute of Science and Technology, Bolshoy Boulevard 30, Building 1, Moscow 121205, Russia
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21
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Sun S, Li JY, Nim HT, Piers A, Ramialison M, Porrello ER, Konstantinov IE, Elefanty AG, Stanley EG. CD90 Marks a Mesenchymal Program in Human Thymic Epithelial Cells In Vitro and In Vivo. Front Immunol 2022; 13:846281. [PMID: 35371075 PMCID: PMC8966383 DOI: 10.3389/fimmu.2022.846281] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2021] [Accepted: 02/18/2022] [Indexed: 11/13/2022] Open
Abstract
Thymic epithelium is critical for the structural integrity of the thymus and for T cell development. Within the fully formed thymus, large numbers of hematopoietic cells shape the thymic epithelium into a scaffold-like structure which bears little similarity to classical epithelial layers, such as those observed in the skin, intestine or pancreas. Here, we show that human thymic epithelial cells (TECs) possess an epithelial identity that also incorporates the expression of mesenchymal cell associated genes, whose expression levels vary between medullary and cortical TECs (m/cTECs). Using pluripotent stem cell (PSC) differentiation systems, we identified a unique population of cells that co-expressed the master TEC transcription factor FOXN1, as well as the epithelial associated marker EPCAM and the mesenchymal associated gene CD90. Using the same serum free culture conditions, we also observed co-expression of EPCAM and CD90 on cultured TECs derived from neonatal human thymus in vitro. Single cell RNA-sequencing revealed these cultured TECs possessed an immature mTEC phenotype and expressed epithelial and mesenchymal associated genes, such as EPCAM, CLDN4, CD90 and COL1A1. Importantly, flow cytometry and single cell RNA-sequencing analysis further confirmed the presence of an EPCAM+CD90+ population in the CD45- fraction of neonatal human thymic stromal cells in vivo. Using the human thymus cell atlas, we found that cTECs displayed more pronounced mesenchymal characteristics than mTECs during embryonic development. Collectively, these results suggest human TECs possess a hybrid gene expression program comprising both epithelial and mesenchymal elements, and provide a basis for the further exploration of thymus development from primary tissues and from the in vitro differentiation of PSCs.
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Affiliation(s)
- Shicheng Sun
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, VIC, Australia.,Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, VIC, Australia.,The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, VIC, Australia
| | - Jacky Y Li
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, VIC, Australia.,Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, VIC, Australia.,The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, VIC, Australia
| | - Hieu T Nim
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, VIC, Australia.,Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, VIC, Australia.,The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, VIC, Australia.,Australian Regenerative Medicine Institute and Systems Biology Institute Australia, Monash University, Clayton, VIC, Australia
| | - Adam Piers
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, VIC, Australia.,Melbourne Centre for Cardiovascular Genomics and Regenerative Medicine, Royal Children's Hospital, Melbourne, VIC, Australia
| | - Mirana Ramialison
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, VIC, Australia.,Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, VIC, Australia.,The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, VIC, Australia.,Australian Regenerative Medicine Institute and Systems Biology Institute Australia, Monash University, Clayton, VIC, Australia
| | - Enzo R Porrello
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, VIC, Australia.,Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, VIC, Australia.,The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, VIC, Australia.,Melbourne Centre for Cardiovascular Genomics and Regenerative Medicine, Royal Children's Hospital, Melbourne, VIC, Australia
| | - Igor E Konstantinov
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, VIC, Australia.,Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, VIC, Australia.,Australian Regenerative Medicine Institute and Systems Biology Institute Australia, Monash University, Clayton, VIC, Australia.,Department of Cardiac Surgery, Royal Children's Hospital, Melbourne, VIC, Australia
| | - Andrew G Elefanty
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, VIC, Australia.,Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, VIC, Australia.,The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, VIC, Australia.,Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC, Australia
| | - Edouard G Stanley
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, VIC, Australia.,Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, VIC, Australia.,The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, VIC, Australia.,Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC, Australia
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22
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Kan S, Grainge C, Nichol K, Reid A, Knight D, Sun Y, Bartlett N, Liang M. TLR7 agonist loaded airway epithelial targeting nanoparticles stimulate innate immunity and suppress viral replication in human bronchial epithelial cells. Int J Pharm 2022; 617:121586. [PMID: 35181464 DOI: 10.1016/j.ijpharm.2022.121586] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/17/2021] [Revised: 02/06/2022] [Accepted: 02/11/2022] [Indexed: 11/29/2022]
Abstract
Nanoparticle-based delivery is a strategy for increasing the therapeutic window of inhaled immunomodulatory drugs that have inflammatory activity. TLR7 agonists are a class of immunomodulators that have been considered for the treatment of virus-induced respiratory diseases. However, due to high immune-stimulatory activity, TLR7 agonists, delivered via direct exposure, generally have a narrow therapeutic window. To address this, we have developed lipid/polymer hybrid nanoparticles (NPs) conjugated with anti-EpCAM monoclonal antibody for targeted delivery of TLR7 agonist (CL264) to airway epithelial cells (AECs)2 - the primary site of respiratory virus infection. These airway epithelial targeting nanoparticles (AEC-NPs)3 showed safety and biocompatibility, and approximately two-fold increased cellular uptake compared to non-targeting NPs. Upon cell entry, AEC-NPs were able to deliver CL264 to cytoplasm and endosomes where TLR7 is located. CL264 delivered by AEC-NPs significantly increased innate immune response through expression of IFN-β, IFN-λ 2/3 and IFN-stimulated genes and suppressed more than 92% of viral load at 48 hours post-infection compared to the drug alone and non-targeting NPs. In conclusion, AEC-NPs exhibited increased cellular uptake leading to enhanced innate immune activation and suppression of viral replication. These findings support the use of AEC-targeting approach for delivering drugs with a narrow therapeutic window.
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Affiliation(s)
- Stanislav Kan
- School of Biomedical Sciences and Pharmacy, The University of Newcastle, Callaghan, New South Wales, Australia; Priority Research Centre for Healthy Lungs, Hunter Medical Research Institute, The University of Newcastle, New South Wales, Australia
| | - Christopher Grainge
- School of Medicine and Public Health, The University of Newcastle, Callaghan, New South Wales, Australia; Priority Research Centre for Healthy Lungs, Hunter Medical Research Institute, The University of Newcastle, New South Wales, Australia
| | - Kristy Nichol
- Priority Research Centre for Healthy Lungs, Hunter Medical Research Institute, The University of Newcastle, New South Wales, Australia
| | - Andrew Reid
- School of Medicine and Public Health, The University of Newcastle, Callaghan, New South Wales, Australia; Priority Research Centre for Healthy Lungs, Hunter Medical Research Institute, The University of Newcastle, New South Wales, Australia
| | - Darryl Knight
- Department of Anaesthesiology, Pharmacology and Therapeutics, Faculty of Medicine, University of British Columbia, Vancouver, Canada
| | - Yong Sun
- Department of Pharmaceutics, School of Pharmacy, Qingdao University, Qingdao, P. R. China
| | - Nathan Bartlett
- School of Biomedical Sciences and Pharmacy, The University of Newcastle, Callaghan, New South Wales, Australia; Priority Research Centre for Healthy Lungs, Hunter Medical Research Institute, The University of Newcastle, New South Wales, Australia
| | - Mingtao Liang
- School of Biomedical Sciences and Pharmacy, The University of Newcastle, Callaghan, New South Wales, Australia.
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23
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Yu W, Ma Y, Shrivastava SK, Srivastava RK, Shankar S. Chronic alcohol exposure induces hepatocyte damage by inducing oxidative stress, SATB2 and stem cell‐like characteristics, and activating lipogenesis. J Cell Mol Med 2022; 26:2119-2131. [PMID: 35152538 PMCID: PMC8980954 DOI: 10.1111/jcmm.17235] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2021] [Revised: 01/25/2022] [Accepted: 01/31/2022] [Indexed: 12/12/2022] Open
Abstract
Alcohol is a risk factor for hepatocellular carcinoma (HCC). However, the molecular mechanism by which chronic alcohol consumption contributes to HCC is not well understood. The purpose of the study was to demonstrate the effects of chronic ethanol exposure on the damage of human normal hepatocytes. Our data showed that chronic exposure of hepatocytes with ethanol induced changes similar to transformed hepatocytes that is, exhibited colonies and anchorage‐independent growth. These damaged hepatocytes contained high levels of reactive oxygen species (ROS) and showed induction of the SATB2 gene. Furthermore, damaged hepatocytes gained the phenotypes of CSCs which expressed stem cell markers (CD133, CD44, CD90, EpCAM, AFP and LGR5), and pluripotency maintaining factors (Sox‐2, POU5F1/Oct4 and KLF‐4). Ethanol exposure also induced Nanog, a pluripotency maintaining transcription factor that functions in concert with Oct4 and SOX‐2. Furthermore, ethanol induced expression of EMT‐related transcription factors (Snail, Slug and Zeb1), N‐Cadherin, and inhibited E‐cadherin expression in damaged hepatocytes. Ethanol enhanced recruitment of SATB2 to promoters of Bcl‐2, Nanog, c‐Myc, Klf4 and Oct4. Ethanol also induced activation of the Wnt/TCF‐LEF1 pathway and its targets (Bcl‐2, Cyclin D1, AXIN2 and Myc). Finally, ethanol induced hepatocellular steatosis, SREBP1 transcription, and modulated the expression of SREBP1c, ACAC, ACLY, FASN, IL‐1β, IL‐6, TNF‐α, GPC3, FLNB and p53. These data suggest that chronic alcohol consumption may contribute towards the development of HCC by damaging normal hepatocytes with the generation of inflammatory environment, induction of SATB2, stem cell‐like characteristics, and cellular steatosis.
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Affiliation(s)
- Wei Yu
- Kansas City VA Medical Center Kansas City Missouri USA
| | - Yiming Ma
- Kansas City VA Medical Center Kansas City Missouri USA
| | - Sushant K. Shrivastava
- Department of Pharmaceutics Indian Institute of Technology Banaras Hindu University Varanasi U.P. India
| | - Rakesh K. Srivastava
- Kansas City VA Medical Center Kansas City Missouri USA
- Department of Genetics Louisiana State University Health Sciences Center New Orleans Louisina USA
- Stanley S. Scott Cancer Center Department of Genetics Louisiana State University Health Sciences Center New Orleans Louisina USA
- A.B. Freeman School of Business Tulane University New Orleans Louisina USA
| | - Sharmila Shankar
- Kansas City VA Medical Center Kansas City Missouri USA
- John W. Deming Department of Medicine Tulane University School of Medicine New Orleans Louisina USA
- Southeast Louisiana Veterans Health Care System New Orleans Louisina USA
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24
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Integration of miRNA-lncRNA-mRNA profiles in liver tissue from EpCAM knockout mice. ARCH BIOL SCI 2022. [DOI: 10.2298/abs211207001l] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/27/2022] Open
Abstract
The epithelial cell adhesion molecule (EpCAM) is highly expressed in the
liver during development and diseases. However, its role in the development
and pathology of liver remains to be explored. The liver tissues of EpCAM-/-
and wildtype (WT) mice at P0 stage were used for RNA sequencing. The
differently expressed miRNAs, lncRNAs and mRNAs were selected and confirmed
by qPCR. The expression of metabolism-related gene SET domain bifurcated 2
(Setdb2) was significantly increased in the liver of EpCAM-/- mice; the
triglyceride (TG) and total cholesterol (TC) levels in the liver were also
markedly decreased in EpCAM-/- mice. The microRNA (miRNA)-long noncoding RNA
(lncRNA)-mRNA regulatory networks indicated that EpCAM may play important
roles in glucose and lipid metabolism of the liver during development and in
disease. The comprehensive miRNA, lncRNA and mRNA expression profiles in the
developing liver of EpCAM-/- mice established here might help to elucidate
functions and mechanisms of EpCAM during development and in diseases of the
liver.
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25
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Choi J, Kang S, Kim B, So S, Han J, Kim GN, Lee MY, Roh S, Lee JY, Oh SJ, Sung YH, Lee Y, Kim SH, Kang E. Efficient hepatic differentiation and regeneration potential under xeno-free conditions using mass-producible amnion-derived mesenchymal stem cells. Stem Cell Res Ther 2021; 12:569. [PMID: 34772451 PMCID: PMC8588618 DOI: 10.1186/s13287-021-02470-y] [Citation(s) in RCA: 3] [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: 11/24/2020] [Accepted: 06/22/2021] [Indexed: 12/18/2022] Open
Abstract
BACKGROUND Amnion-derived mesenchymal stem cells (AM-MSCs) are an attractive source of stem cell therapy for patients with irreversible liver disease. However, there are obstacles to their use due to low efficiency and xeno-contamination for hepatic differentiation. METHODS We established an efficient protocol for differentiating AM-MSCs into hepatic progenitor cells (HPCs) by analyzing transcriptome-sequencing data. Furthermore, to generate the xeno-free conditioned differentiation protocol, we replaced fetal bovine serum (FBS) with polyvinyl alcohol (PVA). We investigated the hepatocyte functions with the expression of mRNA and protein, secretion of albumin, and activity of CYP3A4. Finally, to test the transplantable potential of HPCs, we transferred AM-MSCs along with hepatic progenitors after differentiated days 11, 12, and 13 based on the expression of hepatocyte-related genes and mitochondrial function. Further, we established a mouse model of acute liver failure using a thioacetamide (TAA) and cyclophosphamide monohydrate (CTX) and transplanted AM-HPCs in the mouse model through splenic injection. RESULTS We analyzed gene expression from RNA sequencing data in AM-MSCs and detected downregulation of hepatic development-associated genes including GATA6, KIT, AFP, c-MET, FGF2, EGF, and c-JUN, and upregulation of GSK3. Based on this result, we established an efficient hepatic differentiation protocol using the GSK3 inhibitor, CHIR99021. Replacing FBS with PVA resulted in improved differentiation ability, such as upregulation of hepatic maturation markers. The differentiated hepatocyte-like cells (HLCs) not only synthesized and secreted albumin, but also metabolized drugs by the CYP3A4 enzyme. The best time for translation of AM-HPCs was 12 days from the start of differentiation. When the AM-HPCs were transplanted into the liver failure mouse model, they settled in the damaged livers and differentiated into hepatocytes. CONCLUSION This study offers an efficient and xeno-free conditioned hepatic differentiation protocol and shows that AM-HPCs could be used as transplantable therapeutic materials. Thus, we suggest that AM-MSC-derived HPCs are promising cells for treating liver disease.
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Affiliation(s)
- Jiwan Choi
- Department of Medical Science, Asan Medical Institute of Convergence Science and Technology (AMIST), University of Ulsan College of Medicine, Asan Medical Center, Seoul, 05505, South Korea
- Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, South Korea
- Present Address: Center for Embryo & Stem Cell Research, CHA Advanced Research Institute and Department of Biomedical Science, CHA University, Pocheon-si, Gyeonggi, 13488, South Korea
| | - Seoon Kang
- Department of Medical Science, Asan Medical Institute of Convergence Science and Technology (AMIST), University of Ulsan College of Medicine, Asan Medical Center, Seoul, 05505, South Korea
- Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, South Korea
- Present Address: Center for Embryo & Stem Cell Research, CHA Advanced Research Institute and Department of Biomedical Science, CHA University, Pocheon-si, Gyeonggi, 13488, South Korea
| | - Bitnara Kim
- Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, South Korea
- Present Address: Center for Embryo & Stem Cell Research, CHA Advanced Research Institute and Department of Biomedical Science, CHA University, Pocheon-si, Gyeonggi, 13488, South Korea
| | - Seongjun So
- Department of Medical Science, Asan Medical Institute of Convergence Science and Technology (AMIST), University of Ulsan College of Medicine, Asan Medical Center, Seoul, 05505, South Korea
- Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, South Korea
| | - Jongsuk Han
- Department of Medical Science, Asan Medical Institute of Convergence Science and Technology (AMIST), University of Ulsan College of Medicine, Asan Medical Center, Seoul, 05505, South Korea
- Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, South Korea
- Present Address: Center for Embryo & Stem Cell Research, CHA Advanced Research Institute and Department of Biomedical Science, CHA University, Pocheon-si, Gyeonggi, 13488, South Korea
| | - Gyeong-Nam Kim
- Department of Medical Science, Asan Medical Institute of Convergence Science and Technology (AMIST), University of Ulsan College of Medicine, Asan Medical Center, Seoul, 05505, South Korea
- Convergence Medicine Research Center, Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, South Korea
| | - Mi-Young Lee
- Department of Obstetrics and Gynecology, University of Ulsan College of Medicine, Asan Medical Center, Seoul, 05505, South Korea
| | - Seonae Roh
- Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, South Korea
| | - Ji-Yoon Lee
- Asan Institute for Life Sciences, Asan Medical Center and Department of Convergence Medicine, College of Medicine, University of Ulsan, Seoul, 05505, South Korea
| | - Soo Jin Oh
- Asan Institute for Life Sciences, Asan Medical Center and Department of Convergence Medicine, College of Medicine, University of Ulsan, Seoul, 05505, South Korea
| | - Young Hoon Sung
- Department of Medical Science, Asan Medical Institute of Convergence Science and Technology (AMIST), University of Ulsan College of Medicine, Asan Medical Center, Seoul, 05505, South Korea
- Convergence Medicine Research Center, Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, South Korea
| | - Yeonmi Lee
- Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, South Korea
- Present Address: Center for Embryo & Stem Cell Research, CHA Advanced Research Institute and Department of Biomedical Science, CHA University, Pocheon-si, Gyeonggi, 13488, South Korea
| | - Sung Hoon Kim
- Department of Obstetrics and Gynecology, University of Ulsan College of Medicine, Asan Medical Center, Seoul, 05505, South Korea.
| | - Eunju Kang
- Department of Medical Science, Asan Medical Institute of Convergence Science and Technology (AMIST), University of Ulsan College of Medicine, Asan Medical Center, Seoul, 05505, South Korea.
- Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, South Korea.
- Present Address: Center for Embryo & Stem Cell Research, CHA Advanced Research Institute and Department of Biomedical Science, CHA University, Pocheon-si, Gyeonggi, 13488, South Korea.
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Pan M, Kohlbauer V, Blancke Soares A, Schinke H, Huang Y, Kranz G, Quadt T, Hachmeister M, Gires O. Interactome analysis reveals endocytosis and membrane recycling of EpCAM during differentiation of embryonic stem cells and carcinoma cells. iScience 2021; 24:103179. [PMID: 34693227 PMCID: PMC8517208 DOI: 10.1016/j.isci.2021.103179] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2021] [Revised: 08/16/2021] [Accepted: 09/23/2021] [Indexed: 12/16/2022] Open
Abstract
Transmembrane epithelial cell adhesion molecule (EpCAM) is expressed in epithelia, carcinoma, teratoma, and embryonic stem cells (ESCs). EpCAM displays spatiotemporal patterning during embryogenesis, tissue morphogenesis, cell differentiation, and epithelial-to-mesenchymal transition (EMT) in carcinomas. Potential interactors of EpCAM were identified in murine F9 teratoma cells using a stable isotope labeling with amino acids in cell culture-based proteomic approach (n = 77, enrichment factor >3, p value ≤ 0.05). Kyoto Encyclopedia of Genes and Genomes and gene ontology terms revealed interactions with regulators of endosomal trafficking and membrane recycling, which were further validated for Rab5, Rab7, and Rab11. Endocytosis and membrane recycling of EpCAM were confirmed in mF9 cells, E14TG2α ESC, and Kyse30 carcinoma cells. Reduction of EpCAM during mesodermal differentiation and TGFβ-induced EMT correlated with enhanced endocytosis and block or reduction of recycling in ESCs and esophageal carcinoma cells. Hence, endocytosis and membrane recycling are means of regulation of EpCAM protein levels during differentiation of ESC and EMT induction in carcinoma cells.
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Affiliation(s)
- Min Pan
- Department of Otorhinolaryngology, The First Affiliated Hospital of Chongqing Medical University, Yuzhong District, Chongqing, China
| | - Vera Kohlbauer
- Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital, LMU Munich, Munich, Germany
| | - Alexandra Blancke Soares
- Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital, LMU Munich, Munich, Germany
| | - Henrik Schinke
- Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital, LMU Munich, Munich, Germany
| | - Yuanchi Huang
- Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital, LMU Munich, Munich, Germany
| | - Gisela Kranz
- Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital, LMU Munich, Munich, Germany
| | - Tanja Quadt
- Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital, LMU Munich, Munich, Germany
| | - Matthias Hachmeister
- Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital, LMU Munich, Munich, Germany
| | - Olivier Gires
- Department of Otorhinolaryngology, Head and Neck Surgery, University Hospital, LMU Munich, Munich, Germany.,Clinical Cooperation Group "Personalized Radiotherapy in Head and Neck Cancer", Helmholtz Zentrum München, Neuherberg, Germany
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Abstract
The epithelial cell adhesion molecule (EpCAM) is involved in oncogenesis of hepatoblastomas (HBs). Prior genomic profiling studies showed higher EpCAM expression and worse prognosis in HBs containing primitive histotypes, however, this has not been fully addressed from an immunohistochemical perspective. Our goal is to characterize differential EpCAM immunohistochemistry (EpCAM-IHC) among HBs histotypes. We retrieved 62 HBs from 52 patients. EpCAM-IHC was performed (anti-MOC-31, 1:50 dilution; Cell Marque Corporation, Rocklin, CA) and graded in histotypes using the immunoreactive score. The median age of patients was 2 years (range: 0.4 to 9 y) with a M:F ratio of 1.9. Outcome information was available in 38 patients (alive=30, alive with disease=3, and deceased=5) with median follow-up of 60 months (range: 2 to 171 mo). EpCAM-IHC showed notable overexpression (immunoreactive score >4) in embryonal (89%) and crowded fetal (74%) in contrast to glandular (33%), well-differentiated fetal (32%), and small cell undifferentiated/blastemal (3%) components. Mesenchymal elements were negative. In summary, EpCAM-IHC is helpful to distinguish between epithelial components as it is progressively lost in the transition from embryonal to crowded fetal and into well-differentiated fetal histotypes. Its preferential expression among primitive HBs might have therapeutic and prognostic implications. The significance of its largely negative expression in small cell undifferentiated/blastema is interesting despite its presumed immaturity, deserving further studies.
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Xu T, Vorobyeva A, Schulga A, Konovalova E, Vorontsova O, Ding H, Gräslund T, Tashireva LA, Orlova A, Tolmachev V, Deyev SM. Imaging-Guided Therapy Simultaneously Targeting HER2 and EpCAM with Trastuzumab and EpCAM-Directed Toxin Provides Additive Effect in Ovarian Cancer Model. Cancers (Basel) 2021; 13:3939. [PMID: 34439094 PMCID: PMC8393281 DOI: 10.3390/cancers13163939] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2021] [Revised: 07/23/2021] [Accepted: 08/02/2021] [Indexed: 12/29/2022] Open
Abstract
Efficient treatment of disseminated ovarian cancer (OC) is challenging due to its heterogeneity and chemoresistance. Overexpression of human epidermal growth factor receptor 2 (HER2) and epithelial cell adhesion molecule (EpCAM) in approx. 30% and 70% of ovarian cancers, respectively, allows for co-targeted treatment. The clinical efficacy of the monoclonal antibody trastuzumab in patients with HER2-positive breast, gastric and gastroesophageal cancers makes it readily available as the HER2-targeting component. As the EpCAM-targeting component, we investigated the designed ankyrin repeat protein (DARPin) Ec1 fused to a truncated variant of Pseudomonas exotoxin A with reduced immunogenicity and low general toxicity (LoPE). Ec1-LoPE was radiolabeled, evaluated in ovarian cancer cells in vitro and its biodistribution and tumor-targeting properties were studied in vivo. The therapeutic efficacy of Ec1-LoPE alone and in combination with trastuzumab was studied in mice bearing EpCAM- and HER2-expressing SKOV3 xenografts. SPECT/CT imaging enabled visualization of EpCAM and HER2 expression in the tumors. Co-treatment using Ec1-LoPE and trastuzumab was more effective at reducing tumor growth and prolonged the median survival of mice compared with mice in the control and monotherapy groups. Repeated administration of Ec1-LoPE was well tolerated without signs of hepatic or kidney toxicity. Co-treatment with trastuzumab and Ec1-LoPE might be a potential therapeutic strategy for HER2- and EpCAM-positive OC.
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Affiliation(s)
- Tianqi Xu
- Department of Immunology, Genetics and Pathology, Uppsala University, 751 85 Uppsala, Sweden; (T.X.); (O.V.)
| | - Anzhelika Vorobyeva
- Department of Immunology, Genetics and Pathology, Uppsala University, 751 85 Uppsala, Sweden; (T.X.); (O.V.)
- Research Centrum for Oncotheranostics, Research School of Chemistry and Applied Biomedical Sciences, National Research Tomsk Polytechnic University, 634 050 Tomsk, Russia; (A.S.); (A.O.); (S.M.D.)
| | - Alexey Schulga
- Research Centrum for Oncotheranostics, Research School of Chemistry and Applied Biomedical Sciences, National Research Tomsk Polytechnic University, 634 050 Tomsk, Russia; (A.S.); (A.O.); (S.M.D.)
- Molecular Immunology Laboratory, Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
| | - Elena Konovalova
- Molecular Immunology Laboratory, Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
| | - Olga Vorontsova
- Department of Immunology, Genetics and Pathology, Uppsala University, 751 85 Uppsala, Sweden; (T.X.); (O.V.)
| | - Haozhong Ding
- Department of Protein Science, KTH Royal Institute of Technology, Roslagstullsbacken 21, 114 17 Stockholm, Sweden; (H.D.); (T.G.)
| | - Torbjörn Gräslund
- Department of Protein Science, KTH Royal Institute of Technology, Roslagstullsbacken 21, 114 17 Stockholm, Sweden; (H.D.); (T.G.)
| | - Liubov A. Tashireva
- Cancer Research Institute, Tomsk National Research Medical Center Russian Academy of Sciences, 634009 Tomsk, Russia;
| | - Anna Orlova
- Research Centrum for Oncotheranostics, Research School of Chemistry and Applied Biomedical Sciences, National Research Tomsk Polytechnic University, 634 050 Tomsk, Russia; (A.S.); (A.O.); (S.M.D.)
- Department of Medicinal Chemistry, Uppsala University, 751 23 Uppsala, Sweden
- Science for Life Laboratory, Uppsala University, 751 23 Uppsala, Sweden
| | - Vladimir Tolmachev
- Department of Immunology, Genetics and Pathology, Uppsala University, 751 85 Uppsala, Sweden; (T.X.); (O.V.)
- Research Centrum for Oncotheranostics, Research School of Chemistry and Applied Biomedical Sciences, National Research Tomsk Polytechnic University, 634 050 Tomsk, Russia; (A.S.); (A.O.); (S.M.D.)
| | - Sergey M. Deyev
- Research Centrum for Oncotheranostics, Research School of Chemistry and Applied Biomedical Sciences, National Research Tomsk Polytechnic University, 634 050 Tomsk, Russia; (A.S.); (A.O.); (S.M.D.)
- Molecular Immunology Laboratory, Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
- Bio-Nanophotonic Lab, Institute of Engineering Physics for Biomedicine (PhysBio), National Research Nuclear University ‘MEPhI’, 115409 Moscow, Russia
- Center of Biomedical Engineering, Sechenov University, 119991 Moscow, Russia
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Chen S, Goldsmith JD, Fawaz R, Al-Ibraheemi A, Perez-Atayde AR, Vargas SO. Liver Pathology, Including MOC31 Immunohistochemistry, in Congenital Tufting Enteropathy. Am J Surg Pathol 2021; 45:1091-1097. [PMID: 33756496 DOI: 10.1097/pas.0000000000001710] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
Abstract
Congenital tufting enteropathy (CTE) is a rare heritable cause of intractable diarrhea due to EPCAM mutation. Pathologic findings include intestinal villous atrophy, tufted discohesive tear-drop-shaped epithelium, and a normal brush border. In affected patients, absent intestinal epithelial cell adhesion molecule (EpCAM) expression results in loss of MOC31 immunostaining. CTE liver pathology has not yet been described. We identified CTE patients with liver biopsies and reviewed clinicopathologic material including MOC31 immunohistochemistry. Three CTE patients had 4 liver core biopsies (at ages 1, 5, 7, and 16 y), 2 for preintestinal transplant evaluation, and 2 (from a single patient) for pretreatment assessment of chronic hepatitis C; all had received parenteral nutrition (PN). All samples showed loss of biliary epithelial polarization and mild portal and lobular inflammation. Only the hepatitis C patient demonstrated fibrosis. One patient each had lobular neutrophilic microabscesses and macrovesicular steatosis. Proliferative ductular reactions were absent in CTE patients but present in all controls on PN for other reasons. MOC31 was absent in biliary epithelium and hepatocytes of all CTE patients; controls showed consistent strong membranous biliary epithelial and patchy membranous periportal hepatocyte staining. Our data show that, histologically, hepatopathy in CTE can be difficult to separate from comorbid disease including PN effect; however, the absent ductular reaction may be characteristic. MOC31 localization in the biliary epithelium and zone 1 hepatocytes of controls suggests these compartments of the liver might be most susceptible to effects of EpCAM deficiency. In addition, we validate the liver as suitable tissue for CTE diagnosis using MOC31 immunohistochemistry.
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Affiliation(s)
| | | | - Rima Fawaz
- Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Boston Children's Hospital, Boston, MA
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Wang Z, Kang B, Gao Q, Huang L, Di J, Fan Y, Yu J, Jiang B, Gao F, Wang D, Sun H, Gu Y, Li J, Su X. Quadruple-editing of the MAPK and PI3K pathways effectively blocks the progression of KRAS-mutated colorectal cancer cells. Cancer Sci 2021; 112:3895-3910. [PMID: 34185934 PMCID: PMC8409416 DOI: 10.1111/cas.15049] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2021] [Revised: 06/15/2021] [Accepted: 06/21/2021] [Indexed: 02/06/2023] Open
Abstract
Mutated KRAS promotes the activation of the MAPK pathway and the progression of colorectal cancer (CRC) cells. Aberrant activation of the PI3K pathway strongly attenuates the efficacy of MAPK suppression in KRAS‐mutated CRC. The development of a novel strategy targeting a dual pathway is therefore highly essential for the therapy of KRAS‐mutated CRC. In this study, a quadruple‐depleting system for the KRAS, MEK1, PIK3CA, and MTOR genes based on CRISPR/SaCas9 was developed. Adenovirus serotype 5 (ADV5) was integrated with two engineered proteins, an adaptor and a protector, to form ADV‐protein complex (APC) for systemic delivery of the CRISPR system. Quadruple‐editing could significantly inhibit the MAPK and PI3K pathways in CRC cells with oncogenic mutations of KRAS and PIK3CA or with KRAS mutation and compensated PI3K activation. Compared with MEK and PI3K/MTOR inhibitors, quadruple‐editing induced more significant survival inhibition on primary CRC cells with oncogenic mutations of KRAS and PIK3CA. The adaptor specifically targeting EpCAM and the hexon‐shielding protector could dramatically enhance ADV5 infection efficiency to CRC cells and significantly reduce off‐targeting tropisms to many organs except the colon. Moreover, quadruple‐editing intravenously delivered by APC significantly blocked the dual pathway and tumor growth of KRAS‐mutated CRC cells, without influencing normal tissues in cell‐ and patient‐derived xenograft models. Therefore, APC‐delivered quadruple‐editing of the MAPK and PI3K pathways shows a promising therapeutic potential for KRAS‐mutated CRC.
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Affiliation(s)
- Zaozao Wang
- Department of Gastrointestinal Surgery IV, Key Laboratory of Carcinogenesis and Translational Research, Peking University Cancer Hospital and Institute, Beijing, China
| | | | | | | | - Jiabo Di
- Department of Gastrointestinal Surgery IV, Key Laboratory of Carcinogenesis and Translational Research, Peking University Cancer Hospital and Institute, Beijing, China
| | - Yingcong Fan
- Department of Gastrointestinal Surgery IV, Key Laboratory of Carcinogenesis and Translational Research, Peking University Cancer Hospital and Institute, Beijing, China
| | - Jianhong Yu
- Department of Gastrointestinal Surgery IV, Key Laboratory of Carcinogenesis and Translational Research, Peking University Cancer Hospital and Institute, Beijing, China
| | - Beihai Jiang
- Department of Gastrointestinal Surgery IV, Key Laboratory of Carcinogenesis and Translational Research, Peking University Cancer Hospital and Institute, Beijing, China
| | | | | | | | - Ying Gu
- BGI-Shenzhen, Shenzhen, China
| | - Jian Li
- Department of Gastrointestinal Oncology, Key Laboratory of Carcinogenesis and Translational Research, Peking University Cancer Hospital and Institute, Beijing, China
| | - Xiangqian Su
- Department of Gastrointestinal Surgery IV, Key Laboratory of Carcinogenesis and Translational Research, Peking University Cancer Hospital and Institute, Beijing, China
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Cho CS, Xi J, Si Y, Park SR, Hsu JE, Kim M, Jun G, Kang HM, Lee JH. Microscopic examination of spatial transcriptome using Seq-Scope. Cell 2021; 184:3559-3572.e22. [PMID: 34115981 PMCID: PMC8238917 DOI: 10.1016/j.cell.2021.05.010] [Citation(s) in RCA: 288] [Impact Index Per Article: 72.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2021] [Revised: 03/29/2021] [Accepted: 05/07/2021] [Indexed: 12/18/2022]
Abstract
Spatial barcoding technologies have the potential to reveal histological details of transcriptomic profiles; however, they are currently limited by their low resolution. Here, we report Seq-Scope, a spatial barcoding technology with a resolution comparable to an optical microscope. Seq-Scope is based on a solid-phase amplification of randomly barcoded single-molecule oligonucleotides using an Illumina sequencing platform. The resulting clusters annotated with spatial coordinates are processed to expose RNA-capture moiety. These RNA-capturing barcoded clusters define the pixels of Seq-Scope that are ∼0.5-0.8 μm apart from each other. From tissue sections, Seq-Scope visualizes spatial transcriptome heterogeneity at multiple histological scales, including tissue zonation according to the portal-central (liver), crypt-surface (colon) and inflammation-fibrosis (injured liver) axes, cellular components including single-cell types and subtypes, and subcellular architectures of nucleus and cytoplasm. Seq-Scope is quick, straightforward, precise, and easy-to-implement and makes spatial single-cell analysis accessible to a wide group of biomedical researchers.
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Affiliation(s)
- Chun-Seok Cho
- Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Jingyue Xi
- Department of Biostatistics, University of Michigan School of Public Health, Ann Arbor, MI 48109, USA
| | - Yichen Si
- Department of Biostatistics, University of Michigan School of Public Health, Ann Arbor, MI 48109, USA
| | - Sung-Rye Park
- Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Jer-En Hsu
- Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Myungjin Kim
- Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Goo Jun
- Human Genetics Center, University of Texas Health Science Center at Houston, Houston, TX 77030, USA
| | - Hyun Min Kang
- Department of Biostatistics, University of Michigan School of Public Health, Ann Arbor, MI 48109, USA
| | - Jun Hee Lee
- Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
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Zhu X, Zhang B, He Y, Bao J. Liver Organoids: Formation Strategies and Biomedical Applications. Tissue Eng Regen Med 2021; 18:573-585. [PMID: 34132985 DOI: 10.1007/s13770-021-00357-w] [Citation(s) in RCA: 32] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2020] [Revised: 05/20/2021] [Accepted: 05/22/2021] [Indexed: 02/05/2023] Open
Abstract
The liver is the most important digestive organ in the body. Several studies have explored liver biology and diseases related to the liver. However, most of these studies have only explored liver development, mechanism of liver regeneration and pathophysiology of liver diseases mainly based on two-dimensional (2D) cell lines and animal models. Traditional 2D cell lines do not represent the complex three-dimensional tissue architecture whereas animal models are limited by inter-species differences. These shortcomings limit understanding of liver biology and diseases. Liver organoid technology is effective in elucidating structural and physiological characteristics and basic tissue-level functions of liver tissue. In this review, formation strategies and a wide range of applications in biomedicine of liver organoid are summarized. Liver organoids are derived from single type cell culture, such as induced pluripotent stem cells (iPSCs), adult stem cells, primary hepatocytes, and primary cholangiocytes and multi-type cells co-culture, such as iPSC-derived hepatic endoderm cells co-cultured with mesenchymal stem cells and umbilical cord-derived endothelial cells. In vitro studies report that liver organoids are a promising model for regenerative medicine, organogenesis, liver regeneration, disease modelling, drug screening and personalized treatment. Liver organoids are a promising in vitro model for basic research and for development of clinical therapeutic interventions for hepatopathy.
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Affiliation(s)
- Xinglong Zhu
- Institute of Clinical Pathology, Key Laboratory of Transplant Engineering and Immunology, NHC, West China Hospital, Sichuan University, No. 37, Guoxue Alley, Wuhou District, Chengdu, 610041, Sichuan Province, China
| | - Bingqi Zhang
- Institute of Clinical Pathology, Key Laboratory of Transplant Engineering and Immunology, NHC, West China Hospital, Sichuan University, No. 37, Guoxue Alley, Wuhou District, Chengdu, 610041, Sichuan Province, China
| | - Yuting He
- Institute of Clinical Pathology, Key Laboratory of Transplant Engineering and Immunology, NHC, West China Hospital, Sichuan University, No. 37, Guoxue Alley, Wuhou District, Chengdu, 610041, Sichuan Province, China
| | - Ji Bao
- Institute of Clinical Pathology, Key Laboratory of Transplant Engineering and Immunology, NHC, West China Hospital, Sichuan University, No. 37, Guoxue Alley, Wuhou District, Chengdu, 610041, Sichuan Province, China.
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Bellanti F, di Bello G, Iannelli G, Pannone G, Pedicillo MC, Boulter L, Lu WY, Tamborra R, Villani R, Vendemiale G, Forbes SJ, Serviddio G. Inhibition of nuclear factor (erythroid-derived 2)-like 2 promotes hepatic progenitor cell activation and differentiation. NPJ Regen Med 2021; 6:28. [PMID: 34039998 PMCID: PMC8155039 DOI: 10.1038/s41536-021-00137-z] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2020] [Accepted: 04/28/2021] [Indexed: 02/08/2023] Open
Abstract
The stem cell ability to self-renew and lead regeneration relies on the balance of complex signals in their microenvironment. The identification of modulators of hepatic progenitor cell (HPC) activation is determinant for liver regeneration and may improve cell transplantation for end-stage liver disease. This investigation used different models to point out the Nuclear factor (erythroid-derived 2)-like 2 (NRF2) as a key regulator of the HPC fate. We initially proved that in vivo models of biliary epithelial cells (BECs)/HPC activation show hepatic oxidative stress, which activates primary BECs/HPCs in vitro. NRF2 downregulation and silencing were associated with morphological, phenotypic, and functional modifications distinctive of differentiated cells. Furthermore, NRF2 activation in the biliary tract repressed the ductular reaction in injured liver. To definitely assess the importance of NRF2 in HPC biology, we applied a xenograft model by inhibiting NRF2 in the human derived HepaRG cell line and transplanting into SCID/beige mice administered with anti-Fas antibody to induce hepatocellular apoptosis; this resulted in effective human hepatocyte repopulation with reduced liver injury. To conclude, NRF2 inhibition leads to the activation and differentiation of liver progenitors. This redox-dependent transcription factor represents a potential target to regulate the commitment of undifferentiated hepatic progenitors into specific lineages.
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Affiliation(s)
- Francesco Bellanti
- Centre for Experimental and Regenerative Medicine, Department of Medical and Surgical Sciences, University of Foggia, Foggia, Italy.
| | - Giorgia di Bello
- Centre for Experimental and Regenerative Medicine, Department of Medical and Surgical Sciences, University of Foggia, Foggia, Italy
| | - Giuseppina Iannelli
- Centre for Experimental and Regenerative Medicine, Department of Medical and Surgical Sciences, University of Foggia, Foggia, Italy
| | - Giuseppe Pannone
- Anatomical Pathology Unit, Department of Clinical and Experimental Medicine, University of Foggia, Foggia, Italy
| | - Maria Carmela Pedicillo
- Anatomical Pathology Unit, Department of Clinical and Experimental Medicine, University of Foggia, Foggia, Italy
| | - Luke Boulter
- MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh, UK
| | - Wei-Yu Lu
- Centre for Liver and Gastrointestinal Research, Institute of Immunology and Immunotherapy, University of Birmingham, Edgbaston Birmingham, UK
| | - Rosanna Tamborra
- Centre for Experimental and Regenerative Medicine, Department of Medical and Surgical Sciences, University of Foggia, Foggia, Italy
| | - Rosanna Villani
- Centre for Experimental and Regenerative Medicine, Department of Medical and Surgical Sciences, University of Foggia, Foggia, Italy
| | - Gianluigi Vendemiale
- Centre for Experimental and Regenerative Medicine, Department of Medical and Surgical Sciences, University of Foggia, Foggia, Italy
| | - Stuart J Forbes
- MRC Centre for Regenerative Medicine, University of Edinburgh, Edinburgh, UK
| | - Gaetano Serviddio
- Centre for Experimental and Regenerative Medicine, Department of Medical and Surgical Sciences, University of Foggia, Foggia, Italy
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The Molecular Basis of Different Approaches for the Study of Cancer Stem Cells and the Advantages and Disadvantages of a Three-Dimensional Culture. Molecules 2021; 26:molecules26092615. [PMID: 33947095 PMCID: PMC8124970 DOI: 10.3390/molecules26092615] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2021] [Revised: 04/13/2021] [Accepted: 04/26/2021] [Indexed: 12/12/2022] Open
Abstract
Cancer stem cells (CSCs) are a rare tumor subpopulation with high differentiation, proliferative and tumorigenic potential compared to the remaining tumor population. CSCs were first discovered by Bonnet and Dick in 1997 in acute myeloid leukemia. The identification and isolation of these cells in this pioneering study were carried out through the flow cytometry, exploiting the presence of specific cell surface molecular markers (CD34+/CD38−). In the following years, different strategies and projects have been developed for the study of CSCs, which are basically divided into surface markers assays and functional assays; some of these techniques also allow working with a cellular model that better mimics the tumor architecture. The purpose of this mini review is to summarize and briefly describe all the current methods used for the identification, isolation and enrichment of CSCs, describing, where possible, the molecular basis, the advantages and disadvantages of each technique with a particular focus on those that offer a three-dimensional culture.
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Abstract
Aim of the study CD326 has been used as a single marker to enrich for hepatic stem cell populations in the liver. However, bile duct epithelium is also positive for CD326, which impedes the selection of pure hepatic stem cell populations. Some markers have been proposed to be co-expressed by hepatic stem cells but these have not been systematically compared. Therefore, we determined the percentages and compared the characteristics of human liver cells expressing potential stem cell surface markers. Material and methods We analyzed CD326 expression in human liver tissues from fetal, neonatal, pediatric, and adult stages using immunohistochemistry. In flow cytometry, we quantified fetal liver cells for their co-expression of CD326 with CD56, CD117, CD44, CD90, CD49f, LGR5 and SSEA4. We analyzed the various fractions for their quantitative expression of genes typically associated with progenitors and hepatic lineages. Results 12.5% of cells were positive for CD326; of these, 63.5% co-expressed CD44. The lowest co-expression percentages were for SSEA4 (2.1%) and LGR5 (0.7%). Fractions revealed distinct gene expression patterns. Of all combinations, cells that co-expressed surface CD326 and SSEA4 demonstrated the highest gene expression for the proliferation marker MKi67 and hepatic markers DLK1, AFP and ALB, and were the only fraction negative for the biliary epithelial marker KRT19. Histology of adult and fetal liver showed cells positive for CD326 and SSEA4 but negative for CK19. Conclusions CD326-positive cells represent a heterogeneous population, which in combination with SSEA4 potentially distinguishes bile duct epithelium from hepatic stem cells. These findings can help to further classify human hepatic progenitor stages.
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Kong C, Bobe S, Pilger C, Lachetta M, Øie CI, Kirschnick N, Mönkemöller V, Hübner W, Förster C, Schüttpelz M, Kiefer F, Huser T, Schulte Am Esch J. Multiscale and Multimodal Optical Imaging of the Ultrastructure of Human Liver Biopsies. Front Physiol 2021; 12:637136. [PMID: 33679449 PMCID: PMC7925637 DOI: 10.3389/fphys.2021.637136] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/04/2020] [Accepted: 01/27/2021] [Indexed: 12/30/2022] Open
Abstract
The liver as the largest organ in the human body is composed of a complex macroscopic and microscopic architecture that supports its indispensable function to maintain physiological homeostasis. Optical imaging of the human liver is particularly challenging because of the need to cover length scales across 7 orders of magnitude (from the centimeter scale to the nanometer scale) in order to fully assess the ultrastructure of the entire organ down to the subcellular scale and probe its physiological function. This task becomes even more challenging the deeper within the organ one hopes to image, because of the strong absorption and scattering of visible light by the liver. Here, we demonstrate how optical imaging methods utilizing highly specific fluorescent labels, as well as label-free optical methods can seamlessly cover this entire size range in excised, fixed human liver tissue and we exemplify this by reconstructing the biliary tree in three-dimensional space. Imaging of tissue beyond approximately 0.5 mm length requires optical clearing of the human liver. We present the successful use of optical projection tomography and light-sheet fluorescence microscopy to derive information about the liver architecture on the millimeter scale. The intermediate size range is covered using label-free structural and chemically sensitive methods, such as second harmonic generation and coherent anti-Stokes Raman scattering microscopy. Laser-scanning confocal microscopy extends the resolution to the nanoscale, allowing us to ultimately image individual liver sinusoidal endothelial cells and their fenestrations by super-resolution structured illumination microscopy. This allowed us to visualize the human hepatobiliary system in 3D down to the cellular level, which indicates that reticular biliary networks communicate with portal bile ducts via single or a few ductuli. Non-linear optical microscopy enabled us to identify fibrotic regions extending from the portal field to the parenchyma, along with microvesicular steatosis in liver biopsies from an older patient. Lastly, super-resolution microscopy allowed us to visualize and determine the size distribution of fenestrations in human liver sinusoidal endothelial cells for the first time under aqueous conditions. Thus, this proof-of-concept study allows us to demonstrate, how, in combination, these techniques open up a new chapter in liver biopsy analysis.
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Affiliation(s)
- Cihang Kong
- Department of Physics, Bielefeld University, Bielefeld, Germany
| | - Stefanie Bobe
- European Institute for Molecular Imaging, University of Münster, Münster, Germany
| | | | - Mario Lachetta
- Department of Physics, Bielefeld University, Bielefeld, Germany
| | - Cristina Ionica Øie
- Vascular Biology Research Group, Department of Medical Biology, University of Tromsø - The Arctic University of Norway, Tromsø, Norway
| | - Nils Kirschnick
- European Institute for Molecular Imaging, University of Münster, Münster, Germany
| | | | - Wolfgang Hübner
- Department of Physics, Bielefeld University, Bielefeld, Germany.,Forschungsverbund BioMedizin Bielefeld (FBMB), Bielefeld, Germany
| | | | - Mark Schüttpelz
- Department of Physics, Bielefeld University, Bielefeld, Germany
| | - Friedemann Kiefer
- European Institute for Molecular Imaging, University of Münster, Münster, Germany
| | - Thomas Huser
- Department of Physics, Bielefeld University, Bielefeld, Germany.,Forschungsverbund BioMedizin Bielefeld (FBMB), Bielefeld, Germany
| | - Jan Schulte Am Esch
- Forschungsverbund BioMedizin Bielefeld (FBMB), Bielefeld, Germany.,Department of General and Visceral Surgery, Evangelisches Klinikum Bethel gGmbH, University Hospital OWL of the University of Bielefeld, Bielefeld, Germany
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Wang Z, Faria J, Penning LC, Masereeuw R, Spee B. Tissue-Engineered Bile Ducts for Disease Modeling and Therapy. Tissue Eng Part C Methods 2021; 27:59-76. [PMID: 33267737 DOI: 10.1089/ten.tec.2020.0283] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Recent biotechnical advances in the in vitro culture of cholangiocytes and generation of bioengineered biliary tissue have a high potential for creating biliary tissue to be used for disease modeling, drug screening, and transplantation. For the past few decades, scientists have searched for a source of cholangiocytes, focused on primary cholangiocytes or cholangiocytes derived from hepatocytes or stem cells. At the same time, the development of scaffolds for biliary tissue engineering for transplantation and modeling of cholangiopathies has been explored. In this review, we provide an overview on the current understanding of cholangiocytes sources, the effect of signaling molecules, and transcription factors on cell differentiation, along with the effects of extracellular matrix molecules and scaffolds on bioengineered biliary tissues, and their application in disease modeling and drug screening. Impact statement Over the past few decades, biliary tissue engineering has acquired significant attention, but currently a number of factors hinder this field to eventually generate bioengineered bile ducts that mimic in vivo physiology and are suitable for transplantation. In this review, we present the latest advances with respect to cell source selection, influence of growth factors and scaffolds, and functional characterization, as well as applications in cholangiopathy modeling and drug screening. This review is suited for a broad spectrum of readers, including fundamental liver researchers and clinicians with interest in the current state and application of bile duct engineering and disease modeling.
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Affiliation(s)
- Zhenguo Wang
- Department of Clinical Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.,Division of Pharmacology, Utrecht Institute for Pharmaceutical Sciences, Faculty of Science, Utrecht University, Utrecht, The Netherlands
| | - João Faria
- Division of Pharmacology, Utrecht Institute for Pharmaceutical Sciences, Faculty of Science, Utrecht University, Utrecht, The Netherlands
| | - Louis C Penning
- Department of Clinical Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands
| | - Rosalinde Masereeuw
- Division of Pharmacology, Utrecht Institute for Pharmaceutical Sciences, Faculty of Science, Utrecht University, Utrecht, The Netherlands
| | - Bart Spee
- Department of Clinical Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands
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Kulkeaw K, Tubsuwan A, Tongkrajang N, Whangviboonkij N. Generation of human liver organoids from pluripotent stem cell-derived hepatic endoderms. PeerJ 2020; 8:e9968. [PMID: 33133779 PMCID: PMC7580584 DOI: 10.7717/peerj.9968] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2020] [Accepted: 08/26/2020] [Indexed: 12/18/2022] Open
Abstract
Background The use of a personalized liver organoid derived from human-induced pluripotent stem cells (HuiPSCs) is advancing the use of in vitro disease models for the design of specific, effective therapies for individuals. Collecting patient peripheral blood cells for HuiPSC generation is preferable because it is less invasive; however, the capability of blood cell-derived HuiPSCs for hepatic differentiation and liver organoid formation remains uncertain. Moreover, the currently available methods for liver organoid formation require a multistep process of cell differentiation or a combination of hepatic endodermal, endothelial and mesenchymal cells, which is a major hurdle for the application of personalized liver organoids in high-throughput testing of drug toxicity and safety. To demonstrate the capability of blood cell-derived HuiPSCs for liver organoid formation without support from endothelial and mesenchymal cells. Methods The peripheral blood-derived HuiPSCs first differentiated into hepatic endoderm (HE) in two-dimensional (2D) culture on Matrigel-coated plates under hypoxia for 10 days. The HE was then collected and cultured in 3D culture using 50% Matrigel under ambient oxygen. The maturation of hepatocytes was further induced by adding hepatocyte growth medium containing HGF and oncostatin M on top of the 3D culture and incubating the culture for an additional 12–17 days. The function of the liver organoids was assessed using expression analysis of hepatocyte-specific gene and proteins. Albumin (ALB) synthesis, glycogen and lipid storage, and metabolism of indocyanine were evaluated. The spatial distribution of albumin was examined using immunofluorescence and confocal microscopy. Results CD34+ hematopoietic cell-derived HuiPSCs were capable of differentiating into definitive endoderm expressing SOX17 and FOXA2, hepatic endoderm expressing FOXA2, hepatoblasts expressing AFP and hepatocytes expressing ALB. On day 25 of the 2D culture, cells expressed SOX17, FOXA2, AFP and ALB, indicating the presence of cellular heterogeneity. In contrast, the hepatic endoderm spontaneously formed a spherical, hollow structure in a 3D culture of 50% Matrigel, whereas hepatoblasts and hepatocytes could not form. Microscopic observation showed a single layer of polygonal-shaped cells arranged in a 3D structure. The hepatic endoderm-derived organoid synthesis ALB at a higher level than the 2D culture but did not express definitive endoderm-specific SOX17, indicating the greater maturity of the hepatocytes in the liver organoids. Confocal microscopic images and quantitative ELISA confirmed albumin synthesis in the cytoplasm of the liver organoid and its secretion. Overall, 3D culture of the hepatic endoderm is a relatively fast, simple, and less laborious way to generate liver organoids from HuiPSCs that is more physiologically relevant than 2D culture.
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Affiliation(s)
- Kasem Kulkeaw
- Department of Parasitology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Alisa Tubsuwan
- Stem Cell Research Group, Institute of Molecular Biosciences, Mahidol University, Nakhon Pathom, Thailand
| | - Nongnat Tongkrajang
- Department of Parasitology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Narisara Whangviboonkij
- Department of Parasitology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
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Defining Adult Stem Cell Function at Its Simplest: The Ability to Replace Lost Cells through Mitosis. Cell Stem Cell 2020; 25:174-183. [PMID: 31374197 DOI: 10.1016/j.stem.2019.07.002] [Citation(s) in RCA: 112] [Impact Index Per Article: 22.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Classic studies on hematopoiesis indicate that blood cell numbers are maintained by rare, hard-wired, transplantable stem cells (SCs). Subsequent studies in other organs have implicitly assumed that all SC hierarchies follow the design of the hematopoietic system. Lineage tracing techniques have revolutionized the study of solid tissue SCs. It thus appears that key characteristics of the hematopoietic SC hierarchy (rarity of SCs, specific marker expression, quiescence, asymmetric division, and unidirectional differentiation) are not generalizable to other tissues. In light of these insights, we offer a revised, generalizable definition of SC function: the ability to replace lost tissue through cell division.
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Mao S, He J, Zhao Y, Liu T, Xie F, Yang H, Mao Y, Pang Y, Sun W. Bioprinting of patient-derived in vitro intrahepatic cholangiocarcinoma tumor model: establishment, evaluation and anti-cancer drug testing. Biofabrication 2020; 12:045014. [PMID: 32599574 DOI: 10.1088/1758-5090/aba0c3] [Citation(s) in RCA: 55] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Towards the development of in vivo-mimicking tumor model for extensive study of tumorigenesis and establishment of personalized therapy, patient-derived primary tumor cells were employed in this work for three-dimensional (3D) bioprinting. Intrahepatic cholangiocarcinoma cells isolated from patient were bioprinted using a composite hydrogel system of gelatin-alginate-MatrigelTM into pre-designed grid architecture. ICC cells were observed to process a colony forming ability with high survival rate and active proliferation. Expression levels of tumor markers, cancer stem cell markers, matrix metalloproteinase protein, index of tumor fibrosis, index of liver function, and epithelial-mesenchymal transition regulatory proteins confirmed the development of the invasive and metastatic phenotype of the intrahepatic cholangiocarcinoma cells in the 3D printed tumor microenvironment. Similar results were obtained in anti-cancer drug resistance of the intrahepatic cholangiocarcinoma cells in the 3D bioprinted construct that demonstrated stem-like properties, which suggested the promising potential of current 3D printed tumor model in the development of personalized therapy, especially for discovery of more conducive targeted drugs.
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Affiliation(s)
- Shuangshuang Mao
- Department of Mechanical Engineering, Biomanufacturing Center, Tsinghua University, Beijing, People's Republic of China. Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing, Tsinghua University, Beijing, People's Republic of China. 'Biomanufacturing and Engineering Living Systems' 111 - Innovation International Talents Base, Beijing, People's Republic of China
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Study on the Selection of the Targets of Esophageal Carcinoma and Interventions of Ginsenosides Based on Network Pharmacology and Bioinformatics. EVIDENCE-BASED COMPLEMENTARY AND ALTERNATIVE MEDICINE 2020; 2020:4821056. [PMID: 32714406 PMCID: PMC7333027 DOI: 10.1155/2020/4821056] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 01/16/2020] [Revised: 05/03/2020] [Accepted: 05/27/2020] [Indexed: 12/26/2022]
Abstract
Background Esophageal carcinoma (ESCA) is not only a threat to people's health but also the sixth most common cause of cancer-related mortality worldwide. Methods In this study, the key targets of ESCA are screened through GeneCards and DisGeNET databases combined with the Gene Expression Omnibus (GEO) database (GSE1420 and GSE20347). Then, data associated with ESCA samples are downloaded from The Cancer Genome Atlas (TCGA) database for integrated analysis. Moreover, the effect of epithelial cell adhesion molecule (EpCAM) expression on the survival of patients with ESCA is evaluated by Kaplan–Meier and Cox analyses. The virtual screening is carried out using a Suflex-Dock molecular docking module. The chemical components, which have been well bound to EpCAM, are screened out based on a total score >5 as a threshold. Ginsenosides and EpCAM are analyzed by LigPlot + v.2.2 software to identify the binding sites. Results Four ESCA targets are obtained from GeneCards, DisGeNET, and GEO databases. In this study, it is found that high EpCAM expression is associated with histologic grade, stage, patient age, N classification, T classification, and radiation therapy. The Kaplan–Meier curves for overall survival also show that the higher expression of EpCAM is associated with worse outcomes in patients with ESCA. Univariate and multivariate Cox analyses indicate that EpCAM mRNA expression might be a useful biomarker for ESCA(P < 0.05). Molecular docking technology suggests that ginsenoside Rg3 and ginsenoside Rh2 can easily establish good docking modes and have a high affinity with EpCAM. The 6′-hydroxyl and 6″-hydroxyl on the 3-glycosyl of ginsenoside Rg3 are prone to form hydrogen bonds (Lys151 and Lys221) with the active sites of EpCAM ligand binding domain. The hydroxyl groups on the 12 sites of the ginsenoside Rh2 glycoside framework are found to have hydrogen bonding with Leu240. The formation of hydrogen bonds plays an important role in binding of ginsenoside Rg3 and ginsenoside Rh2 to EpCAM, as well as the stability of EpCAM conformation. Conclusion EpCAM may be determined as a potential biomarker for early diagnosis and prognosis of ESCA. Ginsenoside Rg3 and ginsenoside Rh2 have potential antiesophageal cancer activities. This experiment provides a reference for the study of the chemical compositions of ginsenosides in the treatment of esophageal cancer.
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A Novel Function for KLF4 in Modulating the De-differentiation of EpCAM -/CD133 - nonStem Cells into EpCAM +/CD133 + Liver Cancer Stem Cells in HCC Cell Line HuH7. Cells 2020; 9:cells9051198. [PMID: 32408542 PMCID: PMC7290717 DOI: 10.3390/cells9051198] [Citation(s) in RCA: 33] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/02/2020] [Revised: 04/27/2020] [Accepted: 04/30/2020] [Indexed: 12/13/2022] Open
Abstract
The complex and heterogeneous nature of hepatocellular carcinoma (HCC) hampers the identification of effective therapeutic strategies. Cancer stem cells (CSCs) represent a fraction of cells within tumors with the ability to self-renew and differentiate, and thus significantly contribute to the formation and maintenance of heterogeneous tumor mass. Increasing evidence indicates high plasticity in tumor cells, suggesting that non-CSCs could acquire stem cell properties through de-differentiation or reprogramming processes. In this paper, we reveal KLF4 as a transcription factor that can induce a CSC-like phenotype in non-CSCs through upregulating the EpCAM and E-CAD expression. Our studies indicated that KLF4 could directly bind to the promoter of EpCAM and increase the number of EpCAM+/CD133+ liver cancer stem cells (LCSCs) in the HuH7 HCC cell line. When KLF4 was overexpressed in EpCAM−/CD133− non-stem cells, the expressions of hepatic stem/progenitor cell genes such as CK19, EpCAM and LGR5 were significantly increased. KLF4 overexpressing non-stem cells exhibited greater cell viability upon sorafenib treatment, while the cell migration and invasion capabilities of these cells were suppressed. Importantly, we detected an increased membranous expression and colocalization of β-CAT, E-CAD and EpCAM in the KLF4-overexpressing EpCAM−/CD133− non-stem cells, suggesting that this complex might be required for the cancer stem cell phenotype. Moreover, our in vivo xenograft studies demonstrated that with a KLF4 overexpression, EpCAM−/CD133− non-stem cells attained an in vivo tumor forming ability comparable to EpCAM+/CD133+ LCSCs, and the tumor specimens from KLF4-overexpressing xenografts had increased levels of both the KLF4 and EpCAM proteins. Additionally, we identified a correlation between the KLF4 and EpCAM protein expressions in human HCC tissues independent of the tumor stage and differentiation status. Collectively, our data suggest a novel function for KLF4 in modulating the de-differentiation of tumor cells and the induction of EpCAM+/CD133+ LCSCs in HuH7 HCC cells.
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Tang D, Chen Y, Fu GB, Yuan TJ, Huang WJ, Wang ZY, Li WJ, Jiao YF, Yu WF, Yan HX. EpCAM inhibits differentiation of human liver progenitor cells into hepatocytes in vitro by activating Notch1 signaling. Biochem Biophys Res Commun 2020; 525:S0006-291X(20)30309-0. [PMID: 32087972 DOI: 10.1016/j.bbrc.2020.02.041] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/14/2020] [Accepted: 02/08/2020] [Indexed: 12/26/2022]
Abstract
In both normal turnover of the hepatic tissue and acute hepatic injury, the liver predominantly activates terminally differentiated hepatocytes to proliferate and repair. However, in chronic and severe chronic injury, this capacity fails, and liver progenitor cells (LPCs) can give rise to hepatocytes to restore both hepatic architecture and liver metabolic function. Although the promotion of LPC-to-hepatocyte differentiation to acquire a considerable number of functional hepatocytes could serve as a potentially new therapeutic option for patients with end-stage liver disease, its development first requires the identification of the molecular mechanisms driving this process. Here, we found that the epithelial cell adhesion molecule (EpCAM), a progenitor cell marker, regulates the differentiation of LPCs into hepatocytes through Notch1 signaling pathway. Western blotting (WB) revealed a consistent expression pattern of EpCAM and Notch1 during LPC-to-hepatocyte differentiation in vitro. Additionally, overexpression of EpCAM blocked LPC-to-hepatocyte differentiation, which was in consistent with the repressive role of Notch signaling during hepatic differentiation. WB and immunofluorescence data also showed that the upregulation of EpCAM expression increased the generation of Notch intracellular domain (N1ICD), indicating the promotion of Notch1 activity. Our results established the EpCAM-Notch1 signaling axis as an inhibitory mechanism preventing LPC-to-hepatocyte differentiation in vitro.
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Affiliation(s)
- Dan Tang
- Department of Anesthesiology and Critical Care Medicine, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Yi Chen
- Department of Anesthesiology and Critical Care Medicine, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Gong-Bo Fu
- International Cooperation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, China
| | - Tian-Jie Yuan
- Department of Anesthesiology and Critical Care Medicine, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Wei-Jian Huang
- International Cooperation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, China
| | - Zhen-Yu Wang
- Department of Interventional Oncology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Wei-Jian Li
- Department of Interventional Oncology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Ying-Fu Jiao
- Department of Anesthesiology and Critical Care Medicine, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China.
| | - Wei-Feng Yu
- Department of Anesthesiology and Critical Care Medicine, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China.
| | - He-Xin Yan
- Department of Anesthesiology and Critical Care Medicine, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China; Department of Interventional Oncology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China.
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Ishiguro K, Yan IK, Lewis-Tuffin L, Patel T. Targeting Liver Cancer Stem Cells Using Engineered Biological Nanoparticles for the Treatment of Hepatocellular Cancer. Hepatol Commun 2020; 4:298-313. [PMID: 32025612 PMCID: PMC6996342 DOI: 10.1002/hep4.1462] [Citation(s) in RCA: 53] [Impact Index Per Article: 10.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/21/2019] [Accepted: 12/03/2019] [Indexed: 12/17/2022] Open
Abstract
By exploiting their biological functions, the use of biological nanoparticles such as extracellular vesicles can provide an efficient and effective approach for hepatic delivery of RNA‐based therapeutics for the treatment of liver cancers such as hepatocellular cancer (HCC). Targeting liver cancer stem cells (LCSC) within HCC provide an untapped opportunity to improve outcomes by enhancing therapeutic responses. Cells with tumor‐initiating capabilities such as LCSC can be identified by expression of markers such as epithelial cell adhesion molecule (EpCAM) on their cell surface. EpCAM is a target of Wnt/β‐catenin signaling, a fundamental pathway in stem‐cell growth. Moreover, mutations in the β‐catenin gene are frequently observed in HCC and can be associated with constitutive activation of the Wnt/β‐catenin pathway. However, targeting these pathways for the treatment of HCC has been challenging. Using RNA nanotechnology, we developed engineered biological nanoparticles capable of specific and effective delivery of RNA therapeutics targeting β‐catenin to LCSC. Extracellular vesicles isolated from milk were loaded with small interfering RNA to β‐catenin and decorated with RNA scaffolds to incorporate RNA aptamers capable of binding to EpCAM. Cellular uptake of these EpCAM‐targeting therapeutic milk‐derived nanovesicles in vitro resulted in loss of β‐catenin expression and decreased proliferation. The uptake and therapeutic efficacy of these engineered biological nanotherapeutics was demonstrated in vivo using tumor xenograft mouse models. Conclusion: β‐catenin can be targeted directly to control the proliferation of hepatic cancer stem cells using small interfering RNA delivered using target‐specific biological nanoparticles. Application of this RNA nanotechnology–based approach to engineer biological nanotherapeutics provides a platform for developing cell‐surface molecule–directed targeted therapeutics.
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Affiliation(s)
- Kaori Ishiguro
- Department of Transplantation Mayo Clinic Jacksonville FL.,Department of Cancer Biology Mayo Clinic Jacksonville FL
| | - Irene K Yan
- Department of Transplantation Mayo Clinic Jacksonville FL.,Department of Cancer Biology Mayo Clinic Jacksonville FL
| | | | - Tushar Patel
- Department of Transplantation Mayo Clinic Jacksonville FL.,Department of Cancer Biology Mayo Clinic Jacksonville FL
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Diao L, Yousuf Y, Amini‐Nik S, Jeschke MG. Increased proliferation of hepatic periportal ductal progenitor cells contributes to persistent hypermetabolism after trauma. J Cell Mol Med 2020; 24:1578-1587. [PMID: 31793707 PMCID: PMC6991656 DOI: 10.1111/jcmm.14845] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2019] [Revised: 10/16/2019] [Accepted: 10/28/2019] [Indexed: 12/13/2022] Open
Abstract
Prolonged and persistent hypermetabolism and excessive inflammatory response after severe trauma is detrimental and associated with poor outcome. The predisposing pathology or signals mediating this complex response are essentially unknown. As the liver is the central organ mediating the systemic metabolic responses and considering that adult hepatic stem cells are on top of the hierarchy of cell differentiation and may pass epigenetic information to their progeny, we asked whether liver progenitor cells are activated, signal hypermetabolism upon post-traumatic cellular stress responses, and pass this to differentiated progeny. We generated Sox9CreERT2 : ROSA26 EYFP mice to lineage-trace the periportal ductal progenitor cells (PDPCs) and verify the fate of these cells post-burn. We observed increased proliferation of PDPCs and their progeny peaking around two weeks post-burn, concomitant with the hepatomegaly and the cellular stress responses. We then sorted out PDPCs, PDPC-derived hepatocytes and mature hepatocytes, compared their transcriptome and showed that PDPCs and their progeny present a significant up-regulation in signalling pathways associated with inflammation and metabolic activation, contributing to persistent hypermetabolic and hyper-inflammatory state. Furthermore, concomitant down-regulation of LXR signalling in PDPCs and their progeny implicates the therapeutic potential of early and short-term administration of LXR agonists in ameliorating such persistent hypermetabolism.
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Affiliation(s)
- Li Diao
- Sunnybrook Research InstituteTorontoONCanada
| | | | - Saeid Amini‐Nik
- Sunnybrook Research InstituteTorontoONCanada
- Division of Plastic SurgeryDepartment of SurgeryUniversity of TorontoTorontoONCanada
- Department of Laboratory Medicine and Pathobiology (LMP)University of TorontoTorontoONCanada
| | - Marc G. Jeschke
- Sunnybrook Research InstituteTorontoONCanada
- Division of Plastic SurgeryDepartment of SurgeryUniversity of TorontoTorontoONCanada
- Department of ImmunologyUniversity of TorontoTorontoONCanada
- Ross Tilley Burn CentreSunnybrook Health Sciences CentreTorontoONCanada
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Dai M, Cai Z, Chen N, Li J, Wen J, Tan L, Guo D. [Matrine suppresses stemness of hepatocellular carcinoma cells by regulating β-catenin signaling pathway]. NAN FANG YI KE DA XUE XUE BAO = JOURNAL OF SOUTHERN MEDICAL UNIVERSITY 2019; 39:1239-1245. [PMID: 31801708 DOI: 10.12122/j.issn.1673-4254.2019.10.17] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
OBJECTIVE To explore the effects of matrine on the proliferation, tumor cell stemness, β-catenin transcriptional activity and AKT/GSK3β/β-catenin signaling pathway in human hepatocellular carcinoma (HCC) HepG2 and Huh7 cells. METHODS The proliferation and colony formation ability of HepG2 and Huh7 cells treated with 200, 400, and 800 μg/mL matrine were evaluated with MTT assay and colony formation assay, respectively. Real-time quantitative PCR was performed to detect the mRNA expressions of CD90, epithelial cell adhesion molecule (EpCAM) and CD133, and dual-luciferase assay was used to detect the transcriptional activity of β-catenin in the treated cells. The effects of matrine on the expressions of protein kinase B (AKT), P-AKT, GSK-3β, P-GSK-3β, P-β-catenin and β-catenin proteins in the Wnt/β-catenin signaling pathway were assessed using Western blotting. RESULTS Matrine inhibited the proliferation of the two HCC cell lines in a time- and concentration-dependent manner. The half-inhibitory concentrations of matrine were 2369, 1565 and 909.1 μg/mL at 24, 48 and 72 h in HepG2 cells, respectively, and were 1355, 781.8, and 612.8 μg/mL in Huh7 cells, respectively. Matrine concentrationdependently suppressed colony formation of the HCC cells, producing significant inhibitory effects at 400 μg/mL P < 0.01) and 800 μg/mL P < 0.001) in HepG2 cells and at 200 μg/mL P < 0.05), 400 μg/mL P < 0.01), and 800 μg/mL P < 0.001) in Huh7 cells. Matrine at 400 and 800 μg/mL significantly inhibited the mRNA expression of CD90, EpCAM and CD133 and the transcriptional level of β-catenin in both HepG2 and Huh7 cells P < 0.05 or 0.01). Matrine at 400 and 800 μg/mL also significantly decreased the protein levels of β-catenin, P-AKT and P-GSK-3β and increased the phosphorylation level of β-catenin in both of the cell lines. CONCLUSIONS Matrine inhibits the proliferation, colony formation, and the expressions of tumor stem cell markers CD90, EpCAM and CD133 in both HepG2 and Huh7 cells probably by inhibiting Wnt/β-catenin signaling pathway and the transcriptional activity ofβ-catenin.
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Affiliation(s)
- Meiqin Dai
- Department of Pharmacy, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China
| | - Zhuo Cai
- Department of Pharmacy, Air Force Hospital of Southern Military Command, Guangzhou 510602, China
| | - Nana Chen
- College of Pharmaceutical Science, Southern Medical University, Guangzhou 510515, China
| | - Jinzhou Li
- Department of Pharmacy, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China
| | - Jiayong Wen
- Department of Pharmacy, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China
| | - Lizhuan Tan
- Department of Pharmacy, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China
| | - Dan Guo
- Department of Pharmacy, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China
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Akbari S, Sevinç GG, Ersoy N, Basak O, Kaplan K, Sevinç K, Ozel E, Sengun B, Enustun E, Ozcimen B, Bagriyanik A, Arslan N, Önder TT, Erdal E. Robust, Long-Term Culture of Endoderm-Derived Hepatic Organoids for Disease Modeling. Stem Cell Reports 2019; 13:627-641. [PMID: 31522975 PMCID: PMC6829764 DOI: 10.1016/j.stemcr.2019.08.007] [Citation(s) in RCA: 98] [Impact Index Per Article: 16.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2018] [Revised: 08/10/2019] [Accepted: 08/16/2019] [Indexed: 12/18/2022] Open
Abstract
Organoid technologies have become a powerful emerging tool to model liver diseases, for drug screening, and for personalized treatments. These applications are, however, limited in their capacity to generate functional hepatocytes in a reproducible and efficient manner. Here, we generated and characterized the hepatic organoid (eHEPO) culture system using human induced pluripotent stem cell (iPSC)-derived EpCAM-positive endodermal cells as an intermediate. eHEPOs can be produced within 2 weeks and expanded long term (>16 months) without any loss of differentiation capacity to mature hepatocytes. Starting from patient-specific iPSCs, we modeled citrullinemia type 1, a urea cycle disorder caused by mutations in the argininosuccinate synthetase (ASS1) enzyme. The disease-related ammonia accumulation phenotype in eHEPOs could be reversed by the overexpression of the wild-type ASS1 gene, which also indicated that this model is amenable to genetic manipulation. Thus, eHEPOs are excellent unlimited cell sources to generate functional hepatic organoids in a fast and efficient manner.
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Affiliation(s)
- Soheil Akbari
- Izmir Biomedicine and Genome Center, 35340 Izmir, Turkey; Department of Medical Biology and Genetics, Health Science Institute, Dokuz Eylul University, 35340 Izmir, Turkey
| | | | - Nevin Ersoy
- Department of Histology and Embryology, Health Science Institute, Dokuz Eylul University, 35340 Izmir, Turkey
| | - Onur Basak
- Hubrecht Institute, University Medical Center Utrecht, 3584 CT Utrecht, the Netherlands; Department of Translational Neuroscience, Brain Center Rudolf Magnus, University Medical Center Utrecht, 3584 CG Utrecht, the Netherlands
| | - Kubra Kaplan
- Izmir Biomedicine and Genome Center, 35340 Izmir, Turkey; Department of Genome Sciences and Molecular Biotechnology, Izmir International Biomedicine and Genome Institute, Dokuz Eylul University, 35340 Izmir, Turkey
| | - Kenan Sevinç
- Faculty of Medicine, Koç University, 34450 Istanbul, Turkey
| | - Erkin Ozel
- Faculty of Medicine, Koç University, 34450 Istanbul, Turkey
| | - Berke Sengun
- Faculty of Medicine, Koç University, 34450 Istanbul, Turkey
| | - Eray Enustun
- Faculty of Medicine, Koç University, 34450 Istanbul, Turkey
| | - Burcu Ozcimen
- Faculty of Medicine, Koç University, 34450 Istanbul, Turkey
| | - Alper Bagriyanik
- Izmir Biomedicine and Genome Center, 35340 Izmir, Turkey; Department of Histology and Embryology, Health Science Institute, Dokuz Eylul University, 35340 Izmir, Turkey; Department of Histology and Embryology, Faculty of Medicine, Dokuz Eylul University, 35340 Izmir, Turkey
| | - Nur Arslan
- Izmir Biomedicine and Genome Center, 35340 Izmir, Turkey; Department of Pediatric Gastroenterology and Metabolism, Faculty of Medicine, Dokuz Eylul University, 35340 Izmir, Turkey
| | | | - Esra Erdal
- Izmir Biomedicine and Genome Center, 35340 Izmir, Turkey; Department of Medical Biology and Genetics, Health Science Institute, Dokuz Eylul University, 35340 Izmir, Turkey.
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Zhang Q, Cao WS, Wang XQ, Zhang M, Lu XM, Chen JQ, Chen Y, Ge MM, Zhong CY, Han HY. Genistein inhibits nasopharyngeal cancer stem cells through sonic hedgehog signaling. Phytother Res 2019; 33:2783-2791. [PMID: 31342620 DOI: 10.1002/ptr.6464] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2018] [Revised: 07/07/2019] [Accepted: 07/09/2019] [Indexed: 01/03/2025]
Abstract
Genistein, a soy derived isoflavanoid compound, exerts anticancer effects in various cancers. Nasopharyngeal cancer stem cells (NCSCs) are a small subpopulation of cancer cells which are responsible for initiation, progression, metastasis, and recurrence of nasopharyngeal cancer. The present study aimed to investigate the suppressive effects of genistein on NCSCs and its underlying mechanism. NCSCs were enriched from human nasopharyngeal cancer cell lines CNE2 and HONE1 through tumorsphere-forming assay. It was shown that genistein inhibited the tumorsphere formation capacity, decreased the number of EpCAM+ cells, downregulated the expression of NCSCs markers, suppressed cell proliferation, and induced apoptosis of NCSCs. Genistein suppressed the activity of Sonic hedgehog (SHH) signaling, which was important for the maintenance of NCSCs, while activation of SHH signaling by purmorphamine diminished the inhibitory effects of genistein on NCSCs. Our data suggested that genistein inhibited NCSCs through the suppression of SHH signaling. These findings support the use of genistein for targeting NCSCs.
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Affiliation(s)
- Qi Zhang
- Department of Nutrition and Food Safety, School of Public Health, Nanjing Medical University, Nanjing, China
| | - Wan-Shuang Cao
- Department of Nutrition and Food Safety, School of Public Health, Nanjing Medical University, Nanjing, China
| | - Xue-Qi Wang
- Department of Nutrition and Food Safety, School of Public Health, Nanjing Medical University, Nanjing, China
| | - Min Zhang
- Department of Nutrition and Food Safety, School of Public Health, Nanjing Medical University, Nanjing, China
| | - Xiao-Min Lu
- Department of Nutrition and Food Safety, School of Public Health, Nanjing Medical University, Nanjing, China
| | - Jia-Qi Chen
- Department of Nutrition and Food Safety, School of Public Health, Nanjing Medical University, Nanjing, China
| | - Yue Chen
- Department of Nutrition and Food Safety, School of Public Health, Nanjing Medical University, Nanjing, China
| | - Miao-Miao Ge
- Department of Nutrition and Food Safety, School of Public Health, Nanjing Medical University, Nanjing, China
| | - Cai-Yun Zhong
- Department of Nutrition and Food Safety, School of Public Health, Nanjing Medical University, Nanjing, China
- Center for Global Health, School of Public Health, Nanjing Medical University, Nanjing, China
| | - Hong-Yu Han
- Department of Clinical Nutrition, Sun Yat-sen University Cancer Center, State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Guangzhou, China
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Wang S, Kim J, Lee C, Oh D, Han J, Kim TJ, Kim SW, Seo YS, Oh SH, Jung Y. Tumor necrosis factor-inducible gene 6 reprograms hepatic stellate cells into stem-like cells, which ameliorates liver damage in mouse. Biomaterials 2019; 219:119375. [DOI: 10.1016/j.biomaterials.2019.119375] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2019] [Revised: 07/19/2019] [Accepted: 07/20/2019] [Indexed: 12/12/2022]
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50
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The Emerging Roles of Cancer Stem Cells and Wnt/Beta-Catenin Signaling in Hepatoblastoma. Cancers (Basel) 2019; 11:cancers11101406. [PMID: 31547062 PMCID: PMC6826653 DOI: 10.3390/cancers11101406] [Citation(s) in RCA: 31] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2019] [Revised: 09/11/2019] [Accepted: 09/11/2019] [Indexed: 02/07/2023] Open
Abstract
Hepatoblastoma (HB) is the most common form of primary liver malignancy found in pediatric populations. HB is considered to be clonal and arises from hepatoblasts, or embryonic liver progenitor cells. These less differentiated tumor-initiating progenitor cells, or cancer stem cells (CSCs), may contribute to tumor recurrence and resistance to therapies, and have high metastatic abilities. Phenotypic heterogeneity, undesired genetic and epigenetic alterations, and dysregulated signaling pathways provide CSCs with a survival advantage over current therapies. The molecular and cellular basis of HB and the mechanism of CSC induction are not fully understood. The Wnt/beta-catenin pathway is one of the major developmental pathways and is believed to play an important role in the pathogenesis of HB and CSC formation. This review summarizes the cellular and molecular characteristics of HB with a specific emphasis on CSCs and Wnt/beta-catenin signaling.
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