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Pan Y, Zheng Z, Zhang X, Liu S, Zhuansun S, Gong S, Li S, Wang H, Chen Y, Yang T, Wu H, Xue F, Xia Q, He K. Hybrid Bioactive Hydrogel Promotes Liver Regeneration through the Activation of Kupffer Cells and ECM Remodeling After Partial Hepatectomy. Adv Healthc Mater 2024; 13:e2303828. [PMID: 38608209 DOI: 10.1002/adhm.202303828] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2023] [Revised: 03/31/2024] [Indexed: 04/14/2024]
Abstract
Partial hepatectomy is an essential surgical technique used to treat advanced liver diseases such as liver tumors, as well as for performing liver transplants from living donors. However, postoperative complications such as bleeding, abdominal adhesions, wound infections, and inadequate liver regeneration pose significant challenges and increase morbidity and mortality rates. A self-repairing mixed hydrogel (O5H2/Cu2+/SCCK), containing stem cell derived cytokine (SCCK) derived from human umbilical cord mesenchymal stem cells (HUMSCs) treated with the traditional Chinese remedy Tanshinone IIA (TSA), is developed. This SCCK, in conjunction with O5H2, demonstrates remarkable effects on Kupffer cell activation and extracellular matrix (ECM) remodeling. This leads to the secretion of critical growth factors promoting enhanced proliferation of hepatocytes and endothelial cells, thereby facilitating liver regeneration and repair after partial hepatectomy. Furthermore, the hydrogel, featuring macrophage-regulating properties, effectively mitigates inflammation and oxidative stress damage in the incision area, creating an optimal environment for postoperative liver regeneration. The injectability and strong adhesion of the hydrogel enables rapid hemostasis at the incision site, while its physical barrier function prevents postoperative abdominal adhesions. Furthermore, the hydrogel's incorporation of Cu2+ provides comprehensive antibacterial effects, protecting against a wide range of bacteria types and reducing the chances of infections after surgery.
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Affiliation(s)
- Yixiao Pan
- Department of Liver Surgery and Liver Transplantation, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, P. R. China
- Shanghai Engineering Research Center of Transplantation and Immunology, Shanghai, 200127, P. R. China
- Shanghai Institute of Transplantation, Shanghai, 200127, P. R. China
| | - Zhigang Zheng
- Department of Liver Surgery and Liver Transplantation, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, P. R. China
- Shanghai Engineering Research Center of Transplantation and Immunology, Shanghai, 200127, P. R. China
- Shanghai Institute of Transplantation, Shanghai, 200127, P. R. China
| | - Xueliang Zhang
- School of Chemistry and Chemical Engineering, Shanghai Jiao Tong University, Shanghai, 200240, P. R. China
| | - Shupeng Liu
- Department of Gastroenterology, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, P. R. China
| | - Shiya Zhuansun
- Department of Hematology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, No. 725 Wanping South Road, Xuhui District, Shanghai, P. R. China
| | - Shiming Gong
- Department of Liver Surgery and Liver Transplantation, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, P. R. China
- Shanghai Engineering Research Center of Transplantation and Immunology, Shanghai, 200127, P. R. China
- Shanghai Institute of Transplantation, Shanghai, 200127, P. R. China
| | - Shilun Li
- Department of Vascular Surgery, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, P. R. China
| | - Hongye Wang
- Department of Interventional Oncology, Renji Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, 200127, P. R. China
| | - Yiwen Chen
- School of Medicine, Shanghai Jiao Tong University, Shanghai, 200025, P. R. China
| | - Taihua Yang
- Department of Liver Surgery and Liver Transplantation, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, P. R. China
- Shanghai Engineering Research Center of Transplantation and Immunology, Shanghai, 200127, P. R. China
- Shanghai Institute of Transplantation, Shanghai, 200127, P. R. China
| | - Huimin Wu
- Department of Liver Surgery and Liver Transplantation, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, P. R. China
- Shanghai Engineering Research Center of Transplantation and Immunology, Shanghai, 200127, P. R. China
- Shanghai Institute of Transplantation, Shanghai, 200127, P. R. China
| | - Feng Xue
- Department of Liver Surgery and Liver Transplantation, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, P. R. China
- Shanghai Engineering Research Center of Transplantation and Immunology, Shanghai, 200127, P. R. China
- Shanghai Institute of Transplantation, Shanghai, 200127, P. R. China
| | - Qiang Xia
- Department of Liver Surgery and Liver Transplantation, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, P. R. China
- Shanghai Engineering Research Center of Transplantation and Immunology, Shanghai, 200127, P. R. China
- Shanghai Institute of Transplantation, Shanghai, 200127, P. R. China
| | - Kang He
- Department of Liver Surgery and Liver Transplantation, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, P. R. China
- Shanghai Engineering Research Center of Transplantation and Immunology, Shanghai, 200127, P. R. China
- Shanghai Institute of Transplantation, Shanghai, 200127, P. R. China
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Tavabie OD, Salehi S, Aluvihare VR. The challenges and potential in developing microRNA associated with regeneration as biomarkers to improve prognostication for liver failure syndromes and hepatocellular carcinoma. Expert Rev Mol Diagn 2024; 24:5-22. [PMID: 38059597 DOI: 10.1080/14737159.2023.2292642] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2023] [Accepted: 12/05/2023] [Indexed: 12/08/2023]
Abstract
INTRODUCTION Determining the need for liver transplantation remains critical in the management of hepatocellular carcinoma (HCC) and liver failure syndromes (including acute liver failure and decompensated cirrhosis states). Conventional prognostic models utilize biomarkers of liver and non-liver failure and have limitations in their application. Novel biomarkers which predict regeneration may fulfil this niche. microRNA are implicated in health and disease and are present in abundance in the circulation. Despite this, they have not translated into mainstream clinical biomarkers. AREAS COVERED We will discuss current challenges in the prognostication of patients with liver failure syndromes as well as for patients with HCC. We will discuss biomarkers implicated with liver regeneration. We then provide an overview of the challenges in developing microRNA into clinically tractable biomarkers. Finally, we will provide a scoping review of microRNA which may have potential as prognostic biomarkers in liver failure syndromes and HCC. EXPERT OPINION Novel biomarkers are needed to improve prognostic models in liver failure syndromes and HCC. Biomarkers associated with liver regeneration are currently lacking and may fulfil this niche. microRNA have the potential to be developed into clinically tractable biomarkers but a consensus on standardizing methodology and reporting is required prior to large-scale studies.
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Affiliation(s)
| | - Siamak Salehi
- Institute of Liver Studies, King's College Hospital, London, UK
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Deshmukh K, Apte U. The Role of Endoplasmic Reticulum Stress Response in Liver Regeneration. Semin Liver Dis 2023; 43:279-292. [PMID: 37451282 PMCID: PMC10942737 DOI: 10.1055/a-2129-8977] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 07/18/2023]
Abstract
Exposure to hepatotoxic chemicals is involved in liver disease-related morbidity and mortality worldwide. The liver responds to damage by triggering compensatory hepatic regeneration. Physical agent or chemical-induced liver damage disrupts hepatocyte proteostasis, including endoplasmic reticulum (ER) homeostasis. Post-liver injury ER experiences a homeostatic imbalance, followed by active ER stress response signaling. Activated ER stress response causes selective upregulation of stress response genes and downregulation of many hepatocyte genes. Acetaminophen overdose, carbon tetrachloride, acute and chronic alcohol exposure, and physical injury activate the ER stress response, but details about the cellular consequences of the ER stress response on liver regeneration remain unclear. The current data indicate that inhibiting the ER stress response after partial hepatectomy-induced liver damage promotes liver regeneration, whereas inhibiting the ER stress response after chemical-induced hepatotoxicity impairs liver regeneration. This review summarizes key findings and emphasizes the knowledge gaps in the role of ER stress in injury and regeneration.
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Affiliation(s)
- Kshitij Deshmukh
- Interdisciplinary Graduate Program in Human Toxicology, University of Iowa, Iowa City, Iowa
| | - Udayan Apte
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas
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Zhu X, Zhang B, He Y, Bao J. Liver Organoids: Formation Strategies and Biomedical Applications. Tissue Eng Regen Med 2021; 18:573-585. [PMID: 34132985 DOI: 10.1007/s13770-021-00357-w] [Citation(s) in RCA: 31] [Impact Index Per Article: 7.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2020] [Revised: 05/20/2021] [Accepted: 05/22/2021] [Indexed: 02/05/2023] Open
Abstract
The liver is the most important digestive organ in the body. Several studies have explored liver biology and diseases related to the liver. However, most of these studies have only explored liver development, mechanism of liver regeneration and pathophysiology of liver diseases mainly based on two-dimensional (2D) cell lines and animal models. Traditional 2D cell lines do not represent the complex three-dimensional tissue architecture whereas animal models are limited by inter-species differences. These shortcomings limit understanding of liver biology and diseases. Liver organoid technology is effective in elucidating structural and physiological characteristics and basic tissue-level functions of liver tissue. In this review, formation strategies and a wide range of applications in biomedicine of liver organoid are summarized. Liver organoids are derived from single type cell culture, such as induced pluripotent stem cells (iPSCs), adult stem cells, primary hepatocytes, and primary cholangiocytes and multi-type cells co-culture, such as iPSC-derived hepatic endoderm cells co-cultured with mesenchymal stem cells and umbilical cord-derived endothelial cells. In vitro studies report that liver organoids are a promising model for regenerative medicine, organogenesis, liver regeneration, disease modelling, drug screening and personalized treatment. Liver organoids are a promising in vitro model for basic research and for development of clinical therapeutic interventions for hepatopathy.
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Affiliation(s)
- Xinglong Zhu
- Institute of Clinical Pathology, Key Laboratory of Transplant Engineering and Immunology, NHC, West China Hospital, Sichuan University, No. 37, Guoxue Alley, Wuhou District, Chengdu, 610041, Sichuan Province, China
| | - Bingqi Zhang
- Institute of Clinical Pathology, Key Laboratory of Transplant Engineering and Immunology, NHC, West China Hospital, Sichuan University, No. 37, Guoxue Alley, Wuhou District, Chengdu, 610041, Sichuan Province, China
| | - Yuting He
- Institute of Clinical Pathology, Key Laboratory of Transplant Engineering and Immunology, NHC, West China Hospital, Sichuan University, No. 37, Guoxue Alley, Wuhou District, Chengdu, 610041, Sichuan Province, China
| | - Ji Bao
- Institute of Clinical Pathology, Key Laboratory of Transplant Engineering and Immunology, NHC, West China Hospital, Sichuan University, No. 37, Guoxue Alley, Wuhou District, Chengdu, 610041, Sichuan Province, China.
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Liver regeneration: biological and pathological mechanisms and implications. Nat Rev Gastroenterol Hepatol 2021; 18:40-55. [PMID: 32764740 DOI: 10.1038/s41575-020-0342-4] [Citation(s) in RCA: 521] [Impact Index Per Article: 130.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 06/24/2020] [Indexed: 02/08/2023]
Abstract
The liver is the only solid organ that uses regenerative mechanisms to ensure that the liver-to-bodyweight ratio is always at 100% of what is required for body homeostasis. Other solid organs (such as the lungs, kidneys and pancreas) adjust to tissue loss but do not return to 100% of normal. The current state of knowledge of the regenerative pathways that underlie this 'hepatostat' will be presented in this Review. Liver regeneration from acute injury is always beneficial and has been extensively studied. Experimental models that involve partial hepatectomy or chemical injury have revealed extracellular and intracellular signalling pathways that are used to return the liver to equivalent size and weight to those prior to injury. On the other hand, chronic loss of hepatocytes, which can occur in chronic liver disease of any aetiology, often has adverse consequences, including fibrosis, cirrhosis and liver neoplasia. The regenerative activities of hepatocytes and cholangiocytes are typically characterized by phenotypic fidelity. However, when regeneration of one of the two cell types fails, hepatocytes and cholangiocytes function as facultative stem cells and transdifferentiate into each other to restore normal liver structure. Liver recolonization models have demonstrated that hepatocytes have an unlimited regenerative capacity. However, in normal liver, cell turnover is very slow. All zones of the resting liver lobules have been equally implicated in the maintenance of hepatocyte and cholangiocyte populations in normal liver.
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Gu C, Chai M, Liu J, Wang H, Du W, Zhou Y, Tan WS. Expansion of Transdifferentiated Human Hepatocytes in a Serum-Free Microcarrier Culture System. Dig Dis Sci 2020; 65:2009-2023. [PMID: 31722057 DOI: 10.1007/s10620-019-05925-8] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/14/2019] [Accepted: 10/29/2019] [Indexed: 02/07/2023]
Abstract
BACKGROUND AND AIMS Bioartificial livers (BALs) have attracted much attention as potential supportive therapies for liver diseases. A serum-free microcarrier culture strategy for the in vitro high-density expansion of human-induced hepatocyte-like cells (hiHeps) suitable for BALs was studied in this article. METHODS hiHeps were transdifferentiated from human fibroblasts by the lentiviral overexpression of FOXA3, HNF1A, and HNF4A. Cells were cultured on microcarriers, their proliferation was evaluated by cell count and CCK-8 assays, and their function was evaluated by detecting liver function parameters in the supernatant, including urea secretion, albumin synthesis, and lactate dehydrogenase levels. The expressions of hepatocyte function-associated genes of hiHeps were measured by qRT-PCR in 2D and 3D conditions. The expression of related proteins during fibronectin promotes cell adhesion, and proliferation on microcarrier was detected by western blotting. RESULTS During microcarrier culture, the optimal culture conditions during the adherence period were the use of half-volume high-density inoculation, Cytodex 3 at a concentration of 3 mg/mL, a cell seeding density of 2.0 × 105 cells/mL, and a stirring speed of 45 rpm. The final cell density in self-developed, chemically defined serum-free medium (SFM) reached 2.53 × 106 cells/mL, and the maximum increase in expansion was 12.61-fold. In addition, we found that fibronectin (FN) can promote hiHep attachment and proliferation on Cytodex 3 microcarriers and that this pro-proliferative effect was mediated by the integrin-β1/FAK/ERK/CyclinD1 signaling pathway. Finally, the growth and function of hiHeps on Cytodex 3 in SFM were close to those of hiHeps on Cytodex 3 in hepatocyte maintenance medium (HMM), and cells maintained their morphology and function after harvest on microcarriers. CONCLUSIONS Serum-free microcarrier culture has important implications for the expansion of a sufficient number of hiHeps prior to the clinical application of BALs.
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Affiliation(s)
- Ce Gu
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, People's Republic of China
| | - Miaomiao Chai
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, People's Republic of China
| | - Jiaxing Liu
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, People's Republic of China
| | - Hui Wang
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, People's Republic of China
| | - Wenjing Du
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, People's Republic of China
| | - Yan Zhou
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, People's Republic of China.
| | - Wen-Song Tan
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, People's Republic of China
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Abstract
Introduction: Liver disease is an increasing cause of worldwide mortality, and currently the only curative treatment for end-stage liver disease is whole organ allograft transplantation. Whilst this is an effective treatment, there is a shortage of suitable grafts and consequently some patients die whilst on the waiting list. Cell therapy provides an alternative treatment to increase liver function and potentially ameliorate fibrosis. Areas covered: In this review, we discuss the different cellular sources for therapy investigated to date in humans including mature hepatocytes, hematopoietic stem cells, mesenchymal stromal cells and hepatic progenitor cells. Cells investigated in animals include embryonic stem cells, induced pluripotent stem cells and directly reprogrammed cells. We then appraise the experience and evidence base underlying each cell type. Expert opinion: We discuss how this field may evolve in the years to come focusing on opportunities to enhance the intrinsic regenerative response with therapeutic targets and cell therapies. Growing expertise in tissue engineering will likely lead to increasingly complex bio-reactors and bio-artificial livers, which open a further avenue to restore liver function and delay or prevent the need for transplantation.
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Affiliation(s)
- Alexander Boyd
- a NIHR Birmingham Biomedical Research Centre , University Hospitals Birmingham NHS Foundation Trust and University of Birmingham , Birmingham , UK.,b Centre for Liver and Gastrointestinal Research, Institute of Immunology and Immunotherapy , University of Birmingham , Birmingham , UK.,c Liver Unit , University Hospitals Birmingham NHS Foundation Trust , Birmingham , UK
| | - Philip Newsome
- a NIHR Birmingham Biomedical Research Centre , University Hospitals Birmingham NHS Foundation Trust and University of Birmingham , Birmingham , UK.,b Centre for Liver and Gastrointestinal Research, Institute of Immunology and Immunotherapy , University of Birmingham , Birmingham , UK.,c Liver Unit , University Hospitals Birmingham NHS Foundation Trust , Birmingham , UK
| | - Wei-Yu Lu
- b Centre for Liver and Gastrointestinal Research, Institute of Immunology and Immunotherapy , University of Birmingham , Birmingham , UK
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Hickey RD, Naugler WE. Ectopic expansion of engineered human liver tissue seeds using mature cell populations. Hepatology 2018; 67:2465-2467. [PMID: 29222915 DOI: 10.1002/hep.29718] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/09/2017] [Revised: 12/04/2017] [Accepted: 12/04/2017] [Indexed: 12/07/2022]
Affiliation(s)
- Raymond D Hickey
- Department of Surgery, Department of Molecular Medicine, Mayo Clinic, Rochester, MN
| | - Willscott E Naugler
- Department of Medicine, Division of Gastroenterology and Hepatology, Oregon Health & Science Center, Portland, OR; Oregon Stem Cell Center, Oregon Health & Science Center, Portland, OR
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Gupta S, Kim S, Vemuru R, Aragona E, Yerneni P, Burk R, Rha C. Hepatocyte Transplantation: An Alternative System for Evaluating Cell Survival and Immunoisolation. Int J Artif Organs 2018. [DOI: 10.1177/039139889301600310] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/29/2023]
Abstract
To evaluate systems for barrier immunoisolation of transplanted hepatocytes, we used transgenic mouse hepatocytes that secrete HBsAg. Hepatocytes were rapidly encapsulated in chitosan, a cationic polymer derived by deacetylation of chitin. Chitosan was allowed to electrostatically bond with anionic sodium alginate for creating an outer bipolymer membrane of the capsules. After encapsulation, hepatocyte viability remained unchanged for seven days in vitro with secretion of HBsAg into the culture medium throughout this period. Following intraperitoneal transplantation of encapsulated hepatocytes, HBsAg promptly appeared in blood of recipients. In congeneic recipients, serum HBsAg peaked at two weeks. Hepatocytes were present in recovered chitosan capsules and expressed HBsAg mRNA. In allogeneic recipients, however, serum HBsAg disappeared within one week and recovered chitosan capsules showed lymphomononuclear cells but not hepatocytes. Transplantation of chitosan encapsulatd HbsAg secreting hepatocytes failed to induce an anti-HBs response, suggesting modulation of the host immune response. These results indicate that transplantation systems using genetically modified hepatocytes which secrete gene products in the blood of recipients should facilitate evaluation of hepatocyte encapsulation.
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Affiliation(s)
- S. Gupta
- Departments of Medicine, Obstetrics and Gynecology and the Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, NY
| | - S.K. Kim
- Biomaterials Science and Engineering Laboratory, Massachusetts Institute of Technology, Cambridge, MA - USA
| | - R.P. Vemuru
- Departments of Medicine, Obstetrics and Gynecology and the Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, NY
| | - E. Aragona
- Departments of Medicine, Obstetrics and Gynecology and the Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, NY
| | - P.R. Yerneni
- Departments of Medicine, Obstetrics and Gynecology and the Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, NY
| | - R.D. Burk
- Pediatrics, Microbiology and Immunology, Obstetrics and Gynecology and the Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, NY
| | - C.K. Rha
- Biomaterials Science and Engineering Laboratory, Massachusetts Institute of Technology, Cambridge, MA - USA
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Stange J, Mitzner S. A Carrier-Mediated Transport of Toxins in a Hybrid Membrane. Safety Barrier between a Patients Blood and a Bioartificial Liver. Int J Artif Organs 2018. [DOI: 10.1177/039139889601901109] [Citation(s) in RCA: 100] [Impact Index Per Article: 14.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022]
Abstract
Combination of detoxifying liver support systems with liver cell bioreactors may have additional benefits for the treatment of liver failure due to the replacement of known and unknown metabolic activities of the liver. However, the problem of side effects and possible risks caused by the use of animal hepatocytes or hepatoma cells remains unsolved which underlines the need of a safety barrier between the patients blood and the extracorporeal bioreactor. Passive filters do not meet the requirements of such membranes, because in liver failure desired and undesired molecules in the patients blood share similar physicochemical properties. That challanges the developement of biologically designed separation membranes. A hybrid membrane is formed by implementation of transport proteins into a highly permeable hollow fiber. The transport of free solutes and albumin bound toxins is tested in vitro in comparison with conventional high flux membranes. The transport characteristics for tightly albumin bound toxins are significantly improved for the hybrid membrane. The transport of albumin bound toxins across the membrane is not associated with albumin. The selectivity of the transport is evaluated in vivo. No significant loss of middle molecular weight hormones attached to other carrier proteins was observed. Neither transport of immunologically relevant proteins across the membrane nor loss of valuable proteins was measured. Also in vivo, a significant reduction of protein bound toxins and a transport of metabolically relevant solutes, like amino acids, was shown. The presented hybrid membrane may be used like an “intellegent membrane” as a safety barrier between the patients blood and cell devices.
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Affiliation(s)
- J. Stange
- Department of Internal Medicine/HII, University of Rostock, Rostock - Germany
| | - S. Mitzner
- Department of Internal Medicine/HII, University of Rostock, Rostock - Germany
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Stange J, Mitzner S. Hepatocyte Encapsulation - Initial Intentions and New Aspects for Its Use in Bioartificial Liver Support. Int J Artif Organs 2018. [DOI: 10.1177/039139889601900107] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022]
Affiliation(s)
- J. Stange
- Department of Internal Medicine, University of Rostock, Rostock - Germany
| | - S. Mitzner
- Department of Internal Medicine, University of Rostock, Rostock - Germany
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12
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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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13
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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: 28] [Impact Index Per Article: 3.5] [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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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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15
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Markus PM, Koenig S, Krause P, Becker H. Selective Intraportal Transplantation of DiI-marked Isolated Rat Hepatocytes. Cell Transplant 2017; 6:455-62. [PMID: 9331496 DOI: 10.1177/096368979700600504] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
Abstract
Transplantation of isolated hepatocytes is a promising alternative to orthotopic liver transplantation in experimental animal models with acute hepatic failure and hereditary enzyme defects. Conventional light microscopy identification of hepatocytes within recipient livers has been limited due to the inability to distinguish between donor and recipient liver cells. In this study, we labeled hepatocytes intracellularly with the fluorescent dye DiI-18 prior to selective intraportal or intrasplenic transplantation. Syngeneic LEW rat hepatocytes were isolated and 2 × 107 fluorescence-labeled cells were transplanted by intraportal infusion selectively into 2/3 of the recipient liver lobules to avoid lethal portal hypertension. Rats were sacrificed on postop days 1, 3, 5, 10, 20, and 40. Histological examination was performed using light and fluorescence microscopy counterstained by light green dye. The quantity of transplanted hepatocytes residing within the recipient liver was determined by FACS analysis after enzymatic digestion of the recipient liver lobules. Engrafted hepatocytes were identified in the periportal regions of transplanted liver lobules. The stained hepatocytes were retrieved up to 20 days postop using fluorescent microscopy. Using FACS analysis the number of labeled hepatocytes was found to diminish over time following transplantation from 2.1% on postop day 1 to 0.5% on day 10. Labeled hepatocytes transplanted into the spleen were retrieved in clusters up to 20 days postop (the last day of observation). Furthermore, the migration of labeled hepatocytes from spleen to liver parenchyma was observed following intrasplenic transplantation. However, after selective intraportal transplantation, only fluorescent debris was found in splenic and pulmonary tissue upon examination of various organs. This article describes the method of fluorescent labeling of rat hepatocytes and reports the feasibility and limitations of using DiI-18 as a marker.
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Affiliation(s)
- P M Markus
- Department of General Surgery, Georg August University of Göttingen, Germany
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16
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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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17
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Markus PM, Krause P, Fayyazi A, Honnicke K, Becker H. Allogeneic Hepatocyte Transplantation Using FK 506. Cell Transplant 2017; 6:77-83. [PMID: 9040958 DOI: 10.1177/096368979700600112] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022] Open
Abstract
Hepatocyte transplantation is an intriguing alternative to orthotopic liver transplantation. While engraftment of syngeneic hepatocytes can be achieved with relative ease, engraftment of allogeneic hepatocytes has been far more complicated. We used FK 506 (Tacrolimus), a novel and highly efficient immunosuppressant, which has been reported to augment liver regeneration in rats. Recipients of isolated syngeneic (LEW) and allogeneic (Wistar F.) rat hepatocytes (major histocompatibility barrier) recieved different immunosuppressive regiments with FK 506 or Cyclosporine A (CsA). Mature syngeneic hepatocytes could be retrieved up to post op day 300 with the lowest number of hepatocytes on post op day 20. Following allogeneic transplantation, no mature hepatocytes could be identified after post op day 10, though ductular like structures within the spleen were found in FK 506 but not CsA-treated animals. The epithelial cells of ductular like structures exhibit cytological features of CK-19 positive cells. Our results suggest that under CsA or FK 506 immunosuppression long-term survival of mature allogeneic hepatocytes within the spleen cannot be achieved across a major histocompatibility barrier though FK 506 allows engraftment of allogeneic donor type ductular cells. Copyright © 1997 Elsevier Science Inc.
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Affiliation(s)
- P M Markus
- Department of General Surgery and Pathology, Georg August University of Göttingen, Germany
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18
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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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19
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Vogels BA, Maas MA, Bosma A, Chamuleau RA. Significant Improvement of Survival by Intrasplenic Hepatocyte Transplantation in Totally Hepatectomized Rats. Cell Transplant 2017; 5:369-78. [PMID: 8727005 DOI: 10.1177/096368979600500303] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022] Open
Abstract
The effect of intrasplenic hepatocyte transplantation (HTX) was studied in an experimental model of acute liver failure in rats with chronic liver atrophy. Rats underwent a portacaval shunt operation on Day -14 to induce liver atrophy, and underwent total hepatectomy on Day 0 as a start of acute liver failure. Intrasplenic hepatocyte or sham transplantation was performed on Day -7, -3, or -1 (n = 4 to 6 per group). During the period following hepatectomy, mean arterial blood pressure was maintained above 80 mm Hg and hypoglycaemia was prevented. Severity of hepatic encephalopathy was assessed by clinical grading and EEG spectral analysis, together with determination of blood ammonia and plasma amino acid concentrations, and “survival” time. Histological examination of the spleen and lungs was performed after sacrifice. Intrasplenic hepatocyte transplantation resulted in a significant improvement in clinical grading in all transplanted groups (p < 0.05), whereas a significant improvement in EEG left index was seen only in the group with transplantation on Day -1 (p < 0.05). In contrast to hepatocyte transplantation 1 day before total hepatectomy, rats with hepatocyte transplantation 3 and 7 days before total hepatectomy showed a significant 3- and 2-fold increase in “survival” time compared to sham transplanted controls: HTX at Day -1: 7.5 ± 0.3 h vs. 5.9 ± 0.6 h (p > 0.05), HTX at Day -3:19.7 ± 3.7 h vs. 6.5 ± 0.3 h (p < 0.05), and HTX at Day -7: 13.8 ± 3.2 h vs. 6.3 ± 0.3 h (p < 0.05). Furthermore, rats with hepatocyte transplantation on Day -3 and -7 showed significantly lower blood ammonia concentrations after total hepatectomy (p < 0.0001). Histological examination of the spleens after sacrifice showed clusters of hepatocytes in the red pulp. Hepatocytes present in the spleen for 3 and 7 days showed bile accumulation and spots of beginning necrosis. The present data show that in a hard model of complete liver failure in portacaval shunted rats, intrasplenic hepatocyte transplantation is able to prolong “survival” time significantly 2- to 3-fold. The relevance of this observation for human application is discussed.
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Affiliation(s)
- B A Vogels
- University of Amsterdam, J. van Gool Laboratory for Experimental Internal Medicine, The Netherlands
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20
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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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21
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Kita S, Yasuchika K, Ishii T, Katayama H, Yoshitoshi EY, Ogiso S, Kawai T, Yasuda K, Fukumitsu K, Mizumoto M, Uemoto S. The Protective Effect of Transplanting Liver Cells Into the Mesentery on the Rescue of Acute Liver Failure After Massive Hepatectomy. Cell Transplant 2016; 25:1547-59. [PMID: 26883767 DOI: 10.3727/096368916x690999] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Postoperative liver failure is one of the most critical complications following extensive hepatectomy. Although transplantation of allogeneic hepatocytes is an attractive therapy for posthepatectomy liver failure, transplanting cells via the portal veins typically causes portal vein embolization. The embolization by transplanted cells would be lethal in patients who have undergone massive hepatectomy. Thus, transplant surgeons need to select extrahepatic sites as transplant sites to prevent portal vein embolization. We aimed to investigate the mechanism of how liver cells transplanted into the mesentery protect recipient rats from acute liver failure after massive hepatectomy. We induced posthepatectomy liver failure by 90% hepatectomy in rats. Liver cells harvested from rat livers were transplanted into the mesenteries of hepatectomized rats. Twenty percent of the harvested cells, which consisted of hepatocytes and nonparenchymal cells, were transplanted into each recipient. The survival rate improved significantly in the liver cell transplantation group compared to the control group 7 days after hepatectomy (69 vs. 7%). Histological findings of the transplantation site, in vivo imaging system study findings, quantitative polymerase chain reaction assays of the transplanted cells, and serum albumin measurements of transplanted Nagase analbuminemic rats showed rapid deterioration of viable transplanted cells. Although viable transplanted cells deteriorated in the transplanted site, histological findings and an adenosine-5'-triphosphate (ATP) assay showed that the transplanted cells had a protective effect on the remaining livers. These results indicated that the paracrine effects of transplanted liver cells had therapeutic effects. The same protective effects were observed in the hepatocyte transplantation group, but not in the liver nonparenchymal cell transplantation group. Therefore, this effect on the remnant liver was mainly due to the hepatocytes among the transplanted liver cells. We demonstrated that transplanted liver cells protect the remnant liver from severe damage after massive hepatectomy.
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Affiliation(s)
- Sadahiko Kita
- Department of Surgery, Graduate School of Medicine, Kyoto University, Kyoto, Japan
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22
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Lee JW, Choi YJ, Yong WJ, Pati F, Shim JH, Kang KS, Kang IH, Park J, Cho DW. Development of a 3D cell printed construct considering angiogenesis for liver tissue engineering. Biofabrication 2016; 8:015007. [DOI: 10.1088/1758-5090/8/1/015007] [Citation(s) in RCA: 152] [Impact Index Per Article: 16.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
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23
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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: 47] [Impact Index Per Article: 4.7] [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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24
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Thrombomodulin improves rat survival after extensive hepatectomy. J Surg Res 2015; 194:375-382. [DOI: 10.1016/j.jss.2014.10.048] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2014] [Revised: 10/11/2014] [Accepted: 10/29/2014] [Indexed: 01/08/2023]
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25
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Forbes SJ, Gupta S, Dhawan A. Cell therapy for liver disease: From liver transplantation to cell factory. J Hepatol 2015; 62:S157-69. [PMID: 25920085 DOI: 10.1016/j.jhep.2015.02.040] [Citation(s) in RCA: 218] [Impact Index Per Article: 21.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/29/2014] [Revised: 02/20/2015] [Accepted: 02/27/2015] [Indexed: 02/08/2023]
Abstract
Work over several decades has laid solid foundations for the advancement of liver cell therapy. To date liver cell therapy in people has taken the form of hepatocyte transplantation for metabolic disorders with a hepatic basis, and for acute or chronic liver failure. Although clinical trials using various types of autologous cells have been implemented to promote liver regeneration or reduce liver fibrosis, clear evidence of therapeutic benefits have so far been lacking. Cell types that have shown efficacy in preclinical models include hepatocytes, liver sinusoidal endothelial cells, mesenchymal stem cells, endothelial progenitor cells, and macrophages. However, positive results in animal models have not always translated through to successful clinical therapies and more realistic preclinical models need to be developed. Studies defining the optimal repopulation by transplanted cells, including routes of cell transplantation, superior engraftment and proliferation of transplanted cells, as well as optimal immunosuppression regimens are required. Tissue engineering approaches to transplant cells in extrahepatic locations have also been proposed. The derivation of hepatocytes from pluripotent or reprogrammed cells raises hope that donor organ and cell shortages could be overcome in the future. Critical hurdles to be overcome include the production of hepatocytes from pluripotent cells with equal functional capacity to primary hepatocytes and long-term phenotypic stability in vivo.
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Affiliation(s)
- Stuart J Forbes
- MRC Centre for Regenerative Medicine, Scottish Centre for Regenerative Medicine, 5 Little France Drive, Edinburgh EH16 4UU, United Kingdom.
| | - Sanjeev Gupta
- Departments of Medicine and Pathology, Albert Einstein College of Medicine, Jack and Pearl Resnick Campus, 1300 Morris Park Avenue, Ullmann Building, Room 625, Bronx, NY 10461, United States
| | - Anil Dhawan
- Paediatric Liver GI and Nutrition Center and NIHR/Wellcome Cell Therapy Unit, King's College Hospital at King's College, London SE59RS, United Kingdom
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26
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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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27
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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: 213] [Impact Index Per Article: 19.4] [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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28
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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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29
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Wang Y, Phillips CN, Herrera GS, Sims CE, Yeh JJ, Allbritton NL. Array of Biodegradable Microraftsfor Isolation and Implantation of Living, Adherent Cells. RSC Adv 2013; 3:9264-9272. [PMID: 23930219 PMCID: PMC3733277 DOI: 10.1039/c3ra41764f] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022] Open
Abstract
A new strategy for efficient sorting and implantation of viable adherent cells into animals is described. An array of biodegradable micro-structures (microrafts) was fabricated using a polydimethylsiloxane substrate for micromolding poly(lactic-co-glycolic acid) (PLGA). Screening various forms of PLGA determined that the suitability of PLGA for microraft manufacture, biocompatibility and in vitro degradation was dependent on molecular weight and lactic/glycolic ratio. Cells plated on the array selectively attached to the microrafts and could be identified by their fluorescence, morphology or other criteria. The cells were efficiently dislodged and collected from the array using a microneedle device. The platform was used to isolate specific cells from a mixed population establishing the ability to sort target cells for direct implantation. As a proof of concept, fluorescently conjugated microrafts carrying tumor cells stably expressing luciferase were isolated from an array and implanted subcutaneously into mice. In vivo bio-luminescence imaging confirmed the growth of a tumor in the recipient animals. Imaging of tissue sections from the tumors demonstrated in vivo degradation of the implanted microrafts. The process is a new strategy for isolating and delivering a small number of adherent cells for animal implantation with potential applications in tissue repair, tumor induction, in vivo differentiation of stem cells and other biomedical research.
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Affiliation(s)
- Yuli Wang
- Department of Chemistry, University of North Carolina, Chapel Hill, NC 27599
| | - Colleen N. Phillips
- Department of Chemistry, University of North Carolina, Chapel Hill, NC 27599
| | - Gabriela S. Herrera
- Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599
| | - Christopher E. Sims
- Department of Chemistry, University of North Carolina, Chapel Hill, NC 27599
| | - Jen Jen Yeh
- Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599
- Departments of Surgery and Pharmacology, University of North Carolina, Chapel Hill, NC 27599
| | - Nancy L. Allbritton
- Department of Chemistry, University of North Carolina, Chapel Hill, NC 27599
- Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599
- Department of Biomedical Engineering, University of North Carolina, Chapel Hill, NC 27599 and North Carolina State University, Raleigh, NC 27695
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30
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Palakkan AA, Hay DC, Anil Kumar PR, Kumary TV, Ross JA. Liver tissue engineering and cell sources: issues and challenges. Liver Int 2013; 33:666-76. [PMID: 23490085 DOI: 10.1111/liv.12134] [Citation(s) in RCA: 73] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/11/2012] [Accepted: 01/27/2013] [Indexed: 02/13/2023]
Abstract
Liver diseases are of major concern as they now account for millions of deaths annually. As a result of the increased incidence of liver disease, many patients die on the transplant waiting list, before a donor organ becomes available. To meet the huge demand for donor liver, alternative approaches using liver tissue engineering principles are being actively pursued. Even though adult hepatocytes, the primary cells of the liver are most preferred for tissue engineering of liver, their limited availability, isolation from diseased organs, lack of in vitro propagation and deterioration of function acts as a major drawback to their use. Various approaches have been taken to prevent the functional deterioration of hepatocytes including the provision of an adequate extracellular matrix and co-culture with non-parenchymal cells of liver. Great progress has also been made to differentiate human stem cells to hepatocytes and to use them for liver tissue engineering applications. This review provides an overview of recent challenges, issues and cell sources with regard to liver tissue engineering.
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Affiliation(s)
- Anwar A Palakkan
- Tissue Injury and Repair Group, University of Edinburgh - MRC Centre for Regenerative Medicine, Edinburgh, UK
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Rozga J, Morsiani E, Lepage E, Moscioni AD, Demetriou AA, Giorgio T. Isolated hepatocytes in a bioartificial liver: A single group view and experience. Biotechnol Bioeng 2012; 43:645-53. [PMID: 18615764 DOI: 10.1002/bit.260430714] [Citation(s) in RCA: 56] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
Despite recent advances in medical supportive therapy, patients with severe fulminant hepatic failure (FHF) have mortality rate approaching 90%. Investigators have attempted to improve survival by using various extracorporeal liver support systems loaded with sorbents and liver tissue preparations. None of them succeeded in gaining clinical acceptance and orthotopic liver transplantation (OLT) remains a primary therapeutic option for patients with FHF. In this study, authors discuss the systems which utilize isolated hepatocytes. Most of these devices were tested in vitro and in animals with chemically and surgically induced liver failure. In some studies, signficant levels of detoxification and liver functions were achieved. The authors describe their own hepatocyte-based artificial liver (BAL). It is based on plasma perfusion through a hollow-fiber module seeded with matrix-anchored porcine hepatocytes. The BAL was used 14 times to treat 9 patients with acute liver failure. On 10 occasions, a charcoal column was included in the plasma circuit. Each treatment lasted 7 +/- 1 h. All procedures were tolerated well and 8 patients (including 6 patients with FHF) underwent OLT. Five patients with increased intracranial pressure (ICP) and evidence of decerebration had normalization of ICP and enjoyed full neurologic recovery after OLT. Laboratory data showed evidence for bilirubin conjugation, decrease in blood ammonia, maintenance of low lactic acid levels, and increase in the ration between the branched chain and aromatic amino acids. No allergic reactions to xenogeneic hepatocytes were observed. The authors conclude that BAL treatment with porcine hepatocytes appears to be safe and can help maintain patients alive and neurologically intact until a liver becomes available for transplantation. (c) 1994 John Wiley & Sons, Inc.
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Affiliation(s)
- J Rozga
- Department of Surgery and Liver Support Unit, Cedars-Sinai Medical Center, Room 8215, North Tower, 870 Beverly Boulevard, Los Angeles, California 90048
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Abstract
Despite recent advances in medical therapy, patients with fulminant hepatic failure (FHF) have a mortality rate approaching 90%. Many patients die because of failure to arrest the progression of cerebral edema. Liver transplantation has improved survival to 65% to 75%. However, there is a shortage of donors and approximately one half of the patients with FHF will die while awaiting liver transplantation. There is thus a need to develop an extracorporeal liver assist system to help keep these patients alive and neurologically intact until either an organ becomes available for transplantation or the native liver recovers from injury. Such a system could also be used during the period of functional recovery from massive liver resection or to assist patients with decompensated chronic liver disease. Over the years, various methods utilizing charcoal and resin hemoperfusion, dialysis, plasma exchange, and other methods of blood detoxification have been developed and tested, but none have gained wide acceptance. This was due to: (i) incomplete understanding of the pathophysiology of liver failure; (ii) lack of accurate methods of assessment, quantitation, and stratification of the degree of liver dysfunction; and (iii) inadequate numbers of prospective controlled clinical trials examining the effects of specific therapeutic modalities. Liver support systems utilizing liver tissue preparations were developed in the 1950s, but it was not until recently that advances in hepatocyte isolation and culture, better understanding of hepatocyte-matrix interactions, and improved hollow-fiber technology have resulted in the development of a new generation of liver assist devices. Some of these devices are currently being tested in the clinical setting. In a preliminary clinical study, we have used a porcine hepatocyte-based liver support system to treat patients with acute liver failure as well as patients with acute exacerbation of chronic liver disease. Patients in the first group, who were candidates for transplantation, were successfully bridged to a transplant with excellent survival. No obvious benefit from bioartifical liver treatments was seen in the second group. It is possible that, in this group, patients will have to be treated earlier and for longer periods of time. Prospective controlled trials will be initiated as soon as the current phase I study is concluded to determine the efficacy of this system in both patients populations. (c) 1996 John Wiley & Sons, Inc.
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Affiliation(s)
- A Kamlot
- Department of Surgery and Liver Support Unit, Cedars-Sinai Medical Center, 8700 Beverly Boulevard, D-4018, Los Angeles, California 90048
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Hammond JS, Lobo DN. Emerging roles for biomaterials in the treatment of liver disease. Expert Rev Med Devices 2012; 9:181-8. [PMID: 22404778 DOI: 10.1586/erd.11.70] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
Abstract
This review explores potential roles for biomaterials in the field of liver surgery and hepatology. The studies reviewed are presented in three sections. The first section discusses liver regeneration and strategies to modulate it. The second section outlines the pathophysiology of liver inflammation and fibrosis and highlights novel therapeutic targets. The final section summarises the current challenges in liver surgery and discusses how biomaterials may be used to address these challenges and focuses on early translational applications for biomaterials for drug delivery and liver surgery.
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Affiliation(s)
- John S Hammond
- Division of Gastrointestinal Surgery, Nottingham Digestive Diseases Centre NIHR Biomedical Research Unit, Nottingham University Hospitals, Queen's Medical Centre, Nottingham NG7 2UH, UK.
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The instant blood-mediated inflammatory reaction characterized in hepatocyte transplantation. Transplantation 2011; 91:632-8. [PMID: 21289595 DOI: 10.1097/tp.0b013e31820ae459] [Citation(s) in RCA: 72] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/18/2023]
Abstract
BACKGROUND Hepatocyte transplantation (HcTx) has proven to be a safe procedure, although the functional results have been unsatisfactory, probably due to insufficient engraftment or a loss of transplanted mass or function. In this study, we investigate whether hepatocytes in contact with blood induce an inflammatory reaction leading to, similar to what happens in clinical islet transplantation, an instant blood-mediated inflammatory reaction (IBMIR) resulting in an early loss of transplanted cells. METHODS By using an experimental model that mimics the portal vein blood flow, we could study different parameters reflecting the effects on the innate immunity elicited by hepatocytes in contact with ABO-matched human blood. RESULTS We report that all aspects of the IBMIR such as platelet and granulocyte consumption, coagulation, and complement activation were demonstrated. Addition of various specific inhibitors of coagulation allowed us to clearly delineate the various stages of the hepatocyte-triggered IBMIR and show that the reaction was triggered by tissue factor. Analysis of a case of clinical HcTx showed that hepatocyte-induced IBMIR also occurs in vivo. Both the inflammatory and the coagulation aspects were controlled by low-molecular-weight dextran sulfate. CONCLUSION Isolated hepatocytes in contact with blood induce the IBMIR in vitro, and there are indications that these events are also relevant in vivo. According to these findings, HcTx would benefit from controlling a wider range of signals from the innate immune system.
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Park JH, Yoon YC, Hong TH, You YK, Kim DG. Effect of Liver Cell Transplantation on Acute Hepatic Failure Induced by Massive Liver Resection in the Rat. KOREAN JOURNAL OF HEPATO-BILIARY-PANCREATIC SURGERY 2011; 15:107-16. [PMID: 26421025 PMCID: PMC4582550 DOI: 10.14701/kjhbps.2011.15.2.107] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 01/19/2011] [Accepted: 03/20/2011] [Indexed: 01/13/2023]
Abstract
Purpose This study is designed to ascertain the most effective quantity and injection route of hepatocytes in an acute liver failure model induced by massive liver resection in rats. Methods Rats weighing 450 to 650 gm underwent partial hepatectomy that was 80% of their liver weight, resulting in acute liver failure. Hepatocytes were obtained by perfusing collagenase (Wako, Japan) solution through portal vein into liver of the allogenic rat. These hepatocytes were injected into different places with different dosage. The experimental groups were divided into the Control group, Splenic group I (2×106 cells into splenic capsule), Splenic group II (2×107 cells into splenic capsule), Portal vein group (2×107 cells into portal vein), Subperitoneal group (2×107 cells into subperitoneum). The experimental animals were observed carefully for 5 days for assessment of survival and regeneration of liver. Liver function tests including serum alanine aminotransferase (ALT), total bilirubin, gamma-glutamyl transferase (γ-GTP) on postoperative 1, 2, 3, 5th days and histologic examinations of specimens obtained from each respective groups on postoperative 5th day were performed. Results Serum ALT level on postoperative day 1 peaked and then gradually normalized showing statistical significance (p=0.035). Study groups showing statistically significant difference under repeated anova analysis were between the Splenic group II and Control (p=0.035), and between the Splenic group II and Portal vein group (p=0.001) with respect to serum ALT levels. Also, progression of each study group showed statistical significance. (p=0.02). Serum total bilirubin and r-GTP did not show any significant difference. Conclusion Hepatocyte transplantation of 2×107 cells into spleen showed the best results in the acute hepatic failure rat.
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Affiliation(s)
- Jung Hyun Park
- Department of Surgery, Seoul St. Mary's Hospital, The Catholic University of Korea, Korea
| | - Young Chul Yoon
- Department of Surgery, Seoul St. Mary's Hospital, The Catholic University of Korea, Korea
| | - Tae Ho Hong
- Department of Surgery, Seoul St. Mary's Hospital, The Catholic University of Korea, Korea
| | - Young Kyoung You
- Department of Surgery, Seoul St. Mary's Hospital, The Catholic University of Korea, Korea
| | - Dong Goo Kim
- Department of Surgery, Seoul St. Mary's Hospital, The Catholic University of Korea, Korea
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Schrobback K, Klein TJ, Schuetz M, Upton Z, Leavesley DI, Malda J. Adult human articular chondrocytes in a microcarrier-based culture system: expansion and redifferentiation. J Orthop Res 2011; 29:539-46. [PMID: 20957734 DOI: 10.1002/jor.21264] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/21/2010] [Accepted: 08/18/2010] [Indexed: 02/04/2023]
Abstract
Expanding human chondrocytes in vitro while maintaining their ability to form cartilage remains a key challenge in cartilage tissue engineering. One promising approach to address this is to use microcarriers as substrates for chondrocyte expansion. While microcarriers have shown beneficial effects for expansion of animal and ectopic human chondrocytes, their utility has not been determined for freshly isolated adult human articular chondrocytes. Thus, we investigated the proliferation and subsequent chondrogenic differentiation of these clinically relevant cells on porous gelatin microcarriers and compared them to those expanded using traditional monolayers. Chondrocytes attached to microcarriers within 2 days and remained viable over 4 weeks of culture in spinner flasks. Cells on microcarriers exhibited a spread morphology and initially proliferated faster than cells in monolayer culture, however, with prolonged expansion they were less proliferative. Cells expanded for 1 month and enzymatically released from microcarriers formed cartilaginous tissue in micromass pellet cultures, which was similar to tissue formed by monolayer-expanded cells. Cells left attached to microcarriers did not exhibit chondrogenic capacity. Culture conditions, such as microcarrier material, oxygen tension, and mechanical stimulation require further investigation to facilitate the efficient expansion of clinically relevant human articular chondrocytes that maintain chondrogenic potential for cartilage regeneration applications. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:539-546, 2011.
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Affiliation(s)
- Karsten Schrobback
- Institute of Health and Biomedical Innovation, Queensland University of Technology, Kelvin Grove, Queensland, Australia.
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Hodgson H, Selden C. Liver cell implants: a long road. Liver Transpl 2011; 17:99-101. [PMID: 21280180 DOI: 10.1002/lt.22245] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/12/2023]
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Martin Y, Eldardiri M, Lawrence-Watt DJ, Sharpe JR. Microcarriers and Their Potential in Tissue Regeneration. TISSUE ENGINEERING PART B-REVIEWS 2011; 17:71-80. [PMID: 21083436 DOI: 10.1089/ten.teb.2010.0559] [Citation(s) in RCA: 97] [Impact Index Per Article: 6.9] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/19/2023]
Affiliation(s)
- Yella Martin
- Blond McIndoe Research Foundation, Queen Victoria Hospital, East Grinstead, West Sussex, United kingdom
| | - Mohamed Eldardiri
- Blond McIndoe Research Foundation, Queen Victoria Hospital, East Grinstead, West Sussex, United kingdom
| | - Diana J. Lawrence-Watt
- Blond McIndoe Research Foundation, Queen Victoria Hospital, East Grinstead, West Sussex, United kingdom
| | - Justin R. Sharpe
- Blond McIndoe Research Foundation, Queen Victoria Hospital, East Grinstead, West Sussex, United kingdom
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Sun LY, Lin SZ, Li YS, Harn HJ, Chiou TW. Functional Cells Cultured on Microcarriers for Use in Regenerative Medicine Research. Cell Transplant 2011; 20:49-62. [PMID: 20887678 DOI: 10.3727/096368910x532792] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023] Open
Abstract
Microcarriers have been successfully used for many years for growing anchorage-dependent cells and as a means of delivering cells for tissue repair. When cultured on microcarriers, the number of anchorage-dependent cells, including primary cells, can easily be scaled up and controlled to generate the quantities of cells necessary for therapeutic applications. Recently, stem cell technology has been recognized as a powerful tool in regenerative medicine, but adequate numbers of stem cells that retain their differentiation potential are still difficult to obtain. For anchorage-dependent stem cells, however, microcarrier-based suspension culture using various types of microcarriers has proven to be a good alternative for effective ex vivo expansion. In this article, we review studies reporting the expansion, differentiation, or transplantation of functional anchorage-dependent cells that were expanded with the microcarrier culture system. Thus, the implementation of technological advances in biodegradable microcarriers, the bead-to-bead transfer process, and appropriate stem cell media may soon foster the ability to produce the numbers of stem cells necessary for cell-based therapies and/or tissue engineering.
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Affiliation(s)
- Li-Yi Sun
- Department of Biological Science and Technology, National Chiao Tung University, Hsinchu, Taiwan
| | - Shinn-Zong Lin
- Center for Neuropsychiatry, China Medical University and Hospital and Beigang Hospital, Taichung and Yun-Lin, Taiwan
| | - Yuan-Sheng Li
- Department of Life Science and Graduate Institute of Biotechnology, National Dong Hwa University, Hualien, Taiwan
| | - Horng-Jyh Harn
- Department of Pathology, China Medical University and Hospital, Taichung, Taiwan
- Department of Medicine, China Medical University, Taichung, Taiwan
| | - Tzyy-Wen Chiou
- Department of Life Science and Graduate Institute of Biotechnology, National Dong Hwa University, Hualien, Taiwan
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Li J, Tao R, Wu W, Cao H, Xin J, Li J, Guo J, Jiang L, Gao C, Demetriou AA, Farkas DL, Li L. 3D PLGA scaffolds improve differentiation and function of bone marrow mesenchymal stem cell-derived hepatocytes. Stem Cells Dev 2011; 19:1427-36. [PMID: 20055663 DOI: 10.1089/scd.2009.0415] [Citation(s) in RCA: 78] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
UNLABELLED Liver tissue engineering with hepatic stem cells provides a promising alternative to liver transplantation in patients with acute and chronic hepatic failure. In this study, a three-dimensional (3D) bioscaffold was introduced for differentiation of rat bone marrow mesenchymal stem cells (BMSCs) into hepatocytes. For hepatocyte differentiation, third passage BMSCs isolated from normal adult F344 rats were seeded into collagen-coated poly(lactic-co-glycolic acid) (C-PLGA) 3D scaffolds with hepatocyte differentiation medium for 3 weeks. Hepatogenesis in scaffolds was characterized by reverse transcript PCR, western blot, confocal laser scanning microscopy (CLSM), periodic acid-Schiff staining, histochemistry, and biochemical assays with hepatic-specific genes and markers. A monolayer culture system was used as a control differentiation group. The results showed that isolated cells possessed the basic features of BMSCs. Differentiated hepatocyte-like cells in C-PLGA scaffolds expressed hepatocyte-specific markers [eg, albumin (ALB), alpha-fetoprotein, cytokeratin 18, hepatocyte nuclear factor 4alpha, and cytochrome P450] at mRNA and protein levels. Most markers were expressed in C-PLGA group 1 week earlier than in the control group. Results of biocompatibility indicated that the differentiated hepatocyte-like cells grew more stably in C-PLGA scaffolds than that in controls during a 3-week differentiation period. The significantly higher metabolic functions in hepatocyte-like cells in the C-PLGA scaffold group further demonstrated the important role of the scaffold. CONCLUSION As the phenomenon of transdifferentiation is uncommon, our successful transdifferentiation rates of BMSCs to mature hepatocytes prove the superiority of the C-PLGA scaffold in providing a suitable environment for such a differentiation. This material can possibly be used as a bioscaffold for liver tissue engineering in future clinical therapeutic applications.
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Affiliation(s)
- Jun Li
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Zhejiang University, Hangzhou, China
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Abstract
In vitro hepatocyte models represent very useful systems in both fundamental research and various application areas. Primary hepatocytes appear as the closest model for the liver in vivo. However, they are phenotypically unstable, have a limited life span and in addition, exhibit large interdonor variability when of human origin. Hepatoma cell lines appear as an alternative but only the HepaRG cell line exhibits various functions, including major cytochrome P450 activities, at levels close to those found in primary hepatocytes. In vitro hepatocyte models have brought a substantial contribution to the understanding of the biochemistry, physiology, and cell biology of the normal and diseased liver and in various application domains such as xenobiotic metabolism and toxicity, virology, parasitology, and more generally cell therapies. In the future, new well-differentiated hepatocyte cell lines derived from tumors or from either embryonic or adult stem cells might be expected and although hepatocytes will continue to be used in various fields, these in vitro liver models should allow marked advances, especially in cell-based therapies and predictive and mechanistic hepatotoxicity of new drugs and other chemicals. All models will benefit from new developments in throughput screening based on cell chips coupled with high-content imaging and in toxicogenomics technologies.
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Abstract
Over the last decade the interest in hepatocyte transplantation has been growing continuously and this treatment may represent an alternative clinical approach for patients with acute liver failure and life-threatening liver-based metabolic disorders. The technology also serves as the proof of concept and reference for future development in stem cell technology. This chapter reviews the field of hepatocyte transplantation from bench to bedside.
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Affiliation(s)
- Anil Dhawan
- King's Cell Isolation Unit, King's College Hospital, London, UK
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Foy BD, Lee J, Morgan J, Toner M, Tompkins RG, Yarmush ML. Optimization of hepatocyte attachment to microcarriers: Importance of oxygen. Biotechnol Bioeng 2010; 42:579-88. [PMID: 18613079 DOI: 10.1002/bit.260420505] [Citation(s) in RCA: 32] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Many potential applications of primary hepatocytes cultured on microcarriers, such as an artificial liver or hepatocyte transplantation, would benefit from having a large number of hepatocytes attached to each microcarrier. In addition, the supply of primary hepatocytes is usually limited, so the efficient utilization of hepatocytes during attachment to microcarriers is necessary. Several physical parameters involved in the attachment process have been investigated, and the number of cells attached per microcarrier and the fraction of hepatocytes which attach have been quantitatively monitored. Variation of the partial pressure of gas phase oxygen in the incubation flask produced significant effects on the attachment of hepatocytes to microcarriers, with higher partial pressures of oxygen found to be necessary for attachment. In addition, variation of fluid depth and cell number, both of which influence the partial pressure of oxygen at the cell surface, affected hepatocyte attachment. The partial pressure of oxygen at the cell surface as a function of the physical parameters was analyzed using a simple one-dimensional theoretical model. Variations in the cell-to-microcarrier ratio used for incubation indicate that a compromise must be made in terms of maximizing the number of cells per microcarrier and the fraction of total hepatocytes which attach. The maximum number of hepatocytes per microcarrier obtained in this work was approximately 100. The best attachment fraction, defined as the ratio of the number of hepatocytes attached to the total number added to the incubation, was approximately 90%.
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Affiliation(s)
- B D Foy
- Surgical Services, Massachusetts General Hospital, Department of Surgery, Harvard Medical School, and the Shriners Burns Institute, Boston, Massachusetts, 02114, USA
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Transcriptional profiling and hepatogenic potential of acute hepatic failure-derived bone marrow mesenchymal stem cells. Differentiation 2010; 80:166-74. [PMID: 20427118 DOI: 10.1016/j.diff.2010.04.002] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2010] [Revised: 03/17/2010] [Accepted: 04/08/2010] [Indexed: 01/14/2023]
Abstract
UNLABELLED Liver stem cell (LSC) transplantation is a promising alternate approach to liver transplantation for patients with end-stage liver disease. However, the precise origin of LSCs remains unclear. Herein we determine if bone marrow mesenchymal stem cells (BMSCs) isolated from rats in acute hepatic failure (AHF) possess hepatic characteristics and have differentiation potential. BMSCs were isolated from AHF and sham-operated rats, and primary hepatocytes were isolated from normal rats for comparison. The transcriptomic profile of BMSCs and primary hepatocytes was analyzed using the Affymetrix GeneChip Rat Genome 230 2.0 Array. BMSCs isolated from AHF and normal rats were induced to differentiate into hepatocytes in vitro and the degree of hepatic differentiation was assessed using quantitative real time RT-PCR, immunohistochemistry, and biochemical assays. AHF-derived BMSCs had a significantly different gene expression profile compared to control BMSCs. Thirty-four gene/probe sets were expressed in both AHF-derived BMSCs and primary hepatocytes, but were absent in control-derived BMSCs, including 3 hepatocyte-specific genes. Forty-four genes were up-regulated more than 2-fold in AHF-derived BMSCs compared to controls, including 3 genes involved in hepatocyte metabolism and development. Furthermore, AHF-derived BMSCs expressed more hepatocyte related genes than control BMSCs. Additional experiments to validate the differentiation of AHF-derived BMSCs, compared to control-derived BMSCs, showed that several hepatocyte-specific genes and proteins [such as albumin (ALB) and alpha fetoprotein (AFP)] were expressed earlier, and at higher levels, after 1 week of differentiation. Hepatocyte-specific metabolic functions were also significantly higher in the AHF group compared to control cells. CONCLUSION AHF-derived BMSCs had a hepatic transcriptional profile and expressed hepatocyte specific genes early during differentiation, and possessed greater hepatogenic potency in vitro compared to cells isolated from control animals, further confirming their potential as a stem cell-based therapy for end-stage liver disease.
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Pettersson S, Wetterö J, Tengvall P, Kratz G. Human articular chondrocytes on macroporous gelatin microcarriers form structurally stable constructs with blood-derived biological glues in vitro. J Tissue Eng Regen Med 2009; 3:450-60. [PMID: 19444864 DOI: 10.1002/term.179] [Citation(s) in RCA: 31] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Biodegradable macroporous gelatin microcarriers fixed with blood-derived biodegradable glue are proposed as a delivery system for human autologous chondrocytes. Cell-seeded microcarriers were embedded in four biological glues-recalcified citrated whole blood, recalcified citrated plasma with or without platelets, and a commercially available fibrin glue-and cultured in an in vitro model under static conditions for 16 weeks. No differences could be verified between the commercial fibrin glue and the blood-derived alternatives. Five further experiments were conducted with recalcified citrated platelet-rich plasma alone as microcarrier sealant, using two different in vitro culture models and chondrocytes from three additional donors. The microcarriers supported chondrocyte adhesion and expansion as well as extracellular matrix (ECM) synthesis. Matrix formation occurred predominantly at sample surfaces under the static conditions. The presence of microcarriers proved essential for the glues to support the structural takeover of ECM proteins produced by the embedded chondrocytes, as exclusion of the microcarriers resulted in unstable structures that dissolved before matrix formation could occur. Immunohistochemical analysis revealed the presence of SOX-9- and S-100-positive chondrocytes as well as the production of aggrecan and collagen type I, but not of the cartilage-specific collagen type II. These results imply that blood-derived glues are indeed potentially applicable for encapsulation of chondrocyte-seeded microcarriers. However, the static in vitro models used in this study proved incapable of supporting cartilage formation throughout the engineered constructs.
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Affiliation(s)
- Sofia Pettersson
- Laboratory for Reconstructive Plastic Surgery, Department of Clinical and Experimental Medicine, Linköping University, Linköping SE-581 85, Sweden.
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Abstract
Hepatocyte transplantation is making its transition from bench to bedside for liver-based metabolic disorders and acute liver failure. Over eighty patients have now been transplanted world wide and the safety of the procedure together with medium-term success has been established. A major limiting factor in the field is the availability of good quality cells as hepatocytes are derived from grafts that are deemed unsuitable for transplantation. Alternative sources of cell, including stem cells may provide a sustainable equivalent to primary hepatocytes. There is also a need to develop techniques that will improve the engraftment, survival and function of transplanted hepatocytes. Such developments may allow hepatocyte transplantation to become an accepted and practical alternative to liver transplantation in the near future.
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Affiliation(s)
- E Fitzpatrick
- Paediatric Liver Centre, King's College London School of Medicine at King's College Hospital, UK
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48
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Abstract
The interest in hepatocyte transplantation has been growing continuously in recent years and this treatment may represent an alternative clinical approach for patients with acute liver failure and liver-based metabolic disorders. This chapter presents an overview of liver cell transplantation, from the basic research to human experience. It summarizes the pre-clinical studies and present status of clinical hepatocyte transplantation and identifies some possible areas of future research in this area.
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Affiliation(s)
- Juliana Puppi
- Institute of Liver Studies, King's College London School of Medicine London, UK
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Hung KC, Yong CC, Chen YS, Eng HL, Kuo FY, Lin CC, Young TH, Kobayashi E, Chen CL, Wang CC. A surgical model of fulminant hepatic failure in rabbits. Liver Int 2007; 27:1333-41. [PMID: 18036098 DOI: 10.1111/j.1478-3231.2007.01512.x] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
AIM Animal models of fulminant hepatic failure (FHF) have been developed for characterization of disease progression and to evaluate the effectiveness of liver-assist devices, some by treatment with hepatotoxic drugs, viral hepatitis or surgical procedures. We have developed a model in the rabbit by combining resection of the three anterior lobes with ligation of the pedicle of the right lateral lobes, resulting in liver necrosis; the remnant quadrate lobes are left intact. MATERIALS AND METHODS Adult male New Zealand white rabbits (n=16) were used. Six animals were killed to measure the weight of the separate liver lobes. The others (n=10) underwent left neck central line placement to monitor continuous blood pressure and collect blood for laboratory analysis, and a burr hole on the right parietal bone to monitor the intracranial pressure (ICP). Blood laboratory analysis, clinical hepatic encephalopathy and ICP levels were measured in FHF animals (n=6). Animals (n=4) undergoing a sham operation served as controls. RESULTS All FHF animals died between 12 and 26 h after liver surgery from FHF characterized by a progressive increase in liver enzymes, ammonia, total bilirubin, coagulopathy, hepatic encephalopathy and intracranial hypertension. Histological features of the ischaemic lobes showed coagulative necrosis of hepatocytes with absence of nuclei and collapse of cell plates. Brain histology revealed hypoxic cell damage. CONCLUSION We have developed a simple, reproducible model of FHF in rabbits that has a number of features comparable with clinical FHF patients and is well suited for testing experimental bioartificial liver systems and investigating the pathogenesis of FHF.
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Affiliation(s)
- Kuo-Chen Hung
- Department of Surgery, Chang Gung Memorial Hospital, Kaohsiung Medical Center, Kaohsiung Hsien, Chang Gung University College of Medicine, Taiwan
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Miyazaki M, Hardjo M, Masaka T, Tomiyama K, Mahmut N, Medina RJ, Niida A, Sonegawa H, Du G, Yong R, Takaishi M, Sakaguchi M, Huh NH. Isolation of a bone marrow-derived stem cell line with high proliferation potential and its application for preventing acute fatal liver failure. Stem Cells 2007; 25:2855-63. [PMID: 17702985 DOI: 10.1634/stemcells.2007-0078] [Citation(s) in RCA: 34] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022]
Abstract
Transplantation of hepatocytes or hepatocyte-like cells of extrahepatic origin is a promising strategy for treatment of acute and chronic liver failure. We examined possible utility of hepatocyte-like cells induced from bone marrow cells for such a purpose. Clonal cell lines were established from the bone marrow of two different rat strains. One of these cell lines, rBM25/S3 cells, grew rapidly (doubling time, approximately 24 hours) without any appreciable changes in cell properties for at least 300 population doubling levels over a period of 300 days, keeping normal diploid karyotype. The cells expressed CD29, CD44, CD49b, CD90, vimentin, and fibronectin but not CD45, indicating that they are of mesenchymal cell origin. When plated on Matrigel with hepatocyte growth factor and fibroblast growth factor-4, the cells efficiently differentiated into hepatocyte-like cells that expressed albumin, cytochrome P450 (CYP) 1A1, CYP1A2, glucose 6-phosphatase, tryptophane-2,3-dioxygenase, tyrosine aminotransferase, hepatocyte nuclear factor (HNF)1 alpha, and HNF4alpha. Intrasplenic transplantation of the differentiated cells prevented fatal liver failure in 90%-hepatectomized rats. In conclusion, a clonal stem cell line derived from adult rat bone marrow could differentiate into hepatocyte-like cells, and transplantation of the differentiated cells could prevent fatal liver failure in 90%-hepatectomized rats. The present results indicate a promising strategy for treating human fatal liver diseases.
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Affiliation(s)
- Masahiro Miyazaki
- Department of Cell Biology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Schikata-cho, Okayama 700-8558, Japan
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