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Gibson J, Dhungana A, Pokhrel M, Arthur B, Suresh P, Adebayo O, Cottle RN. Validation of Clinical-Grade Electroporation Systems for CRISPR-Cas9-Mediated Gene Therapy in Primary Hepatocytes for the Correction of Inherited Metabolic Liver Disease. Cells 2025; 14:711. [PMID: 40422214 PMCID: PMC12109753 DOI: 10.3390/cells14100711] [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: 03/31/2025] [Revised: 05/05/2025] [Accepted: 05/08/2025] [Indexed: 05/28/2025] Open
Abstract
Hepatocyte transplantation (HTx) combined with ex vivo gene therapy has garnered significant interest due to its potential for treating many inherited metabolic liver diseases. The biggest obstacle for HTx is achieving sufficient engraftment levels to rescue diseased phenotypes, which becomes more challenging when combined with ex vivo gene editing techniques. However, recent technological advancements have improved electroporation delivery efficiency, cell viability, and scalability for cell therapy. We recently demonstrated the impacts of electroporation for cell-based gene therapy in a mouse model of hereditary tyrosinemia type 1 (HT1). Here, we explore the use of the clinical-grade electroporator, the MaxCyte ExPERT GTx, utilized in the first FDA-approved CRISPR therapy, Casgevy, and evaluate its potential in primary hepatocytes in terms of delivery efficiency and cell viability. We assessed the gene editing efficiency and post-transplantation engraftment of hepatocytes from mTmG mice electroporated with CRISPR-Cas9-ribonucleoproteins (RNPs) targeting 4-hydroxyphenylpyruvate dioxygenase (Hpd) in a fumarylacetoacetate hydrolase (Fah)-deficient mouse model of HT1. After surgery, Fah-/- graft recipients were cycled off and on nitisinone to achieve independence from drug-induced Hpd inhibition, an indicator of HT1 disease correction. Transplanted hepatocytes subjected to electroporation using the GTx system had a cell viability of 89.9% and 100% on-target gene editing efficiency. Recipients transplanted with GTx-electroporated cells showed a smaller weight reduction than controls transplanted with untransfected cells (7.9% and 13.8%, respectively). Further, there were no mortalities in the GTx-recipient mice, whereas there was 25% mortality in the control recipients. Mean donor cell engraftment was significantly higher in GTx-recipient mice compared to untransfected control recipients (97.9% and 81.6%, respectively). Our results indicate that the GTx system does not negatively impact hepatocyte functionality and engraftment potential, thereby demonstrating the promise of GTx electroporation in hepatocytes as a viable cell therapy for treating genetic diseases that affect the liver.
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Affiliation(s)
| | | | | | | | | | | | - Renee N. Cottle
- Department of Bioengineering, Clemson University, Clemson, SC 29634, USA; (J.G.); (A.D.); (M.P.); (B.A.); (P.S.); (O.A.)
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2
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Selvanathan A, Kankananarachchi I, Bansal S, Fitzpatrick E, Lemonde H, Turner C, Fairbanks L, White FJ, Dhawan A, Schwahn BC. Early postnatal hepatocyte transplantation in a child with molybdenum cofactor deficiency type B. Mol Genet Metab 2025; 145:109079. [PMID: 40121797 DOI: 10.1016/j.ymgme.2025.109079] [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: 01/14/2025] [Revised: 02/27/2025] [Accepted: 03/04/2025] [Indexed: 03/25/2025]
Abstract
Molybdenum cofactor deficiencies (MoCD) are a group of inborn errors of metabolism that result in impaired synthesis of molybdenum cofactor, crucial for the function of three oxidases (sulfite oxidase, xanthine oxidase and aldehyde oxidase). Most patients present with severe neonatal-onset epileptic encephalopathy, hypotonia, poor feeding and apnoea, with death typically occurring within the first three years of life. Whilst there is now an emerging therapy for MoCD Type A (cPMP/fosdenopterin), this treatment is not effective for MoCD Type B and there is no treatment for isolated sulfite oxidase deficiency (ISOD). Liver directed gene delivery is a potential alternative therapy for sulfite intoxication disorders. We report an attempt to use hepatocyte transplantation as a treatment option for MoCD Type B, in an infant with a strong family history of neonatal-onset disease and early mortality. Six transfusions of hepatocytes were given between Day 1 and Day 18 of life, totalling around 1 × 109 cells with immunosuppressive cover. Concomitantly dietary protein restriction was maintained at 2 g/kg, including 0.7 g/kg of methionine- and cyst(e)ine-free amino acid mixture. The aim was to utilize hepatocyte transplantation as a bridge to liver transplantation. Whilst there was evidence of biochemical stabilization with reduction in concentrations of sulfite and S-sulfocysteine and a moderate increase in urate levels compared to the sibling, the treatment was not able to prevent acute brain injury from sulfite toxicity which was evident in neuroimaging at 35 h of age. This correlated clinically with ongoing seizures as well as minimal developmental progress.
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Affiliation(s)
- A Selvanathan
- Willink Metabolic Unit, Manchester Centre for Genomic Medicine, St Mary's Hospital, Manchester, United Kingdom
| | - I Kankananarachchi
- Willink Metabolic Unit, Manchester Centre for Genomic Medicine, St Mary's Hospital, Manchester, United Kingdom
| | - S Bansal
- Paediatric Liver GI and Nutrition Centre and Mowat Labs, King's College Hospital, London, United Kingdom
| | - E Fitzpatrick
- Paediatric Liver GI and Nutrition Centre and Mowat Labs, King's College Hospital, London, United Kingdom
| | - H Lemonde
- Paediatric Metabolic Unit, Evelina London Children's Hospital, London, United Kingdom
| | - C Turner
- WellChild Laboratory, Evelina London Children's Hospital, St Thomas' Hospital, London, United Kingdom
| | - L Fairbanks
- Purine Research Lab, Biochemical Sciences, Synnovis, Guys & St Thomas' NHS Foundation Trust, London, UK
| | - F J White
- Willink Metabolic Unit, Manchester Centre for Genomic Medicine, St Mary's Hospital, Manchester, United Kingdom
| | - A Dhawan
- Paediatric Liver GI and Nutrition Centre and Mowat Labs, King's College Hospital, London, United Kingdom
| | - B C Schwahn
- Willink Metabolic Unit, Manchester Centre for Genomic Medicine, St Mary's Hospital, Manchester, United Kingdom; Division of Evolution and Genomic Sciences, Faculty of Biology, Medicine and Health, University of Manchester, UK.
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Ortuño-Costela MC, Pinzani M, Vallier L. Cell therapy for liver disorders: past, present and future. Nat Rev Gastroenterol Hepatol 2025; 22:329-342. [PMID: 40102584 DOI: 10.1038/s41575-025-01050-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 02/11/2025] [Indexed: 03/20/2025]
Abstract
The liver fulfils a plethora of vital functions and, due to their importance, liver dysfunction has life-threatening consequences. Liver disorders currently account for more than two million deaths annually worldwide and can be classified broadly into three groups, considering their onset and aetiology, as acute liver diseases, inherited metabolic disorders and chronic liver diseases. In the most advanced and severe forms leading to liver failure, liver transplantation is the only treatment available, which has many associated drawbacks, including a shortage of organ donors. Cell therapy via fully mature cell transplantation is an advantageous alternative that may be able to restore a damaged organ's functionality or serve as a bridge until regeneration can occur. Pioneering work has shown that transplanting adult hepatocytes can support liver recovery. However, primary hepatocytes cannot be grown extensively in vitro as they rapidly lose their metabolic activity. Therefore, different cell sources are currently being tested as alternatives to primary cells. Human pluripotent stem cell-derived cells, chemically induced liver progenitors, or 'liver' organoids, hold great promise for developing new cell therapies for acute and chronic liver diseases. This Review focuses on the advantages and drawbacks of distinct cell sources and the relative strategies to address different therapeutic needs in distinct liver diseases.
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Affiliation(s)
- M Carmen Ortuño-Costela
- Berlin Institute of Health, BIH Centre for Regenerative Therapies, Charité-Universitätsmedizin, Berlin, Germany
- Max Planck Institute for Molecular Genetics, Berlin, Germany
| | - Massimo Pinzani
- University College London Institute for Liver and Digestive Health, Division of Medicine, Royal Free Hospital, London, UK
- University of Pittsburgh Medical Center-Mediterranean Institute for Transplantation and Highly Specialized Therapies (UPMC-ISMETT), Palermo, Italy
| | - Ludovic Vallier
- Berlin Institute of Health, BIH Centre for Regenerative Therapies, Charité-Universitätsmedizin, Berlin, Germany.
- Max Planck Institute for Molecular Genetics, Berlin, Germany.
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4
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Shi H, Ding Y, Sun P, Lv Z, Wang C, Ma H, Lu J, Yu B, Li W, Wang C. Chemical approaches targeting the hurdles of hepatocyte transplantation: mechanisms, applications, and advances. Front Cell Dev Biol 2024; 12:1480226. [PMID: 39544361 PMCID: PMC11560891 DOI: 10.3389/fcell.2024.1480226] [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: 08/13/2024] [Accepted: 10/21/2024] [Indexed: 11/17/2024] Open
Abstract
Hepatocyte transplantation (HTx) has been a novel cell-based therapy for severe liver diseases, as the donor livers for orthotopic liver transplantation are of great shortage. However, HTx has been confronted with two main hurdles: limited high-quality hepatocyte sources and low cell engraftment and repopulation rate. To cope with, researchers have investigated on various strategies, including small molecule drugs with unique advantages. Small molecules are promising chemical tools to modulate cell fate and function for generating high quality hepatocyte sources. In addition, endothelial barrier, immune responses, and low proliferative efficiency of donor hepatocytes mainly contributes to low cell engraftment and repopulation rate. Interfering these biological processes with small molecules is beneficial for improving cell engraftment and repopulation. In this review, we will discuss the applications and advances of small molecules in modulating cell differentiation and reprogramming for hepatocyte resources and in improving cell engraftment and repopulation as well as its underlying mechanisms.
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Affiliation(s)
- Huanxiao Shi
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Yi Ding
- Experimental Teaching Center, Naval Medical University, Shanghai, China
| | - Pingxin Sun
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Zhuman Lv
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Chunyan Wang
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Haoxin Ma
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Junyu Lu
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Bing Yu
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Wenlin Li
- Department of Cell Biology, Naval Medical University, Shanghai, China
- Shanghai Key Laboratory of Cell Engineering, Naval Medical University, Shanghai, China
| | - Chao Wang
- Department of Cell Biology, Naval Medical University, Shanghai, China
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Sambati V, Laudisio S, Motta M, Esposito S. Therapeutic Options for Crigler-Najjar Syndrome: A Scoping Review. Int J Mol Sci 2024; 25:11006. [PMID: 39456788 PMCID: PMC11508002 DOI: 10.3390/ijms252011006] [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: 09/13/2024] [Revised: 10/01/2024] [Accepted: 10/11/2024] [Indexed: 10/28/2024] Open
Abstract
Crigler-Najjar Syndrome (CNS) is a rare genetic disorder caused by mutations in the UGT1A1 gene, leading to impaired bilirubin conjugation and severe unconjugated hyperbilirubinemia. CNS presents in the following forms: CNS type 1 (CNS1), the more severe form with the complete absence of UGT1A1 activity, and CNS type 2 (CNS2), with partial enzyme activity. This narrative review aims to provide a detailed overview of CNS, highlighting its clinical significance and the need for new, more effective treatments. By summarizing current knowledge and discussing future treatments, this article seeks to encourage further research and advancements that can improve outcomes for CNS patients. The literature analysis showed that CNS1 requires aggressive management, including phototherapy and plasmapheresis, but liver transplantation (LT) remains the only definitive cure. The timing of LT is critical, as it must be performed before the onset of irreversible brain damage (kernicterus), making early intervention essential. However, LT poses risks such as graft rejection and lifelong immunosuppression. CNS2 is milder, with patients responding well to phenobarbital and having a lower risk of kernicterus. Recent advancements in gene therapy and autologous hepatocyte transplantation offer promising alternatives to LT. Gene therapy using adeno-associated virus (AAV) vectors has shown potential in preclinical studies, though challenges remain in pediatric applications due to liver growth and pre-existing immunity. Autologous hepatocyte transplantation avoids the risk of rejection but requires further research. These emerging therapies provide hope for more effective and less invasive treatment options, aiming to improve the quality of life for CNS patients and reduce reliance on lifelong interventions.
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Affiliation(s)
| | | | | | - Susanna Esposito
- Pediatric Clinic, Department of Medicine and Surgery, University Hospital of Parma, 43126 Parma, Italy; (V.S.); (S.L.)
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Udagawa D, Nagata S, Yagi H, Nishi K, Morisaku T, Adachi S, Nakano Y, Tanaka M, Hori S, Hasegawa Y, Abe Y, Kitago M, Kitagawa Y. A Novel Approach to Orthotopic Hepatocyte Transplantation Engineered With Liver Hydrogel for Fibrotic Livers, Enhancing Cell-Cell Interaction and Angiogenesis. Cell Transplant 2024; 33:9636897241253700. [PMID: 38770981 PMCID: PMC11110510 DOI: 10.1177/09636897241253700] [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: 10/08/2023] [Revised: 04/22/2024] [Accepted: 04/22/2024] [Indexed: 05/22/2024] Open
Abstract
Hepatocyte transplantation (HCT) is a potential bridging therapy or an alternative to liver transplantation. Conventionally, single-cell hepatocytes are injected via the portal vein. This strategy, however, has yet to overcome poor cell engraftment and function. Therefore, we developed an orthotopic HCT method using a liver-derived extracellular matrix (L-ECM) gel. PXB cells (flesh mature human hepatocytes) were dispersed into the hydrogel solution in vitro, and the gel solution was immediately gelated in 37°C incubators to investigate the affinity between mature human hepatocyte and the L-ECM gel. During the 3-day cultivation in hepatocyte medium, PXB cells formed cell aggregates via cell-cell interactions. Quantitative analysis revealed human albumin production in culture supernatants. For the in vivo assay, PXB cells were encapsulated in the L-ECM gel and transplanted between the liver lobes of normal rats. Pathologically, the L-ECM gel was localized at the transplant site and retained PXB cells. Cell survival and hepatic function marker expression were verified in another rat model wherein thioacetamide was administered to induce liver fibrosis. Moreover, cell-cell interactions and angiogenesis were enhanced in the L-ECM gel compared with that in the collagen gel. Our results indicate that L-ECM gels can help engraft transplanted hepatocytes and express hepatic function as a scaffold for cell transplantation.
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Affiliation(s)
- Daisuke Udagawa
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Shogo Nagata
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Hiroshi Yagi
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Kotaro Nishi
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | | | - Shungo Adachi
- Fundamental Innovative Oncology Core, National Cancer Center Research Institute, Tokyo, Japan
| | - Yutaka Nakano
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Masayuki Tanaka
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Shutaro Hori
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Yasushi Hasegawa
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Yuta Abe
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Minoru Kitago
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Yuko Kitagawa
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
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7
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Sun Z, Yuan X, Wu J, Wang C, Zhang K, Zhang L, Hui L. Hepatocyte transplantation: The progress and the challenges. Hepatol Commun 2023; 7:e0266. [PMID: 37695736 PMCID: PMC10497249 DOI: 10.1097/hc9.0000000000000266] [Citation(s) in RCA: 8] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/13/2023] [Accepted: 07/26/2023] [Indexed: 09/13/2023] Open
Abstract
Numerous studies have shown that hepatocyte transplantation is a promising approach for liver diseases, such as liver-based metabolic diseases and acute liver failure. However, it lacks strong evidence to support the long-term therapeutic effects of hepatocyte transplantation in clinical practice. Currently, major hurdles include availability of quality-assured hepatocytes, efficient engraftment and repopulation, and effective immunosuppressive regimens. Notably, cell sources have been advanced recently by expanding primary human hepatocytes by means of dedifferentiation in vitro. Moreover, the transplantation efficiency was remarkably improved by the established preparative hepatic irradiation in combination with hepatic mitogenic stimuli regimens. Finally, immunosuppression drugs, including glucocorticoid and inhibitors for co-stimulating signals of T cell activation, were proposed to prevent innate and adaptive immune rejection of allografted hepatocytes. Despite remarkable progress, further studies are required to improve in vitro cell expansion technology, develop clinically feasible preconditioning regimens, and further optimize immunosuppression regimens or establish ex vivo gene correction-based autologous hepatocyte transplantation.
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Affiliation(s)
- Zhen Sun
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
- School of Life Science and Technology, ShanghaiTech University, Shanghai, China
| | - Xiang Yuan
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Jingqi Wu
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Chenhua Wang
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Kun Zhang
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Ludi Zhang
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Lijian Hui
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
- School of Life Science and Technology, ShanghaiTech University, Shanghai, China
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8
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Chen L, Zhang N, Huang Y, Zhang Q, Fang Y, Fu J, Yuan Y, Chen L, Chen X, Xu Z, Li Y, Izawa H, Xiang C. Multiple Dimensions of using Mesenchymal Stem Cells for Treating Liver Diseases: From Bench to Beside. Stem Cell Rev Rep 2023; 19:2192-2224. [PMID: 37498509 DOI: 10.1007/s12015-023-10583-5] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 06/26/2023] [Indexed: 07/28/2023]
Abstract
Liver diseases impose a huge burden worldwide. Although hepatocyte transplantation has long been considered as a potential strategy for treating liver diseases, its clinical implementation has created some obvious limitations. As an alternative strategy, cell therapy, particularly mesenchymal stem cell (MSC) transplantation, is widely used in treating different liver diseases, including acute liver disease, acute-on-chronic liver failure, hepatitis B/C virus, autoimmune hepatitis, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, alcoholic liver disease, liver fibrosis, liver cirrhosis, and hepatocellular carcinoma. Here, we summarize the status of MSC transplantation in treating liver diseases, focusing on the therapeutic mechanisms, including differentiation into hepatocyte-like cells, immunomodulating function with a variety of immune cells, paracrine effects via the secretion of various cytokines and extracellular vesicles, and facilitation of homing and engraftment. Some improved perspectives and current challenges are also addressed. In summary, MSCs have great potential in the treatment of liver diseases based on their multi-faceted characteristics, and more accurate mechanisms and novel therapeutic strategies stemming from MSCs will facilitate clinical practice.
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Affiliation(s)
- Lijun Chen
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, National Medical Center for Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, 310003, People's Republic of China
- Research Units of Infectious Disease and Microecology, Chinese Academy of Medical Sciences, Hangzhou, Zhejiang, 310003, People's Republic of China
| | - Ning Zhang
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, National Medical Center for Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, 310003, People's Republic of China
- Research Units of Infectious Disease and Microecology, Chinese Academy of Medical Sciences, Hangzhou, Zhejiang, 310003, People's Republic of China
| | - Yuqi Huang
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, National Medical Center for Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, 310003, People's Republic of China
- Research Units of Infectious Disease and Microecology, Chinese Academy of Medical Sciences, Hangzhou, Zhejiang, 310003, People's Republic of China
| | - Qi Zhang
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, National Medical Center for Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, 310003, People's Republic of China
- Research Units of Infectious Disease and Microecology, Chinese Academy of Medical Sciences, Hangzhou, Zhejiang, 310003, People's Republic of China
| | - Yangxin Fang
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, National Medical Center for Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, 310003, People's Republic of China
- Research Units of Infectious Disease and Microecology, Chinese Academy of Medical Sciences, Hangzhou, Zhejiang, 310003, People's Republic of China
| | - Jiamin Fu
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, National Medical Center for Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, 310003, People's Republic of China
- Research Units of Infectious Disease and Microecology, Chinese Academy of Medical Sciences, Hangzhou, Zhejiang, 310003, People's Republic of China
| | - Yin Yuan
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, National Medical Center for Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, 310003, People's Republic of China
- Research Units of Infectious Disease and Microecology, Chinese Academy of Medical Sciences, Hangzhou, Zhejiang, 310003, People's Republic of China
| | - Lu Chen
- Innovative Precision Medicine (IPM) Group, Hangzhou, Zhejiang, 311215, People's Republic of China
| | - Xin Chen
- Department of Hematology, Affiliated Hangzhou First People's Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, 310027, People's Republic of China
| | - Zhenyu Xu
- Innovative Precision Medicine (IPM) Group, Hangzhou, Zhejiang, 311215, People's Republic of China
| | - Yifei Li
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, National Medical Center for Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, 310003, People's Republic of China
- Research Units of Infectious Disease and Microecology, Chinese Academy of Medical Sciences, Hangzhou, Zhejiang, 310003, People's Republic of China
| | - Hiromi Izawa
- Jingugaien Woman Life Clinic, Jingu-Gaien 3-39-5 2F, Shibuya-Ku, Tokyo, Japan
| | - Charlie Xiang
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, National Medical Center for Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, 310003, People's Republic of China.
- Research Units of Infectious Disease and Microecology, Chinese Academy of Medical Sciences, Hangzhou, Zhejiang, 310003, People's Republic of China.
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9
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Wakil A, Niazi M, Lunsford KE, Pyrsopoulos N. Future Approaches and Therapeutic Modalities for Acute-on-Chronic Liver Failure. Clin Liver Dis 2023; 27:777-790. [PMID: 37380297 DOI: 10.1016/j.cld.2023.03.017] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/30/2023]
Abstract
Acute-on-chronic liver failure (ACLF) results from an acute decompensation of cirrhosis due to exogenous insult. The condition is characterized by a severe systemic inflammatory response, inappropriate compensatory anti-inflammatory response, multisystem extrahepatic organ failure, and high short-term mortality. Here, the authors evaluate the current status of potential treatments for ACLF and assess their efficacy and therapeutic potential.
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Affiliation(s)
- Ali Wakil
- Division of Gastroenterology and Hepatology, Department of Medicine, Rutgers New Jersey Medical School, 185 South Orange Avenue, MSB H536, Newark, NJ 07103, USA
| | - Mumtaz Niazi
- Division of Gastroenterology and Hepatology, Department of Medicine, Rutgers New Jersey Medical School, 185 South Orange Avenue, MSB H536, Newark, NJ 07103, USA
| | - Keri E Lunsford
- Department of Surgery, Division of Liver Transplant and HPB Surgery, Rutgers New Jersey Medical School, 185 South Orange Avenue, MSB H536, Newark, NJ 07103, USA
| | - Nikolaos Pyrsopoulos
- Division of Gastroenterology and Hepatology, Department of Medicine, Rutgers New Jersey Medical School, 185 South Orange Avenue, MSB H536, Newark, NJ 07103, USA.
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10
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Nocentini A, Bonardi A, Pratesi S, Gratteri P, Dani C, Supuran CT. Pharmaceutical strategies for preventing toxicity and promoting antioxidant and anti-inflammatory actions of bilirubin. J Enzyme Inhib Med Chem 2022; 37:487-501. [PMID: 34986721 PMCID: PMC8741241 DOI: 10.1080/14756366.2021.2020773] [Citation(s) in RCA: 23] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2021] [Revised: 12/02/2021] [Accepted: 12/03/2021] [Indexed: 12/25/2022] Open
Abstract
Bilirubin (BR) is the final product of haem catabolism. Disruptions along BR metabolic/transport pathways resulting from inherited disorders can increase plasma BR concentration (hyperbilirubinaemia). Unconjugated hyperbilirubinemia may induce BR accumulation in brain, potentially causing irreversible neurological damage, a condition known as BR encephalopathy or kernicterus, to which newborns are especially vulnerable. Numerous pharmaceutical strategies, mostly based on hemoperfusion, have been proposed over the last decades to identify new valid, low-risk alternatives for BR removal from plasma. On the other hand, accumulating evidence indicates that BR produces health benefits due to its potent antioxidant, anti-inflammatory and immunomodulatory action with a significant potential for the treatment of a multitude of diseases. The present manuscript reviews both such aspects of BR pharmacology, gathering literature data on applied pharmaceutical strategies adopted to: (i) reduce the plasma BR concentration for preventing neurotoxicity; (ii) produce a therapeutic effect based on BR efficacy in the treatment of many disorders.
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Affiliation(s)
- Alessio Nocentini
- Department of Neurosciences, Psychology, Drug Research and Child Health, Pharmaceutical and Nutraceutical Section, University of Florence, Florence, Italy
| | - Alessandro Bonardi
- Department of Neurosciences, Psychology, Drug Research and Child Health, Pharmaceutical and Nutraceutical Section, University of Florence, Florence, Italy
| | - Simone Pratesi
- Department of Neurosciences, Psychology, Drug Research and Child Health, Careggi University, Hospital of Florence, Florence, Italy
| | - Paola Gratteri
- Department of Neurosciences, Psychology, Drug Research and Child Health, Pharmaceutical and Nutraceutical Section, Laboratory of Molecular Modelling Cheminformatics & QSAR, University of Florence, Florence, Italy
| | - Carlo Dani
- Department of Neurosciences, Psychology, Drug Research and Child Health, Careggi University, Hospital of Florence, Florence, Italy
| | - Claudiu T. Supuran
- Department of Neurosciences, Psychology, Drug Research and Child Health, Pharmaceutical and Nutraceutical Section, University of Florence, Florence, Italy
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Liver Regeneration and Cell Transplantation for End-Stage Liver Disease. Biomolecules 2021; 11:biom11121907. [PMID: 34944550 PMCID: PMC8699389 DOI: 10.3390/biom11121907] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2021] [Revised: 12/12/2021] [Accepted: 12/14/2021] [Indexed: 02/06/2023] Open
Abstract
Liver transplantation is the only curative option for end-stage liver disease; however, the limitations of liver transplantation require further research into other alternatives. Considering that liver regeneration is prevalent in liver injury settings, regenerative medicine is suggested as a promising therapeutic strategy for end-stage liver disease. Upon the source of regenerating hepatocytes, liver regeneration could be divided into two categories: hepatocyte-driven liver regeneration (typical regeneration) and liver progenitor cell-driven liver regeneration (alternative regeneration). Due to the massive loss of hepatocytes, the alternative regeneration plays a vital role in end-stage liver disease. Advances in knowledge of liver regeneration and tissue engineering have accelerated the progress of regenerative medicine strategies for end-stage liver disease. In this article, we generally reviewed the recent findings and current knowledge of liver regeneration, mainly regarding aspects of the histological basis of regeneration, histogenesis and mechanisms of hepatocytes' regeneration. In addition, this review provides an update on the regenerative medicine strategies for end-stage liver disease. We conclude that regenerative medicine is a promising therapeutic strategy for end-stage liver disease. However, further studies are still required.
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12
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Anand H, Nulty J, Dhawan A. Cell therapy in congenital inherited hepatic disorders. Best Pract Res Clin Gastroenterol 2021; 56-57:101772. [PMID: 35331403 DOI: 10.1016/j.bpg.2021.101772] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/05/2021] [Revised: 10/18/2021] [Accepted: 10/27/2021] [Indexed: 01/31/2023]
Abstract
Congenital inherited hepatic disorders (CIHDs) are a set of diverse and heterogeneous group of genetic disorders leading to a defect in an enzyme or transporter. Most of these disorders are currently treated by liver transplantation as standard of care. Improved surgical techniques and post-operative care has led to a wider availability and success of liver transplantation program worldwide. However liver transplantation has its own limitations due to invasive surgery and lifelong use of immunosuppressive agents. Our experience from auxiliary liver transplantation (where right or the left lobe of the patient liver is replaced with a healthy liver donor) demonstrated successful treatment of the underlying defect of noncirrhotic metabolic disorder suggesting that whole liver replacement may not be necessary to achieve a change in phenotype. Large number of animal studies in human models of CIHD have shown success of hepatocyte transplantation leading to its human use. This review addresses the current state of human hepatocyte transplantation in the management of CIHDs with bottlenecks to its wider application and future perspectives.
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Affiliation(s)
- Hanish Anand
- King's College Hospital NHS Trust: King's College Hospital NHS Foundation Trust, United Kingdom; DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
| | - Jessica Nulty
- King's College Hospital NHS Trust: King's College Hospital NHS Foundation Trust, United Kingdom; DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
| | - Anil Dhawan
- King's College Hospital NHS Trust: King's College Hospital NHS Foundation Trust, United Kingdom; DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK.
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13
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Pluta KD, Ciezkowska M, Wisniewska M, Wencel A, Pijanowska DG. Cell-based clinical and experimental methods for assisting the function of impaired livers – Present and future of liver support systems. Biocybern Biomed Eng 2021. [DOI: 10.1016/j.bbe.2021.06.005] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
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14
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Parsons RF, Baquerizo A, Kirchner VA, Malek S, Desai CS, Schenk A, Finger EB, Brennan TV, Parekh KR, MacConmara M, Brayman K, Fair J, Wertheim JA. Challenges, highlights, and opportunities in cellular transplantation: A white paper of the current landscape. Am J Transplant 2021; 21:3225-3238. [PMID: 34212485 DOI: 10.1111/ajt.16740] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2020] [Revised: 06/22/2021] [Accepted: 06/22/2021] [Indexed: 02/05/2023]
Abstract
Although cellular transplantation remains a relatively small field compared to solid organ transplantation, the prospects for advancement in basic science and clinical care remain bountiful. In this review, notable historical events and the current landscape of the field of cellular transplantation are reviewed with an emphasis on islets (allo- and xeno-), hepatocytes (including bioartificial liver), adoptive regulatory immunotherapy, and stem cells (SCs, specifically endogenous organ-specific and mesenchymal). Also, the nascent but rapidly evolving field of three-dimensional bioprinting is highlighted, including its major processing steps and latest achievements. To reach its full potential where cellular transplants are a more viable alternative than solid organ transplants, fundamental change in how the field is regulated and advanced is needed. Greater public and private investment in the development of cellular transplantation is required. Furthermore, consistent with the call of multiple national transplant societies for allo-islet transplants, the oversight of cellular transplants should mirror that of solid organ transplants and not be classified under the unsustainable, outdated model that requires licensing as a drug with the Food and Drug Administration. Cellular transplantation has the potential to bring profound benefit through progress in bioengineering and regenerative medicine, limiting immunosuppression-related toxicity, and providing markedly reduced surgical morbidity.
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Affiliation(s)
- Ronald F Parsons
- Department of Surgery, Emory Transplant Center, Emory University School of Medicine, Atlanta, Georgia
| | - Angeles Baquerizo
- Scripps Center for Cell and Organ Transplantation, La Jolla, California
| | - Varvara A Kirchner
- Division of Transplantation, Department of Surgery, University of Minnesota, Minneapolis, Minnesota
| | - Sayeed Malek
- Division of Transplant Surgery, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts
| | - Chirag S Desai
- Division of Transplantation, Department of Surgery, University of North Carolina, Chapel Hill, North Carolina
| | - Austin Schenk
- Division of Transplantation, Department of Surgery, Ohio State University, Columbus, Ohio
| | - Erik B Finger
- Division of Transplantation, Department of Surgery, University of Minnesota, Minneapolis, Minnesota
| | - Todd V Brennan
- Department of Surgery, Comprehensive Transplant Center, Cedars-Sinai Medical Center, Los Angeles, California
| | - Kalpaj R Parekh
- Division of Cardiothoracic Surgery, Department of Surgery, Carver College of Medicine, University of Iowa, Iowa City, Iowa
| | - Malcolm MacConmara
- Division of Surgical Transplantation, Department of Surgery, University of Texas Southwestern Medical Center, Dallas, Texas
| | - Kenneth Brayman
- Division of Transplantation, Department of Surgery, University of Virginia, Charlottesville, Virginia
| | - Jeffrey Fair
- Division of Transplant Surgery, Department of Surgery, University of Texas Medical Branch, Galveston, Texas
| | - Jason A Wertheim
- Departments of Surgery and Biomedical Engineering, University of Arizona Health Sciences, Tucson, Arizona
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15
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Bram Y, Nguyen DHT, Gupta V, Park J, Richardson C, Chandar V, Schwartz RE. Cell and Tissue Therapy for the Treatment of Chronic Liver Disease. Annu Rev Biomed Eng 2021; 23:517-546. [PMID: 33974812 PMCID: PMC8864721 DOI: 10.1146/annurev-bioeng-112619-044026] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/04/2022]
Abstract
Liver disease is an important clinical problem, impacting 600 million people worldwide. It is the 11th-leading cause of death in the world. Despite constant improvement in treatment and diagnostics, the aging population and accumulated risk factors led to increased morbidity due to nonalcoholic fatty liver disease and steatohepatitis. Liver transplantation, first established in the 1960s, is the second-most-common solid organ transplantation and is the gold standard for the treatment of liver failure. However, less than 10% of the global need for liver transplantation is met at the current rates of transplantation due to the paucity of available organs. Cell- and tissue-based therapies present an alternative to organ transplantation. This review surveys the approaches and tools that have been developed, discusses the distinctive challenges that exist for cell- and tissue-based therapies, and examines the future directions of regenerative therapies for the treatment of liver disease.
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Affiliation(s)
- Yaron Bram
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA;
| | - Duc-Huy T Nguyen
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA;
| | - Vikas Gupta
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA;
| | - Jiwoon Park
- Department of Physiology, Biophysics and Systems Biology, Weill Cornell Medical College, New York, NY 10065, USA
| | - Chanel Richardson
- Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA
| | - Vasuretha Chandar
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA;
| | - Robert E Schwartz
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA; .,Department of Physiology, Biophysics and Systems Biology, Weill Cornell Medical College, New York, NY 10065, USA
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16
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Demaret T, Evraerts J, Ravau J, Roumain M, Muccioli GG, Najimi M, Sokal EM. High Dose Versus Low Dose Syngeneic Hepatocyte Transplantation in Pex1-G844D NMRI Mouse Model is Safe but Does Not Achieve Long Term Engraftment. Cells 2020; 10:cells10010040. [PMID: 33396635 PMCID: PMC7823729 DOI: 10.3390/cells10010040] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2020] [Revised: 12/21/2020] [Accepted: 12/23/2020] [Indexed: 12/17/2022] Open
Abstract
Genetic alterations in PEX genes lead to peroxisome biogenesis disorder. In humans, they are associated with Zellweger spectrum disorders (ZSD). No validated treatment has been shown to modify the dismal natural history of ZSD. Liver transplantation (LT) improved clinical and biochemical outcomes in mild ZSD patients. Hepatocyte transplantation (HT), developed to overcome LT limitations, was performed in a mild ZSD 4-year-old child with encouraging short-term results. Here, we evaluated low dose (12.5 million hepatocytes/kg) and high dose (50 million hepatocytes/kg) syngeneic male HT via intrasplenic infusion in the Pex1-G844D NMRI mouse model which recapitulates a mild ZSD phenotype. HT was feasible and safe in growth retarded ZSD mice. Clinical (weight and food intake) and biochemical parameters (very long-chain fatty acids, abnormal bile acids, etc.) were in accordance with ZSD phenotype but they were not robustly modified by HT. As expected, one third of the infused cells were detected in the liver 24 h post-HT. No liver nor spleen microchimerism was detected after 7, 14 and 30 days. Future optimizations are required to improve hepatocyte engraftment in Pex1-G844D NMRI mouse liver. The mouse model exhibited the robustness required for ZSD liver-targeted therapies evaluation.
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Affiliation(s)
- Tanguy Demaret
- Laboratoire d’Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique (IREC), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (J.E.); (J.R.); (M.N.); (E.M.S.)
- Correspondence:
| | - Jonathan Evraerts
- Laboratoire d’Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique (IREC), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (J.E.); (J.R.); (M.N.); (E.M.S.)
| | - Joachim Ravau
- Laboratoire d’Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique (IREC), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (J.E.); (J.R.); (M.N.); (E.M.S.)
| | - Martin Roumain
- Bioanalysis and Pharmacology of Bioactive Lipids Research Group (BPBL), Louvain Drug Research Institute (LDRI), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (M.R.); (G.G.M.)
| | - Giulio G. Muccioli
- Bioanalysis and Pharmacology of Bioactive Lipids Research Group (BPBL), Louvain Drug Research Institute (LDRI), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (M.R.); (G.G.M.)
| | - Mustapha Najimi
- Laboratoire d’Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique (IREC), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (J.E.); (J.R.); (M.N.); (E.M.S.)
| | - Etienne M. Sokal
- Laboratoire d’Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique (IREC), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (J.E.); (J.R.); (M.N.); (E.M.S.)
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17
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Ultrasound-guided in Utero Transplantation of Placental Stem Cells into the Liver of Crigler-Najjar Syndrome Model Rat. Transplantation 2020; 103:e182-e187. [PMID: 30985583 DOI: 10.1097/tp.0000000000002735] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
BACKGROUND Advances in prenatal screening and early diagnosis of genetic disease will potentially allow for preemptive treatment of anticipated postnatal disease by in utero cell transplantation (IUCT). This strategy carries potential benefits over postnatal treatment, which might allow for improved engraftment and function of the transplanted cells. Congenital metabolic disorders may be an ideal target for this type of therapy, as in most cases, they require replacement of a single deficient hepatic enzyme, and multiple small-animal models exist for preclinical testing. METHODS The Gunn rat, a Crigler-Najjar syndrome model animal lacking UDP-glucuronosyltransferase (UGT1A1), was used as recipient. Human amniotic epithelial cells (hAECs), which possess hepatic differentiation potential, were transplanted into the midgestation fetal Gunn rat liver via ultrasound-guided IUCT. The impact of IUCT on live birth and postnatal survival was evaluated. Human cell engraftment was immunohistochemically analyzed on postnatal day 21. RESULTS Ultrasound-guided IUCT was conducted in rat fetuses on embryonic day 16. Following IUCT, the antihuman mitochondria-positive cells were detected in the liver of recipient rats at postnatal day 21. CONCLUSIONS Here, we have introduced ultrasound-guided IUCT of hAEC using a small-animal model of a congenital metabolic disorder without immunosuppression. The immunological advantage of IUCT was demonstrated with xenogeneic IUCT. This procedure is suitable to conduct preclinical studies for exploring the feasibility and efficacy of ultrasound-guided transuterine cell injection using rodent disease models.
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18
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Cotovio JP, Fernandes TG. Production of Human Pluripotent Stem Cell-Derived Hepatic Cell Lineages and Liver Organoids: Current Status and Potential Applications. Bioengineering (Basel) 2020; 7:E36. [PMID: 32283585 PMCID: PMC7356351 DOI: 10.3390/bioengineering7020036] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2020] [Revised: 04/03/2020] [Accepted: 04/07/2020] [Indexed: 02/06/2023] Open
Abstract
Liver disease is one of the leading causes of death worldwide, leading to the death of approximately 2 million people per year. Current therapies include orthotopic liver transplantation, however, donor organ shortage remains a great challenge. In addition, the development of novel therapeutics has been limited due to the lack of in vitro models that mimic in vivo liver physiology. Accordingly, hepatic cell lineages derived from human pluripotent stem cells (hPSCs) represent a promising cell source for liver cell therapy, disease modelling, and drug discovery. Moreover, the development of new culture systems bringing together the multiple liver-specific hepatic cell types triggered the development of hPSC-derived liver organoids. Therefore, these human liver-based platforms hold great potential for clinical applications. In this review, the production of the different hepatic cell lineages from hPSCs, including hepatocytes, as well as the emerging strategies to generate hPSC-derived liver organoids will be assessed, while current biomedical applications will be highlighted.
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Affiliation(s)
| | - Tiago G. Fernandes
- iBB-Institute for Bioengineering and Biosciences and Department of Bioengineering, Instituto Superior Técnico, Universidade de Lisboa, Av. Rovisco Pais, 1049-001 Lisbon, Portugal;
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19
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Nguyen MP, Jain V, Iansante V, Mitry RR, Filippi C, Dhawan A. Clinical application of hepatocyte transplantation: current status, applicability, limitations, and future outlook. Expert Rev Gastroenterol Hepatol 2020; 14:185-196. [PMID: 32098516 DOI: 10.1080/17474124.2020.1733975] [Citation(s) in RCA: 29] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/25/2022]
Abstract
Introduction: Hepatocyte transplantation (HT) is a promising alternative to liver transplantation for the treatment of liver-based metabolic diseases and acute liver failure (ALF). However, shortage of good-quality liver tissues, early cell loss post-infusion, reduced cell engraftment and function restricts clinical application.Areas covered: A comprehensive literature search was performed to cover pre-clinical and clinical HT studies. The review discusses the latest developments to address HT limitations: cell sources from marginal/suboptimal donors to neonatal livers, differentiating pluripotent stem cells into hepatocyte-like cells, in vitro expansion, prevention of immune response to transplanted cells by encapsulation or using innate immunity-inhibiting agents, and enhancing engraftment through partial hepatectomy or irradiation.Expert opinion: To date, published data are highly encouraging specially the alginate-encapsulated hepatocyte treatment of children with ALF. Hepatocyte functions can be further improved through co-culturing with mesenchymal stromal cells. Moreover, ex-vivo genetic correction will enable the use of autologous cells in future personalized medicine.
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Affiliation(s)
- Minh Phuong Nguyen
- Dhawan Lab. at the Mowat Labs, Institute of Liver Studies, King's College Hospital, London, United Kingdom
| | - Vandana Jain
- Dhawan Lab. at the Mowat Labs, Institute of Liver Studies, King's College Hospital, London, United Kingdom
| | - Valeria Iansante
- Dhawan Lab. at the Mowat Labs, Institute of Liver Studies, King's College Hospital, London, United Kingdom
| | - Ragai R Mitry
- Dhawan Lab. at the Mowat Labs, Institute of Liver Studies, King's College Hospital, London, United Kingdom
| | - Celine Filippi
- Dhawan Lab. at the Mowat Labs, Institute of Liver Studies, King's College Hospital, London, United Kingdom
| | - Anil Dhawan
- Dhawan Lab. at the Mowat Labs, Institute of Liver Studies, King's College Hospital, London, United Kingdom
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20
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Mesenchymal Stem Cells in the Adult Human Liver: Hype or Hope? Cells 2019; 8:cells8101127. [PMID: 31546729 PMCID: PMC6830330 DOI: 10.3390/cells8101127] [Citation(s) in RCA: 32] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2019] [Revised: 09/18/2019] [Accepted: 09/21/2019] [Indexed: 02/06/2023] Open
Abstract
Chronic liver diseases constitute a significant economic, social, and biomedical burden. Among commonly adopted approaches, only organ transplantation can radically help patients with end-stage liver pathologies. Cell therapy with hepatocytes as a treatment for chronic liver disease has demonstrated promising results. However, quality human hepatocytes are in short supply. Stem/progenitor cells capable of differentiating into functionally active hepatocytes provide an attractive alternative approach to cell therapy for liver diseases, as well as to liver-tissue engineering, drug screening, and basic research. The application of methods generally used to isolate mesenchymal stem cells (MSCs) and maintain them in culture to human liver tissue provides cells, designated here as liver MSCs. They have much in common with MSCs from other tissues, but differ in two aspects-expression of a range of hepatocyte-specific genes and, possibly, inherent commitment to hepatogenic differentiation. The aim of this review is to analyze data regarding liver MSCs, probably another type of liver stem/progenitor cells different from hepatic stellate cells or so-called hepatic progenitor cells. The review presents an analysis of the phenotypic characteristics of liver MSCs, their differentiation and therapeutic potential, methods for isolating these cells from human liver, and discusses issues of their origin and heterogeneity. Human liver MSCs are a fascinating object of fundamental research with a potential for important practical applications.
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21
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Clinical hepatocyte transplantation. GASTROENTEROLOGIA Y HEPATOLOGIA 2019; 42:202-208. [DOI: 10.1016/j.gastrohep.2018.10.007] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/23/2018] [Revised: 09/21/2018] [Accepted: 10/10/2018] [Indexed: 12/18/2022]
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22
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Willadsen M, Chaise M, Yarovoy I, Zhang AQ, Parashurama N. Engineering molecular imaging strategies for regenerative medicine. Bioeng Transl Med 2018; 3:232-255. [PMID: 30377663 PMCID: PMC6195904 DOI: 10.1002/btm2.10114] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/10/2018] [Revised: 08/30/2018] [Accepted: 09/01/2018] [Indexed: 12/15/2022] Open
Abstract
The reshaping of the world's aging population has created an urgent need for therapies for chronic diseases. Regenerative medicine offers a ray of hope, and its complex solutions include material, cellular, or tissue systems. We review basics of regenerative medicine/stem cells and describe how the field of molecular imaging, which is based on quantitative, noninvasive, imaging of biological events in living subjects, can be applied to regenerative medicine in order to interrogate tissues in innovative, informative, and personalized ways. We consider aspects of regenerative medicine for which molecular imaging will benefit. Next, genetic and nanoparticle-based cell imaging strategies are discussed in detail, with modalities like magnetic resonance imaging, optical imaging (near infra-red, bioluminescence), raman microscopy, and photoacoustic microscopy), ultrasound, computed tomography, single-photon computed tomography, and positron emission tomography. We conclude with a discussion of "next generation" molecular imaging strategies, including imaging host tissues prior to cell/tissue transplantation.
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Affiliation(s)
- Matthew Willadsen
- Department of Chemical and Biological Engineering University at Buffalo, State University of New York, Furnas Hall Buffalo New York 14228
| | - Marc Chaise
- Jacobs School of Medicine and Biomedical Sciences University at Buffalo State University of New York 955 Main St., Buffalo, New York 14203
| | - Iven Yarovoy
- Department of Chemical and Biological Engineering University at Buffalo, State University of New York, Furnas Hall Buffalo New York 14228
| | - An Qi Zhang
- Department of Chemical and Biological Engineering University at Buffalo, State University of New York, Furnas Hall Buffalo New York 14228
| | - Natesh Parashurama
- Department of Chemical and Biological Engineering University at Buffalo, State University of New York, Furnas Hall Buffalo New York 14228.,Department of Biomedical Engineering University at Buffalo, State University of New York, Bonner Hall Buffalo New York 14228.,Clinical and Translation Research Center (CTRC) University at Buffalo, State University of New York 875 Ellicott St., Buffalo, New York 14203
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23
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Iansante V, Mitry RR, Filippi C, Fitzpatrick E, Dhawan A. Human hepatocyte transplantation for liver disease: current status and future perspectives. Pediatr Res 2018; 83:232-240. [PMID: 29149103 DOI: 10.1038/pr.2017.284] [Citation(s) in RCA: 150] [Impact Index Per Article: 21.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/15/2017] [Accepted: 10/02/2017] [Indexed: 12/16/2022]
Abstract
Liver transplantation is the accepted treatment for patients with acute liver failure and liver-based metabolic disorders. However, donor organ shortage and lifelong need for immunosuppression are the main limitations to liver transplantation. In addition, loss of the native liver as a target organ for future gene therapy for metabolic disorders limits the futuristic treatment options, resulting in the need for alternative therapeutic strategies. A potential alternative to liver transplantation is allogeneic hepatocyte transplantation. Over the last two decades, hepatocyte transplantation has made the transition from bench to bedside. Standardized techniques have been established for isolation, culture, and cryopreservation of human hepatocytes. Clinical hepatocyte transplantation safety and short-term efficacy have been proven; however, some major hurdles-mainly concerning shortage of donor organs, low cell engraftment, and lack of a long-lasting effect-need to be overcome to widen its clinical applications. Current research is aimed at addressing these problems, with the ultimate goal of increasing hepatocyte transplantation efficacy in clinical applications.
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Affiliation(s)
- V Iansante
- DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
| | - R R Mitry
- DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
| | - C Filippi
- DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
| | - E Fitzpatrick
- DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
| | - A Dhawan
- DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
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24
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Heath RD, Ertem F, Romana BS, Ibdah JA, Tahan V. Hepatocyte transplantation: Consider infusion before incision. World J Transplant 2017; 7:317-323. [PMID: 29312860 PMCID: PMC5743868 DOI: 10.5500/wjt.v7.i6.317] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/28/2017] [Revised: 11/27/2017] [Accepted: 12/05/2017] [Indexed: 02/05/2023] Open
Abstract
Human hepatocyte transplantation is undergoing study as a bridge, or even alternative, to orthotopic liver transplantation (OLT). This technique has undergone multiple developments over the past thirty years in terms of mode of delivery, source and preparation of cell cultures, monitoring of graft function, and use of immunosuppression. Further refinements and improvements in these techniques will likely allow improved graft survival and function, granting patients higher yield from this technique and potentially significantly delaying need for OLT.
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Affiliation(s)
- Ryan D Heath
- Division of Gastroenterology and Hepatology, University of Missouri, Columbia, MO 65212, United States
| | - Furkan Ertem
- Department of Internal Medicine, University of Pittsburgh Medical Center, Pittsburgh, PA 15260, United States
| | - Bhupinder S Romana
- Division of Gastroenterology and Hepatology, University of Missouri, Columbia, MO 65212, United States
| | - Jamal A Ibdah
- Division of Gastroenterology and Hepatology, University of Missouri, Columbia, MO 65212, United States
| | - Veysel Tahan
- Division of Gastroenterology and Hepatology, University of Missouri, Columbia, MO 65212, United States
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25
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Hickey RD, Mao SA, Glorioso J, Elgilani F, Amiot B, Chen H, Rinaldo P, Marler R, Jiang H, DeGrado TR, Suksanpaisan L, O'Connor MK, Freeman BL, Ibrahim SH, Peng KW, Harding CO, Ho CS, Grompe M, Ikeda Y, Lillegard JB, Russell SJ, Nyberg SL. Curative ex vivo liver-directed gene therapy in a pig model of hereditary tyrosinemia type 1. Sci Transl Med 2017; 8:349ra99. [PMID: 27464750 DOI: 10.1126/scitranslmed.aaf3838] [Citation(s) in RCA: 48] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/04/2016] [Accepted: 07/05/2016] [Indexed: 12/23/2022]
Abstract
We tested the hypothesis that ex vivo hepatocyte gene therapy can correct the metabolic disorder in fumarylacetoacetate hydrolase-deficient (Fah(-/-)) pigs, a large animal model of hereditary tyrosinemia type 1 (HT1). Recipient Fah(-/-) pigs underwent partial liver resection and hepatocyte isolation by collagenase digestion. Hepatocytes were transduced with one or both of the lentiviral vectors expressing the therapeutic Fah and the reporter sodium-iodide symporter (Nis) genes under control of the thyroxine-binding globulin promoter. Pigs received autologous transplants of hepatocytes by portal vein infusion. After transplantation, the protective drug 2-(2-nitro-4-trifluoromethylbenzyol)-1,3 cyclohexanedione (NTBC) was withheld from recipient pigs to provide a selective advantage for expansion of corrected FAH(+) cells. Proliferation of transplanted cells, assessed by both immunohistochemistry and noninvasive positron emission tomography imaging of NIS-labeled cells, demonstrated near-complete liver repopulation by gene-corrected cells. Tyrosine and succinylacetone levels improved to within normal range, demonstrating complete correction of tyrosine metabolism. In addition, repopulation of the Fah(-/-) liver with transplanted cells inhibited the onset of severe fibrosis, a characteristic of nontransplanted Fah(-/-) pigs. This study demonstrates correction of disease in a pig model of metabolic liver disease by ex vivo gene therapy. To date, ex vivo gene therapy has only been successful in small animal models. We conclude that further exploration of ex vivo hepatocyte genetic correction is warranted for clinical use.
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Affiliation(s)
- Raymond D Hickey
- Department of Surgery, Mayo Clinic, Rochester, MN 55905, USA. Department of Molecular Medicine, Mayo Clinic, Rochester, MN 55905, USA.
| | - Shennen A Mao
- Department of Surgery, Mayo Clinic, Rochester, MN 55905, USA
| | - Jaime Glorioso
- Department of Surgery, Mayo Clinic, Rochester, MN 55905, USA
| | - Faysal Elgilani
- Department of Surgery, Mayo Clinic, Rochester, MN 55905, USA
| | - Bruce Amiot
- Brami Biomedical Inc., Coon Rapids, MN 55433, USA
| | - Harvey Chen
- Department of Surgery, Mayo Clinic, Rochester, MN 55905, USA
| | - Piero Rinaldo
- Division of Laboratory Genetics, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN 55905, USA
| | - Ronald Marler
- Department of Comparative Medicine, Mayo Clinic, Scottsdale, AZ 85259, USA
| | - Huailei Jiang
- Department of Radiology, Mayo Clinic, Rochester, MN 55905, USA
| | | | - Lukkana Suksanpaisan
- Department of Radiology, Mayo Clinic, Rochester, MN 55905, USA. Imanis Life Sciences, Rochester, MN 55902, USA
| | | | - Brittany L Freeman
- Division of Pediatric Gastroenterology, Mayo Clinic, Rochester, MN 55905, USA
| | - Samar H Ibrahim
- Division of Pediatric Gastroenterology, Mayo Clinic, Rochester, MN 55905, USA
| | - Kah Whye Peng
- Department of Molecular Medicine, Mayo Clinic, Rochester, MN 55905, USA
| | - Cary O Harding
- Department of Molecular and Medical Genetics and Department of Pediatrics, Oregon Health and Science University, Portland, OR 97239, USA
| | - Chak-Sum Ho
- Histocompatibility Laboratory, Gift of Life Michigan, Ann Arbor, MI 48108, USA
| | - Markus Grompe
- Papé Family Pediatric Research Institute, Oregon Health and Science University, Portland, OR 97239, USA
| | - Yasuhiro Ikeda
- Department of Molecular Medicine, Mayo Clinic, Rochester, MN 55905, USA
| | - Joseph B Lillegard
- Department of Surgery, Mayo Clinic, Rochester, MN 55905, USA. Midwest Fetal Care Center, Children's Hospitals and Clinics of Minnesota, Minneapolis, MN 55404, USA
| | - Stephen J Russell
- Department of Molecular Medicine, Mayo Clinic, Rochester, MN 55905, USA
| | - Scott L Nyberg
- Department of Surgery, Mayo Clinic, Rochester, MN 55905, USA
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26
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Ogoke O, Oluwole J, Parashurama N. Bioengineering considerations in liver regenerative medicine. J Biol Eng 2017; 11:46. [PMID: 29204185 PMCID: PMC5702480 DOI: 10.1186/s13036-017-0081-4] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2017] [Accepted: 09/25/2017] [Indexed: 12/19/2022] Open
Abstract
Background Liver disease contributes significantly to global disease burden and is associated with rising incidence and escalating costs. It is likely that innovative approaches, arising from the emerging field of liver regenerative medicine, will counter these trends. Main body Liver regenerative medicine is a rapidly expanding field based on a rich history of basic investigations into the nature of liver structure, physiology, development, regeneration, and function. With a bioengineering perspective, we discuss all major subfields within liver regenerative medicine, focusing on the history, seminal publications, recent progress within these fields, and commercialization efforts. The areas reviewed include fundamental aspects of liver transplantation, liver regeneration, primary hepatocyte cell culture, bioartificial liver, hepatocyte transplantation and liver cell therapies, mouse liver repopulation, adult liver stem cell/progenitor cells, pluripotent stem cells, hepatic microdevices, and decellularized liver grafts. Conclusion These studies highlight the creative directions of liver regenerative medicine, the collective efforts of scientists, engineers, and doctors, and the bright outlook for a wide range of approaches and applications which will impact patients with liver disease.
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Affiliation(s)
- Ogechi Ogoke
- Department of Chemical and Biological Engineering, University at Buffalo (State University of New York), Furnas Hall, Buffalo, NY 14260 USA.,Clinical and Translation Research Center (CTRC), University at Buffalo (State University of New York), 875 Ellicott St., Buffalo, NY 14203 USA
| | - Janet Oluwole
- Clinical and Translation Research Center (CTRC), University at Buffalo (State University of New York), 875 Ellicott St., Buffalo, NY 14203 USA.,Department of Biomedical Engineering, University at Buffalo (State University of New York), Furnas Hall, 907 Furnas Hall, Buffalo, NY 14260 USA
| | - Natesh Parashurama
- Department of Chemical and Biological Engineering, University at Buffalo (State University of New York), Furnas Hall, Buffalo, NY 14260 USA.,Clinical and Translation Research Center (CTRC), University at Buffalo (State University of New York), 875 Ellicott St., Buffalo, NY 14203 USA.,Department of Biomedical Engineering, University at Buffalo (State University of New York), Furnas Hall, 907 Furnas Hall, Buffalo, NY 14260 USA
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27
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Hughes RD, Mitry RR, Dhawan A. Hepatocyte Transplantation for Metabolic Liver Disease: UK Experience. J R Soc Med 2017; 98:341-5. [PMID: 16055896 PMCID: PMC1181831 DOI: 10.1177/014107680509800803] [Citation(s) in RCA: 29] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022] Open
Affiliation(s)
- Robin D Hughes
- Institute of Liver Studies, King's College London & King's College Hospital, London, UK
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28
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In vivo and ex vivo methods of growing a liver bud through tissue connection. Sci Rep 2017; 7:14085. [PMID: 29074999 PMCID: PMC5658340 DOI: 10.1038/s41598-017-14542-2] [Citation(s) in RCA: 67] [Impact Index Per Article: 8.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2017] [Accepted: 10/12/2017] [Indexed: 12/28/2022] Open
Abstract
Cell-based therapy has been proposed as an alternative to orthotopic liver transplantation. The novel transplantation of an in vitro-generated liver bud might have therapeutic potential. In vivo and ex vivo methods for growing a liver bud are essential for paving the way for the clinical translation of liver bud transplantation. We herein report a novel transplantation method for liver buds that are grown in vivo involving orthotopic transplantation on the transected parenchyma of the liver, which showed long engraftment and marked growth in comparison to heterotopic transplantation. Furthermore, this study demonstrates a method for rapidly fabricating scalable liver-like tissue by fusing hundreds of liver bud-like spheroids using a 3D bioprinter. Its system to fix the shape of the 3D tissue with the needle-array system enabled the fabrication of elaborate geometry and the immediate execution of culture circulation after 3D printing—thereby avoiding an ischemic environment ex vivo. The ex vivo-fabricated human liver-like tissue exhibited self-tissue organization ex vivo and engraftment on the liver of nude rats. These achievements conclusively show both in vivo and ex vivo methods for growing in vitro-generated liver buds. These methods provide a new approach for in vitro-generated liver organoids transplantation.
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29
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Spinelli SV, Rodríguez JV, Quintana AB, Mediavilla MG, Guibert EE. Engraftment and Function of Intrasplenically Transplanted Cold Stored Rat Hepatocytes. Cell Transplant 2017. [DOI: 10.3727/096020198389889] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/28/2023] Open
Abstract
Hepatocellular transplant may potentially be efficacious for the treatment of selected liver metabolic disorders and acute hepatic failure. On the other hand, the use of hepatocyte cold preservation techniques in these transplantation protocols would allow to have available cells at the right time and place and, consequently, make an optimal use of scarce human hepatocytes. In our experiments we evaluated the biodistribution and functionality of cold preserved hepatocytes transplanted in the spleen of syngeneic rats. Isolated hepatocytes were labeled with the fluorescent dye 5(6)-carboxyfluorescein diacetate succinimidyl-ester, cold-preserved in modified University of Wisconsin (UW) solution for 48 or 96 h, and then transplanted into the spleen. Recipient animals were euthanized at 0 and 3 h, and at 1, 2, 3, 5, 10, and 14 days after transplantation for tissue analysis. Labeled hepatocytes were clearly identifiable in the recipient tissues up to 14 days later. Fluorescence microscopy also showed no significant differences in biodistribution when either cold stored or freshly isolated hepatocytes were transplanted. In addition, functional activity of transplanted cells was demonstrated by immunohistochemical detection of albumin at levels comparable to those found in normal hepatocytes. Our findings establish that cold preserved hepatocytes appear morphologically and biochemically normal after intrasplenic transplantation. Consequently, it indicates that modified UW solution makes it possible to safety preserve hepatocytes for up to 96 h before transplantation, perhaps providing sufficient time for hepatocyte allocation and potential recipient preparation, if applicable clinically.
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Affiliation(s)
- Silvana V. Spinelli
- Biología Molecular, Dto. Cs. Biológicas, Facultad de Cs. Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario
| | - Joaquín V. Rodríguez
- Farmacología, Dto. Cs. Fisiológicas, Facultad de Cs. Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario
| | - Alejandra B. Quintana
- Morfología, Dto. Cs. Biológicas, Facultad de Cs. Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario
| | - María G. Mediavilla
- Biología Molecular, Dto. Cs. Biológicas, Facultad de Cs. Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario
| | - Edgardo E. Guibert
- Biología Molecular, Dto. Cs. Biológicas, Facultad de Cs. Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario
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30
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Fujino M, Li XK, Kitazawa Y, Funeshima N, Guo L, Okuyama T, Amano T, Amemiya H, Suzuki S. Selective Repopulation of Mice Liver after Fas-Resistant Hepatocyte Transplantation. Cell Transplant 2017. [DOI: 10.3727/000000001783986701] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
Hepatocyte transplantation has been proposed as a potential therapeutic method to treat irreversible liver failure and inherited hepatic disorders, although transplanted cells do not easily reconstruct the liver tissue under intact conditions. This study was aimed at modulating the recipient liver conditions to promote repopulation of the liver after hepatocyte transplantation. Hepatocytes isolated from male MRL-lpr/lpr (lpr) mice with a mutation of Fas antigen were transplanted in a number of 1 × 106 cells in female MRL-+/+ (wildtype mice) by intrasplenic injection. An agonistic anti-Fas antibody (0.15 mg/kg) was administered intravenously 24 h after cell transplantation. We also administrated the antibody at 0.3 mg/kg 1 week after grafting and at 0.6 mg/kg 2 weeks after transplantation. The liver specimens were taken at different time intervals for histological examination. The reconstructed male lpr hepatocytes in the female wild-type mice were determined by a real-time quantitative PCR assay using the primers and probe for the sry gene. The pathologic findings of the recipient livers after treatment with anti-Fas antibody revealed a large number of apoptotic hepatocytes. The grafted lpr hepatocytes were observed to reconstruct as much as 6.9% of the recipient liver in the anti-Fas antibody-treated group 3 months after transplantation. In contrast, we observed the transplanted cells at lower than 0.1% in the nontreated livers. These findings demonstrated that repeated induction of apoptosis in recipient hepatocytes shifts the environment of the liver to a regenerative condition. This method may be useful to promote the reconstruction of transplanted hepatocytes in a recipient liver.
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Affiliation(s)
- Masayuki Fujino
- Department of Experimental Surgery and Bioengineering, Tokyo, Japan
- Department of Zootechnical Science, Tokyo University of Agriculture, Tokyo, Japan
| | - Xiao-Kang Li
- Department of Experimental Surgery and Bioengineering, Tokyo, Japan
| | - Yusuke Kitazawa
- Department of Experimental Surgery and Bioengineering, Tokyo, Japan
| | - Naoko Funeshima
- Department of Experimental Surgery and Bioengineering, Tokyo, Japan
| | - Lei Guo
- Department of Experimental Surgery and Bioengineering, Tokyo, Japan
| | - Torayuki Okuyama
- Genetics, National Children's Medical Research Center, Tokyo, Japan
| | - Takashi Amano
- Department of Zootechnical Science, Tokyo University of Agriculture, Tokyo, Japan
| | - Hiroshi Amemiya
- Department of Experimental Surgery and Bioengineering, Tokyo, Japan
| | - Seiichi Suzuki
- Department of Experimental Surgery and Bioengineering, Tokyo, Japan
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31
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Moscioni AD, Rozga J, Chen S, Naim A, Scott HS, Demetriou AA. Long-Term Correction of Albumin Levels in the Nagase Analbuminemic Rat: Repopulation of the Liver by Transplanted Normal Hepatocytes under a Regeneration Response. Cell Transplant 2017; 5:499-503. [PMID: 8800518 DOI: 10.1177/096368979600500409] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022] Open
Abstract
Numerous studies have reported successful transplantation of hepatocytes with demonstration of function. However, none have shown long-term correction of a liver-related metabolic defect. Male Nagase analbuminemic rats, immunosuppressed with cyclosporin-A, were transplanted with normal hepatocytes (2 × 107 cells/rat) isolated from allogeneic male Sprague–Dawley rat donors. Hepatocytes were selectively transplanted via the portal vein tributary into the posterior liver lobes of Nagase analbuminemic rats. Following 2 wk, to allow engraftment, selected transplanted rats (Group I) were reoperated and the portal venous branch supplying the anterior liver lobes was permanently ligated, resulting in their atrophy and induction of regeneration in the residual transplant-bearing lobes. Control rats consisted of: Group II—transplanted with normal hepatocytes without portal branch ligation; Group III—transplanted with analbuminemic hepatocytes with portal branch ligation; and Group IV—nontransplanted analbuminemic rats with portal branch ligation. The experimental period extended to 3 mo posttransplantation. All rats transplanted with normal hepatocytes demonstrated a significant elevation in serum albumin levels (ELISA). Group I rats had dramatic elevations in serum albumin to near normal levels (1.78 ± 0.20 g/dl), and maintained these levels until the end of the experiment Albumin levels in Group II rats reached 0.26 ± 0.07 g/dl (p < 0.001), whereas Group III and IV rats showed no changes in serum albumin levels throughout the experiment Immunohistology of liver tissue obtained from Group I rats, demonstrated large numbers (22.6 ± 7.5%) of albumin-positive hepatocytes populating the recipient liver. This is the first report of near-total and sustained correction of a genetic defect in liver function in an experimental animal model following allogeneic hepatocyte transplantation.
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Affiliation(s)
- A D Moscioni
- Department of Surgery, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
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32
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Arkadopoulos N, Chen SC, Khalili TM, Detry O, Hewitt WR, Lilja H, Kamachi H, Petrovic L, Mullon CJ, Demetriou AA, Rozga J. Transplantation of Hepatocytes for Prevention of Intracranial Hypertension in Pigs with Ischemic Liver Failure. Cell Transplant 2017; 7:357-63. [PMID: 9710304 DOI: 10.1177/096368979800700403] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022] Open
Abstract
Intracranial hypertension leading to brain stem herniation is a major cause of death in fulminant hepatic failure (FHF). Mannitol, barbiturates, and hyperventilation have been used to treat brain swelling, but most patients are either refractory to medical management or cannot be treated because of concurrent medical problems or side effects. In this study, we examined whether allogeneic hepatocellular transplantation may prevent development of intracranial hypertension in pigs with experimentally induced liver failure. Of the two preparations tested—total hepatectomy (n = 47), and liver devascularization (n = 16)—only pigs with liver ischemia developed brain edema provided, however, that animals were maintained normothermic throughout the postoperative period. This model was then used in transplantation studies, in which six pigs received intrasplenic injection of allogeneic hepatocytes (2.5 × 109 cells/pig) and 3 days later acute liver failure was induced. In both models (anhepatic state, liver devascularization), pigs allowed to become hypothermic had significantly longer survival compared to those maintained normothermic. Normothermic pigs with liver ischemia had, at all time points studied, ICP greater than 20 mmHg. Pigs that received hepatocellular transplants had ICP below 15 mmHg until death; at the same time, cerebral perfusion pressure (CPP) in transplanted pigs was consistently higher than in controls (45 ± 11 mmHg vs. 16 ± 18 mmHg; p < 0.05). Spleens of transplanted pigs contained clusters of viable hepatocytes (hematoxylin-eosin, CAM 5.2). It was concluded that removal of the liver does not result in intracranial hypertension; hypothermia prolongs survival time in both anhepatic pigs and pigs with liver devascularization, and intrasplenic transplantation of allogeneic hepatocytes prevents development of intracranial hypertension in pigs with acute ischemic liver failure.
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Affiliation(s)
- N Arkadopoulos
- Department of Surgery, Allen and Burns Research Institute, Cedars-Sinai Medical Center, UCLA School of Medicine, Los Angeles, CA 90048, USA
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Abstract
Recent advances in liver transplantation have caused a serious shortage of donor livers, and a consensus on determining brain death has not been reached by the medical community in several countries, including Japan. To overcome these circumstances, hepatocellular transplantation (HCTX) has been attempted because HCTX requires no vascular anastomosis and donor hepatocytes are easy to obtain from living donors, and easy to preserve for a long time. In many experimental studies on HCTX some promising findings have been obtained, but clinically there has been little progress. Our success with survival of autotransplanted monkey hepatocytes and the development of a preparation of human hepatocytes has renewed interest in clinical HCTX. A multipuncture perfusion method or collagenase perfusion via the umbilical vein enables us to obtain approximately 1 × 108 hepatocytes from partially resected liver (60 g). The clinical trial of HCTX into the spleen was performed in 10 patients in Japan. A CT image, taken 1 mo after HCTX, showed a low density area corresponding to the inoculated site, which suggested that the transplanted hepatocytes survived and possessed hepatocellular function. One patient who sustained hepatic encephalopathy and massive ascites has now returned to work 11 mo after HCTX and hepatic artery ligation. However, there were no definite findings for functional support of damaged livers by HCTX in the spleen. We will review our experiments on HCTX, and describe the current and future aspects of HCTX.
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Affiliation(s)
- Michio Mito
- Department of Surgery, Asahikawa Medical College, 4-5, Nishi-kagura, Asahikawa, 078 Japan
| | - Mitsuo Kusano
- Department of Surgery, Asahikawa Medical College, 4-5, Nishi-kagura, Asahikawa, 078 Japan
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34
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Krishna Vanaja D, Sivakumar B, Jesudasan RA, Singh L, Janardanasarma MK, Habibullah CM. In Vivo Identification, Survival, and Functional Efficacy of Transplanted Hepatocytes in Acute Liver Failure Mice Model by Fish Using Y-Chromosome Probe. Cell Transplant 2017; 7:267-73. [PMID: 9647436 DOI: 10.1177/096368979800700305] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022] Open
Abstract
Hepatocyte transplantation has excited much interest in lending temporary metabolic support to a failing liver following acute liver injury. The exact site from which they act and the clinical, biochemical, and histological changes in the recipient body following hepatocyte transplantation is yet to be worked out. The present study is an attempt to delineate location and function of transplanted hepatocytes and also the overall survival of these cells with a fluorescent in situ hybridization (FISH) technique using a Y-chromosome–specific probe in a carbon tetrachloride (CCl4)-induced mice model of fulminant hepatic failure. Fifty-five syngenic adult Swiss female mice of approximately the same age and body weight were divided into three groups. Group-1 (n = 15), which received mineral oil, served as a negative control. Group-II (n = 15) received CCl4 (3 mL/kg) 40% vol/vol in mineral oil, by gavage served as positive control for hepatic failure. Group-III (n = 25) received intrasplenic transplantation of syngenic single cell suspension of hepatocytes in Hanks medium, after 30 h of CCl4 administration. Male Swiss adult mice (n = 15) served as donors of hepatocytes. The overall survival of animals in groups I to III was 100, 0, and 70%, respectively, by 2 wk of the study period. Transplanted hepatocytes were identified by Periodic Acid Schiff (PAS) staining and confirmed with a FISH technique using the Y-chromosome probe. The majority of exogenously transplanted hepatocytes were found in the liver and spleen sections even after 1 wk of hepatocyte transplantation. Transplanted cells were mostly found to be translocated into the sinusoids of the liver. Transplanted hepatocytes were found to be beneficial as a temporary liver support in a failing liver, significantly improving the survival of the animals. In the present study, the FISH technique was used to unequivocally distinguish the transplanted cells from the host, and thus describes a model for studying the distribution and survival of the transplanted cells.
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35
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Rozga J, Holzman M, Moscioni AD, Fujioka H, Morsiani E, Demetriou AA. Repeated Intraportal Hepatocyte Transplantation in Analbuminemic Rats. Cell Transplant 2017; 4:237-43. [PMID: 7773557 DOI: 10.1177/096368979500400207] [Citation(s) in RCA: 32] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022] Open
Abstract
The optimal site for implantation of isolated hepatocytes has not been established. We have developed a novel technique which allows repeated infusion of hepatocytes into the portal system via an indwelling catheter. Seven Nagase Analbuminemic rats (NAR) underwent single intra-portal infusion of 2 × 107 isolated normal albumin-producing rat hepatocytes. Another seven NAR rats underwent placement of indwelling catheters into the portal venous system via the gastroduodenal vein. Each of them received six batches of 5 × 106 normal albumin producing hepatocytes. Seven control NAR rats were infused repeatedly (intraportally) with saline only. Plasma albumin (ELISA) showed significant increase in experimental animals and was more pronounced (p < 0.05) in rats transplanted repeatedly than in those given a single dose of cells. Immunohistochemical staining of the liver sections confirmed the presence of transplanted albumin producing hepatocytes. Rats transplanted with a single large batch of isolated hepatocytes showed liver tissue damage, whereas those subjected to repeated cell infusions had normal liver histology. We have developed a novel intraportal transplantation method which allows successful engraftment of a large number of isolated hepatocytes.
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Affiliation(s)
- J Rozga
- Department of Surgery, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
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36
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Jauregui HO, Chowdhury NR, Chowdhury JR. Use of Mammalian Liver Cells for Artificial Liver Support. Cell Transplant 2017; 5:353-67. [PMID: 8727004 DOI: 10.1177/096368979600500302] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023] Open
Abstract
Advances in orthotopic liver transplantation have improved the survival rate of both acute and chronic liver failure patients to nearly 70%. However, the success of this treatment modality has created an international organ shortage. Many patients die while awaiting transplantation in part due to the minimal capacity to store viable transplantable livers beyond 24 h. Additionally, for many areas of the world, routine use of whole liver transplantation to treat liver disease is impractical due to the demands on both financial and technical resources. Potentially, these issues may be alleviated, at least in part, by the use of liver cell transplantation or cellular-based liver assist devices. The well-documented regenerative capacity of the liver may obviate the need for whole organ transplantation in some instances of acute failure, if the patient may be provided temporary metabolic support. Although other patients ultimately may require transplantation, a longer period of time to find a suitable organ for transplantation may be gained by that supportive therapy. The field of liver cell transplantation may offer solutions to patients with inherited metabolic deficiencies or chronic liver disease. The potential to treat an hepatic disorder by using only a fraction of the whole liver would increase the number of whole organs available for orthotopic liver transplantation. Research in the fields of hepatocyte based intra- and extra-corporeal liver support is providing evidence that these therapeutic modalities may ultimately become routine in the treatment of severe liver disease. A historic overview of that technology along with its current status is discussed.
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Affiliation(s)
- H O Jauregui
- Department of Pathology, Rhode Island Hospital, Providence 02903, USA
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37
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Ito M, Nagata H, Yamamoto T, Yoshihara D, Fox IJ, Miyakawa S. Intrasplenic Hepatocyte Transplantation Prolonged the Survival in Nagase Analbuminemic Rats with Liver Failure Induced by Common Bile Duct Ligation. Cell Transplant 2017; 16:547-53. [PMID: 17708344 DOI: 10.3727/000000007783464894] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
It has already been established that hepatocyte transplantation (HTx) in animal models, such as both chemically and surgically induced acute liver failure, liver-based metabolic disease, and cirrhosis, resulted in significant improvement of liver function and survival. However, the efficacy of hepatocyte transplantation in secondary cholestatic liver disease is not well known. In this study, we transplanted hepatocytes into the spleen of Nagase analbuminemic rats (NARs) with common bile duct ligation (CBDL) to evaluate the function of transplanted hepatocytes by both of serum albumin levels and total bilirubin levels. CBDL was carried out on NARs to induce liver failure. Lewis rat hepatocytes were transplanted in NARs 7 days after CBDL. Animals, in groups of four, underwent the following interventions: group 1—intrasplenic transplantation of 30 × 106 primary Lewis rat hepatocytes in NARs with CBDL (n = 4), group 2—intrasplenic injection of 0.5 ml DMEM in NARs with CBDL (n = 4); group 3—CBDL only (n = 4); group 4—intrasplenic transplantation of 30 × 106 primary Lewis rat hepatocytes in NARs (n = 4). Both bilirubin levels and albumin levels in NARs with CBDL were significantly improved post-HTx. Animals receiving hepatocyte transplantation survived longer than animals in nontransplant control groups. This study indicates that hepatocytes can be transplanted to temporarily provide life-supporting liver-specific metabolic function and prolong the survival in recipient rats with liver failure induced by CBDL.
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Affiliation(s)
- Masahiro Ito
- Department of Surgery, Fujita-Health University, Toyoake, Aichi, Japan.
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38
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Nicolas CT, Hickey RD, Chen HS, Mao SA, Lopera Higuita M, Wang Y, Nyberg SL. Concise Review: Liver Regenerative Medicine: From Hepatocyte Transplantation to Bioartificial Livers and Bioengineered Grafts. Stem Cells 2017; 35:42-50. [PMID: 27641427 PMCID: PMC5529050 DOI: 10.1002/stem.2500] [Citation(s) in RCA: 79] [Impact Index Per Article: 9.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2016] [Revised: 07/27/2016] [Accepted: 08/21/2016] [Indexed: 12/13/2022]
Abstract
Donor organ shortage is the main limitation to liver transplantation as a treatment for end-stage liver disease and acute liver failure. Liver regenerative medicine may in the future offer an alternative form of therapy for these diseases, be it through cell transplantation, bioartificial liver (BAL) devices, or bioengineered whole organ liver transplantation. All three strategies have shown promising results in the past decade. However, before they are incorporated into widespread clinical practice, the ideal cell type for each treatment modality must be found, and an adequate amount of metabolically active, functional cells must be able to be produced. Research is ongoing in hepatocyte expansion techniques, use of xenogeneic cells, and differentiation of stem cell-derived hepatocyte-like cells (HLCs). HLCs are a few steps away from clinical application, but may be very useful in individualized drug development and toxicity testing, as well as disease modeling. Finally, safety concerns including tumorigenicity and xenozoonosis must also be addressed before cell transplantation, BAL devices, and bioengineered livers occupy their clinical niche. This review aims to highlight the most recent advances and provide an updated view of the current state of affairs in the field of liver regenerative medicine. Stem Cells 2017;35:42-50.
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Affiliation(s)
- Clara T Nicolas
- William J Von Liebig Transplant Center, Mayo Clinic, Rochester, Minnesota, USA
- Department of Surgery, Mayo Clinic, Rochester, Minnesota, USA
| | - Raymond D Hickey
- Department of Molecular Medicine, Mayo Clinic, Rochester, Minnesota, USA
- Department of Surgery, Mayo Clinic, Rochester, Minnesota, USA
| | - Harvey S Chen
- Department of Surgery, Mayo Clinic, Rochester, Minnesota, USA
| | - Shennen A Mao
- Department of Surgery, Mayo Clinic, Rochester, Minnesota, USA
| | - Manuela Lopera Higuita
- Department of Physiology and Biomedical Engineering, Mayo Clinic College of Medicine, Rochester, Minnesota, USA
| | - Yujia Wang
- William J Von Liebig Transplant Center, Mayo Clinic, Rochester, Minnesota, USA
| | - Scott L Nyberg
- William J Von Liebig Transplant Center, Mayo Clinic, Rochester, Minnesota, USA
- Department of Surgery, Mayo Clinic, Rochester, Minnesota, USA
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Yang J, Zhu Y, Xu T, Pan C, Cai N, Huang H, Zhang L. The preservation of living cells with biocompatible microparticles. NANOTECHNOLOGY 2016; 27:265101. [PMID: 27189861 DOI: 10.1088/0957-4484/27/26/265101] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/05/2023]
Abstract
Biomedical applications of living cells have rapidly expanded in many fields such as toxic detection, drug screening, and regenerative medicine, etc. Efficient methods to support cell survival and maintain activity in vitro have become increasingly important. However, traditional cryopreservation for living cell-based applications is limited by several problems. Here, we report that magnetic hydrogel microparticles can physically assemble into a 3D environment for efficient cell preservation in physiological conditions, avoiding any chemical reactions that would damage the cells. Two representative cell lines (loosely and firmly adherent) were tested to evaluate the versatility of this method. The results showed that cell longevity was significantly extended to at least 15 days, while the control cell samples without microparticles quickly died within 3 days. Moreover, after preservation, cells can be easily retrieved by applying a magnet to separate the magnetic particles. This strategy can also inhibit cell over-proliferation while avoiding the use of temperature extremes or toxic cryoprotectants that are essential in cryopreservation.
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Affiliation(s)
- Jing Yang
- Department of Biochemical Engineering, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China. Key Laboratory of Systems Bioengineering of the Ministry of Education, Tianjin University, Tianjin 300072, China. Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin University, Tianjin, 300072, China
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Abstract
Liver disease is a leading cause of morbidity and mortality. Liver transplantation remains the only proven treatment for end-stage liver failure but is limited by the availability of donor organs. Hepatocyte cell therapy, either with bioartificial liver devices or hepatocyte transplantation, may help address this by delaying or preventing liver transplantation. Early clinical studies have shown promising results, however in most cases, the benefit has been short lived and so further research into these therapies is required. Alternative sources of hepatocytes, including stem cell-derived hepatocytes, are being investigated as the isolation of primary human hepatocytes is limited by the same shortage of donor organs. This review summarises the current clinical experience of hepatocyte cell therapy together with an overview of possible alternative sources of hepatocytes. Current and future areas for research that might lead towards the realisation of the full potential of hepatocyte cell therapy are discussed.
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Affiliation(s)
- David Christopher Bartlett
- a NIHR Centre for Liver Research and Biomedical Research Unit, University of Birmingham, Birmingham, UK.,b Liver Unit, University Hospital Birmingham NHS Foundation Trust, Birmingham, UK
| | - Philip N Newsome
- a NIHR Centre for Liver Research and Biomedical Research Unit, University of Birmingham, Birmingham, UK.,b Liver Unit, University Hospital Birmingham NHS Foundation Trust, Birmingham, UK
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Multi-OMICs and Genome Editing Perspectives on Liver Cancer Signaling Networks. BIOMED RESEARCH INTERNATIONAL 2016; 2016:6186281. [PMID: 27403431 PMCID: PMC4923561 DOI: 10.1155/2016/6186281] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 01/01/2016] [Revised: 04/23/2016] [Accepted: 05/08/2016] [Indexed: 12/26/2022]
Abstract
The advent of the human genome sequence and the resulting ~20,000 genes provide a crucial framework for a transition from traditional biology to an integrative “OMICs” arena (Lander et al., 2001; Venter et al., 2001; Kitano, 2002). This brings in a revolution for cancer research, which now enters a big data era. In the past decade, with the facilitation by next-generation sequencing, there have been a huge number of large-scale sequencing efforts, such as The Cancer Genome Atlas (TCGA), the HapMap, and the 1000 genomes project. As a result, a deluge of genomic information becomes available from patients stricken by a variety of cancer types. The list of cancer-associated genes is ever expanding. New discoveries are made on how frequent and highly penetrant mutations, such as those in the telomerase reverse transcriptase (TERT) and TP53, function in cancer initiation, progression, and metastasis. Most genes with relatively frequent but weakly penetrant cancer mutations still remain to be characterized. In addition, genes that harbor rare but highly penetrant cancer-associated mutations continue to emerge. Here, we review recent advances related to cancer genomics, proteomics, and systems biology and suggest new perspectives in targeted therapy and precision medicine.
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Heparinization of cell surfaces with short peptide-conjugated PEG-lipid regulates thromboinflammation in transplantation of human MSCs and hepatocytes. Acta Biomater 2016; 35:194-205. [PMID: 26876877 DOI: 10.1016/j.actbio.2016.02.018] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2015] [Revised: 02/05/2016] [Accepted: 02/11/2016] [Indexed: 12/14/2022]
Abstract
Infusion of therapeutic cells into humans is associated with immune responses, including thromboinflammation, which result in a large loss of transplanted cells. To address these problems, heparinization of the cell surfaces was achieved by a cell-surface modification technique using polyethylene glycol-conjugated phospholipid (PEG-lipid) derivatives. A short heparin-binding peptide was conjugated to the PEG-lipid for immobilization of heparin conjugates on the surface of human mesenchymal stem cells (hMSCs) and human hepatocytes. Here three kinds of heparin-binding peptides were used for immobilizing heparin conjugates and examined for the antithrombogenic effects on the cell surface. The heparinized cells were incubated in human whole blood to evaluate their hemocompatibility by measuring blood parameters such as platelet count, coagulation markers, complement markers, and Factor Xa activity. We found that one of the heparin-binding peptides did not show cytotoxicity after the immobilization with heparin conjugates. The degree of binding of the heparin conjugates on the cell surface (analyzed by flow cytometer) depended on the ratio of the active peptide to control peptide. For both human MSCs and hepatocytes in whole-blood experiments, no platelet aggregation was seen in the heparin conjugate-immobilized cell group vs. the controls (non-coated cells or control peptide). Also, the levels of thrombin-antithrombin complex (TAT), C3a, and sC5b-9 were significantly lower than those of the controls, indicating a lower activation of coagulation and complement. Factor Xa analysis indicated that the heparin conjugate was still active on the cell surface at 24h post-coating. It is possible to immobilize heparin conjugates onto hMSC and human hepatocyte surfaces and thereby protect the cell surfaces from damaging thromboinflammation. STATEMENT OF SIGNIGFICANCE We present a promising approach to enhance the biocompatibility of therapeutic cells. Here we used short peptide-conjugated PEG-lipid for cell surface modification and heparin conjugates for the coating of human hepatocytes and MSCs. We screened the short peptides to find higher affinity for heparinization of cell surface and performed hemocompatibility assay of heparinized human hepatocytes and human MSCs in human whole blood. Using heparin-binding peptide with higher affinity, not only coagulation activation but also complement activation was significantly suppressed. Thus, it was possible to protect human hepatocytes and human MSCs from the attack of thromboinflammatory activation, which can contribute to the improvement graft survival.
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Na GH, Kim DG, Jung ES. Culture with Growth Factor Supplements Improves the Viability and Function of Rat Hepatocytes. KOREAN JOURNAL OF TRANSPLANTATION 2015. [DOI: 10.4285/jkstn.2015.29.3.139] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/04/2022] Open
Affiliation(s)
- Gun Hyung Na
- Department of Surgery, Seoul St. Mary’s Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea
| | - Dong Goo Kim
- Department of Surgery, Seoul St. Mary’s Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea
| | - Eun Sun Jung
- Department of Pathology, Seoul St. Mary’s Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea
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Stevens KR, Miller JS, Blakely BL, Chen CS, Bhatia SN. Degradable hydrogels derived from PEG-diacrylamide for hepatic tissue engineering. J Biomed Mater Res A 2015; 103:3331-8. [PMID: 25851120 PMCID: PMC4890565 DOI: 10.1002/jbm.a.35478] [Citation(s) in RCA: 53] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/05/2015] [Revised: 04/02/2015] [Accepted: 04/02/2015] [Indexed: 01/19/2023]
Abstract
Engineered tissue constructs have the potential to augment or replace whole organ transplantation for the treatment of liver failure. Poly(ethylene glycol) (PEG)‐based systems are particularly promising for the construction of engineered liver tissue due to their biocompatibility and amenability to modular addition of bioactive factors. To date, primary hepatocytes have been successfully encapsulated in non‐degradable hydrogels based on PEG‐diacrylate (PEGDA). In this study, we describe a hydrogel system based on PEG‐diacrylamide (PEGDAAm) containing matrix‐metalloproteinase sensitive (MMP‐sensitive) peptide in the hydrogel backbone that is suitable for hepatocyte culture both in vitro and after implantation. By replacing hydrolytically unstable esters in PEGDA with amides in PEGDAAm, resultant hydrogels resisted non‐specific hydrolysis, while still allowing for MMP‐mediated hydrogel degradation. Optimization of polymerization conditions, hepatocellular density, and multicellular tissue composition modulated both the magnitude and longevity of hepatic function in vitro. Importantly, hepatic PEGDAAm‐based tissues survived and functioned for over 3 weeks after implantation ectopically in the intraperitoneal (IP) space of nude mice. Together, these studies suggest that MMP‐sensitive PEGDAAm‐based hydrogels may be a useful material system for applications in tissue engineering and regenerative medicine. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 103A: 3331–3338, 2015.
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Affiliation(s)
- Kelly R Stevens
- Harvard-MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, Massachusetts, 02139
| | - Jordan S Miller
- Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania, 19104
| | - Brandon L Blakely
- Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania, 19104
| | - Christopher S Chen
- Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania, 19104
| | - Sangeeta N Bhatia
- Harvard-MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, Massachusetts, 02139.,Howard Hughes Medical Institute, Cambridge, Massachusetts, 02139.,Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology, Cambridge, Massachusetts, 02139
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Hickey RD, Mao SA, Amiot B, Suksanpaisan L, Miller A, Nace R, Glorioso J, Peng KW, Ikeda Y, Russell SJ, Nyberg SL. Noninvasive 3-dimensional imaging of liver regeneration in a mouse model of hereditary tyrosinemia type 1 using the sodium iodide symporter gene. Liver Transpl 2015; 21:442-53. [PMID: 25482651 PMCID: PMC5957080 DOI: 10.1002/lt.24057] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/26/2014] [Accepted: 11/30/2014] [Indexed: 12/24/2022]
Abstract
Cell transplantation is a potential treatment for the many liver disorders that are currently only curable by organ transplantation. However, one of the major limitations of hepatocyte (HC) transplantation is an inability to monitor cells longitudinally after injection. We hypothesized that the thyroidal sodium iodide symporter (NIS) gene could be used to visualize transplanted HCs in a rodent model of inherited liver disease: hereditary tyrosinemia type 1. Wild-type C57Bl/6J mouse HCs were transduced ex vivo with a lentiviral vector containing the mouse Slc5a5 (NIS) gene controlled by the thyroxine-binding globulin promoter. NIS-transduced cells could robustly concentrate radiolabeled iodine in vitro, with lentiviral transduction efficiencies greater than 80% achieved in the presence of dexamethasone. Next, NIS-transduced HCs were transplanted into congenic fumarylacetoacetate hydrolase knockout mice, and this resulted in the prevention of liver failure. NIS-transduced HCs were readily imaged in vivo by single-photon emission computed tomography, and this demonstrated for the first time noninvasive 3-dimensional imaging of regenerating tissue in individual animals over time. We also tested the efficacy of primary HC spheroids engrafted in the liver. With the NIS reporter, robust spheroid engraftment and survival could be detected longitudinally after direct parenchymal injection, and this thereby demonstrated a novel strategy for HC transplantation. This work is the first to demonstrate the efficacy of NIS imaging in the field of HC transplantation. We anticipate that NIS labeling will allow noninvasive and longitudinal identification of HCs and stem cells in future studies related to liver regeneration in small and large preclinical animal models.
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Affiliation(s)
- Raymond D. Hickey
- Department of Molecular Medicine, Mayo Clinic, Rochester, MN, USA
- Department of Surgery, Mayo Clinic, Rochester, MN, USA
| | | | - Bruce Amiot
- Department of Surgery, Mayo Clinic, Rochester, MN, USA
| | | | - Amber Miller
- Department of Molecular Medicine, Mayo Clinic, Rochester, MN, USA
| | - Rebecca Nace
- Department of Molecular Medicine, Mayo Clinic, Rochester, MN, USA
| | | | - Kah Whye Peng
- Department of Molecular Medicine, Mayo Clinic, Rochester, MN, USA
| | - Yasuhiro Ikeda
- Department of Molecular Medicine, Mayo Clinic, Rochester, MN, USA
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Hansel MC, Gramignoli R, Skvorak KJ, Dorko K, Marongiu F, Blake W, Davila J, Strom SC. The history and use of human hepatocytes for the treatment of liver diseases: the first 100 patients. CURRENT PROTOCOLS IN TOXICOLOGY 2014; 62:14.12.1-23. [PMID: 25378242 PMCID: PMC4343212 DOI: 10.1002/0471140856.tx1412s62] [Citation(s) in RCA: 62] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/26/2023]
Abstract
Orthotopic liver transplantation remains the only curative treatment for many end-stage liver diseases, yet the number of patients receiving liver transplants remains limited by the number of organs available for transplant. There is a need for alternative therapies for liver diseases. The transplantation of isolated hepatocytes (liver cells) has been used as an experimental therapy for liver disease in a limited number of cases. Recently, the 100th case of hepatocyte transplantation was reported. This review discusses the history of the hepatocyte transplant field, the major discoveries that supported and enabled the first hepatocyte transplants, and reviews the cases and outcomes of the first 100 clinical transplants. Some of the problems that limit the application or efficacy of hepatocyte transplantation are discussed, as are possible solutions to these problems. In conclusion, hepatocyte transplants have proven effective particularly in cases of metabolic liver disease where reversal or amelioration of the characteristic symptoms of the disease is easily quantified. However, no patients have been completely corrected of a metabolic liver disease for a significant amount of time by hepatocyte transplantation alone. It is likely that future developments in new sources of cells for transplantation will be required before this cellular therapy can be fully implemented and available for large numbers of patients.
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Affiliation(s)
- Marc C Hansel
- Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania; McGowan Institute for Regenerative Medicine, Pittsburgh, Pennsylvania
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Cell-based therapy for acute and chronic liver failures: distinct diseases, different choices. Sci Rep 2014; 4:6494. [PMID: 25263068 PMCID: PMC4178291 DOI: 10.1038/srep06494] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2014] [Accepted: 09/08/2014] [Indexed: 02/06/2023] Open
Abstract
Cell-based therapies (CBTs) are considered the effective approaches to treat liver failure. However, which cell type is the most suitable source of CBTs for acute liver failure (ALF) or chronic liver failure (CLF) remains unclear. To investigate this, mature hepatocytes in adult liver (adult HCs), fetal liver cells (FLCs), induced hepatic stem cells (iHepSCs) and bone marrow derived mesenchymal stromal cells (BMSCs) were used to CBTs for ConA-induced ALF and Fah-deficient induced CLF in mice. The results showed that only BMSCs remitted liver damage and rescued ALF in ConA-treated mice. In this process, BMSCs inhibited ConA-induced inflammatory response by decreasing the mRNA expressions of TNF-α, IFN-γ and FasL and increasing IL-10 mRNA expression. However, in the CLF model, not BMSCs but adult HCs transplantation lessened liver injury, recovered liver function and rescued the life of Fah-/- mice after NTBC withdrawal. Further study showed that adult HCs offered more effective liver regeneration compared to other cells in Fah-/- mice without NTBC. These results demonstrated that BMSCs and adult HCs are the optimal sources of CBTs for ConA-induced ALF and Fah-deficient induced CLF in mice, respectively. This finding deepens our understanding about how to select a proper CBT for different liver failure.
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Fox IJ, Daley GQ, Goldman SA, Huard J, Kamp TJ, Trucco M. Stem cell therapy. Use of differentiated pluripotent stem cells as replacement therapy for treating disease. Science 2014; 345:1247391. [PMID: 25146295 PMCID: PMC4329726 DOI: 10.1126/science.1247391] [Citation(s) in RCA: 217] [Impact Index Per Article: 19.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
Pluripotent stem cells (PSCs) directed to various cell fates holds promise as source material for treating numerous disorders. The availability of precisely differentiated PSC-derived cells will dramatically affect blood component and hematopoietic stem cell therapies and should facilitate treatment of diabetes, some forms of liver disease and neurologic disorders, retinal diseases, and possibly heart disease. Although an unlimited supply of specific cell types is needed, other barriers must be overcome. This review of the state of cell therapies highlights important challenges. Successful cell transplantation will require optimizing the best cell type and site for engraftment, overcoming limitations to cell migration and tissue integration, and occasionally needing to control immunologic reactivity, as well as a number of other challenges. Collaboration among scientists, clinicians, and industry is critical for generating new stem cell-based therapies.
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Affiliation(s)
- Ira J Fox
- Department of Surgery, Children's Hospital of Pittsburgh and McGowan Institute for Regenerative Medicine, University of Pittsburgh, Pittsburgh, PA, USA.
| | - George Q Daley
- Boston Children's Hospital and Dana Farber Cancer Institute, Boston, MA, USA. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School Broad Institute, Cambridge, MA, USA. Howard Hughes Medical Institute, Chevy Chase, MD, USA
| | - Steven A Goldman
- Center for Translational Neuromedicine, The University of Rochester Medical Center, Rochester, NY, USA. Center for Basic and Translational Neuroscience, University of Copenhagen, Denmark
| | - Johnny Huard
- Stem Cell Research Center, Department of Orthopaedic Surgery, University of Pittsburgh, School of Medicine, Pittsburgh, PA, USA
| | - Timothy J Kamp
- Stem Cell and Regenerative Medicine Center, Cellular and Molecular Arrhythmia Research Program, Department of Medicine, School of Medicine and Public Health, University of Wisconsin, Madison, WI, USA
| | - Massimo Trucco
- Division of Immunogenetics, Children's Hospital of Pittsburgh, University of Pittsburgh, Pittsburgh, PA, USA
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Na GH, Kim DG, Kim YH, Han JH, Jung ES. Effects of glucose concentration in the medium on rat hepatocyte culture. Ann Surg Treat Res 2014; 87:53-60. [PMID: 25114883 PMCID: PMC4127907 DOI: 10.4174/astr.2014.87.2.53] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2014] [Revised: 04/07/2014] [Accepted: 04/28/2014] [Indexed: 11/30/2022] Open
Abstract
Purpose To determine the optimum culture conditions by investigating isolated rat hepatocytes cultured in medium containing different glucose concentrations. Methods Hepatocytes were isolated from rats using a two-step perfusion technique and divided into the following two groups cultured in medium containing different glucose concentrations: (1) low-glucose group and (2) high-glucose group. Total cell count and viability of cultured rat hepatocytes and liver function parameters (i.e., concentrations of albumin, ammonia, and urea in the culture medium) were measured. The morphology of cultured rat hepatocytes was examined by staining with hematoxylin and eosin, and albumin receptor expression was confirmed by immunofluorescence. Results Total cell count and viability showed smaller increases in the low-glucose group than the high-glucose group, although the difference was not statistically significant (P = 0.112 and P = 0.147, respectively). The levels of albumin (P = 0.943), ammonia (P = 0.744), and urea (P = 0.709) were not significantly different between the two groups. In both groups, the function of cultured hepatocytes decreased significantly over time. The morphology of hepatocytes was well maintained in both groups at 3 days. On day 7, the cytoplasm was transformed into a spindle shape. On day 10, these changes were exaggerated, and were more prominent in the high-glucose group. Conclusion Morphological assessment indicated that low-glucose culture medium is better than high-glucose culture medium for culturing of hepatocytes, although there was not significantly different in functional assessment. The cultured hepatocytes with low-glucose culture medium could be maintained for 7 days.
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Affiliation(s)
- Gun Hyung Na
- Department of Surgery, Seoul St. Mary's Hospital, The Catholic University of Korea College of Medicine, Seoul, Korea
| | - Dong Goo Kim
- Department of Surgery, Seoul St. Mary's Hospital, The Catholic University of Korea College of Medicine, Seoul, Korea
| | - Young Hui Kim
- Department of Surgery, Seoul St. Mary's Hospital, The Catholic University of Korea College of Medicine, Seoul, Korea
| | - Jae Hyun Han
- Department of Surgery, Seoul St. Mary's Hospital, The Catholic University of Korea College of Medicine, Seoul, Korea
| | - Eun Sun Jung
- Department of Pathology, Seoul St. Mary's Hospital, The Catholic University of Korea College of Medicine, Seoul, Korea
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Abstract
Despite the tremendous hurdles presented by the complexity of the liver's structure and function, advances in liver physiology, stem cell biology and reprogramming, and the engineering of tissues and devices are accelerating the development of cell-based therapies for treating liver disease and liver failure. This State of the Art Review discusses both the near- and long-term prospects for such cell-based therapies and the unique challenges for clinical translation.
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Affiliation(s)
- Sangeeta N Bhatia
- Institute for Medical Engineering & Science at MIT, Department of Electrical Engineering and Computer Science, David H. Koch Institute at MIT, and the Howard Hughes Medical Institute, Cambridge, MA 02139, USA. Division of Medicine, Brigham and Women's Hospital, Boston, MA 02115, USA.
| | - Gregory H Underhill
- Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA
| | - Kenneth S Zaret
- Institute for Regenerative Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA
| | - Ira J Fox
- Department of Surgery, Children's Hospital of Pittsburgh of UPMC, University of Pittsburgh School of Medicine, and McGowan Institute for Regenerative Medicine, Pittsburgh, PA 15224, USA
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