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1
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Li T, Hu Z, Wang L, Lv GY. Details determining the success in establishing a mouse orthotopic liver transplantation model. World J Gastroenterol 2020; 26:3889-3898. [PMID: 32774064 PMCID: PMC7385559 DOI: 10.3748/wjg.v26.i27.3889] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/17/2020] [Revised: 05/03/2020] [Accepted: 06/16/2020] [Indexed: 02/06/2023] Open
Abstract
Liver transplantation (LT) is currently the only effective treatment option for end-stage liver disease. The importance of animal models in transplantation is widely recognized among researchers. Because of the well-characterized mouse genome and the greater diversity and availability of both genetically modified animals and research reagents, mouse orthotopic LT (MOLT) has become an ideal model for the investigation of liver biology, tissue injury, regulation of alloimmunity and tolerance induction, and the pathogenesis of specific liver diseases. However, due to its complicated and technically demanding procedure, the model has merely been used by only a few research groups in the world for years. For a new learner, training lasting at least a couple of months or even years is required. Most of the investigators have emphasized the importance of elaborate techniques and dedicated instruments in establishing a MOLT model, but some details are often neglected. The nontechnical details are also significant, especially for researchers who have little experience in mouse microsurgery. Here, we review and summarize the crucial technical and nontechnical details in establishing the model of MOLT based on scientific articles and our experience in six aspects: animal selection, anesthesia, perioperative management, organ procurement, back-table preparation, and implantation surgery. We aim to enable research groups to shorten the learning curve and implement the mouse LT procedure with high technical success.
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Affiliation(s)
- Ting Li
- Department of Hepatobiliary and Pancreatic Surgery, The First Hospital of Jilin University, Changchun 130021, Jilin Province, China
| | - Zheng Hu
- Key Laboratory of Organ Regeneration and Transplantation of Ministry of Education, The First Hospital of Jilin University, Changchun 130061, Jilin Province, China
- National-Local Joint Engineering Laboratory of Animal Models for Human Diseases, Changchun 130061, Jilin Province, China
| | - Lei Wang
- Department of Gastrointestinal Surgery, The First Hospital of Jilin University, Changchun 130021, Jilin Province, China
| | - Guo-Yue Lv
- Department of Hepatobiliary and Pancreatic Surgery, The First Hospital of Jilin University, Changchun 130021, Jilin Province, China
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2
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Van Hul N, Lendahl U, Andersson ER. Mouse Models for Diseases in the Cholangiocyte Lineage. Methods Mol Biol 2019; 1981:203-236. [PMID: 31016657 DOI: 10.1007/978-1-4939-9420-5_14] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Cholangiopathies are an important group of liver diseases affecting the biliary system, and the purpose of this review is to describe how diseases in the biliary system can be studied in mouse models. A particular focus is placed on mouse models for Alagille syndrome, a cholangiopathy with a strong genetic link to dysfunctional Notch signaling. Recently, a number of different genetic mouse models based on various manipulations of the Notch signaling pathway have been generated to study Alagille syndrome, and we discuss the resulting phenotypes, and possible causes for the phenotypic heterogeneity among the various models. In the final section, we provide a more general discussion on how well mouse models can be expected to mimic human liver disease, as well as an outlook toward the need for new technologies that can help us to gain new insights from mouse models for liver disease.
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Affiliation(s)
- Noémi Van Hul
- Department of Biosciences and Nutrition, Karolinska Institutet, Huddinge, Sweden
| | - Urban Lendahl
- Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden.
| | - Emma R Andersson
- Department of Biosciences and Nutrition, Karolinska Institutet, Huddinge, Sweden.,Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden
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3
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Zeng X, Wang S, Li S, Yang Y, Fang Z, Huang H, Wang Y, Fan X, Ye Q. Hypothermic oxygenated machine perfusion alleviates liver injury in donation after circulatory death through activating autophagy in mice. Artif Organs 2019; 43:E320-E332. [PMID: 31237688 DOI: 10.1111/aor.13525] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2019] [Revised: 06/12/2019] [Accepted: 06/13/2019] [Indexed: 12/13/2022]
Abstract
Hypothermic oxygenated machine perfusion (HOPE) is a safe and reliable method that could alleviate liver injury in donation after circulatory death (DCD). This study focuses on the role of autophagy in HOPE's protective effect on DCD liver injury. A 30-minute warm ischemic liver model was established in mice. After 4 hours of cold storage (CS), 1 hour of hypothermic machine perfusion (HMP) with 100% O2 or 100% N2 was employed. During 2 hours of reperfusion, liver tissue and perfusate were collected to evaluate liver function, oxidative stress level, apoptosis, and necrosis. Western blotting was used to explore the level of autophagy. When the liver experienced warm ischemic injury, LC3B-II expression was significantly enhanced. Compared with the CS, HOPE induced lower release of AST and ALT, as well as lower oxidative stress levels, apoptosis, and necrosis cell numbers, and led to higher tissue ATP content. Meanwhile, expression of autophagy-related proteins, such as ULK1, Atg5, and LC3B-II, increased. When oxygen was completely replaced by nitrogen, the washout effect of HMP did not activate autophagy and did not relieve DCD liver injury. When the autophagy inhibitor 3-methyladenine was used in HOPE, the protective effect of HOPE was attenuated. In conclusion, DCD liver injury activated autophagy compared with healthy liver, while HOPE alleviated DCD liver injury by increasing autophagy levels further in this mouse model. However, HMP with 100% of N2 had no beneficial effect on DCD liver injury or on autophagy levels compared with CS. The research on autophagy may provide a new strategy for alleviating DCD liver injury in clinical practice.
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Affiliation(s)
- Xianpeng Zeng
- Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Wuhan, China
| | - Shengjie Wang
- Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Wuhan, China
| | - Shiyi Li
- Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Wuhan, China
| | - Yunying Yang
- Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Wuhan, China
| | - Zehong Fang
- The Third General Surgery Department of Jiangxi Provincial People's Hospital, Organ Transplant Department of Jiangxi Provincial People's Hospital, Jiangxi Provincial People's Hospital, Nanchang, China
| | - Honglei Huang
- Nuffield Department of Surgical Sciences, Oxford University, Oxford, United Kingdom
| | - Yanfeng Wang
- Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Wuhan, China
| | - Xiaoli Fan
- Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Wuhan, China
| | - Qifa Ye
- Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Wuhan, China.,Research Center of National Health Ministry on Transplantation Medicine Engineering and Technology, The 3rd Xiangya Hospital of Central South University, Changsha, China
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Costello R, Kissenpfennig A, Martins PN, McDaid J. Development of transplant immunosuppressive agents - considerations in the use of animal models. Expert Opin Drug Discov 2018; 13:1041-1053. [PMID: 30332905 DOI: 10.1080/17460441.2018.1535589] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/28/2022]
Abstract
INTRODUCTION The development of all immunosuppressant agents to date has involved the experimental use of large and small animal models. Over the last half-century, immunosuppressive drugs have extended the lives of transplant patients worldwide. However, the use of animal models in the development of these drugs is not perfect, and this has brought to light a number of issues including idiosyncratic reactions that are found in animal models but not in humans. The 2006 highly publicized case of the 'elephant man' TGN 1412 drug trial highlights the importance of being cogent of the limitations of animal models. Areas covered: This review covers the utility and limitations of the use of animal models for the development of immunosuppressant agents. This includes both large and small animal models, particularly rodent models in the transplant setting. Expert opinion: The use of animal models represents a critical stage in the development of immunosuppressive drugs. Limitations include physiological differences to humans; this is especially true of immunologically naïve lab rodents with small memory cell populations. Toxic drug levels may differ widely between species. Animal models are also costly and raise ethical concerns. However, there is currently no way to recreate the complex environment of the human immune system purely in vitro.
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Affiliation(s)
- Russell Costello
- a Wellcome Wolfson Institute for Experimental Medicine , Queen's University , Belfast , UK
| | - Adrien Kissenpfennig
- a Wellcome Wolfson Institute for Experimental Medicine , Queen's University , Belfast , UK
| | - Paulo N Martins
- b Department of Surgery, Division of Transplantation, UMass Memorial Medical Center , University of Massachusetts , Worchester , MA , USA
| | - James McDaid
- c Department of Transplant Surgery , City Hospital , Belfast , UK
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Liu Y, Zhang W, Cheng Y, Miao C, Gong J, Wang M. Activation of PPARγ by Curcumin protects mice from ischemia/reperfusion injury induced by orthotopic liver transplantation via modulating polarization of Kupffer cells. Int Immunopharmacol 2018; 62:270-276. [PMID: 30036770 DOI: 10.1016/j.intimp.2018.07.013] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2018] [Revised: 06/18/2018] [Accepted: 07/13/2018] [Indexed: 02/07/2023]
Abstract
Curcumin shows protective effects on various diseases due to its anti-inflammatory and anti-oxidative functions; however, its effect on organ transplantation has not been fully elucidated. To understand its role in liver ischemia/reperfusion (I/R) injury, we studied its impact on orthotopic liver transplantation (OLT) and Kupffer cells (KCs) polarization and its underlying mechanisms. We first investigated the reactive oxygen species (ROS) accumulation and cytokines profile of KCs, intracellular ROS and the mRNA level of pro-inflammatory cytokines were downregulated while the mRNA level of anti-inflammatory cytokine was upregulated by the pretreatment of Curcumin; Then the liver injury was detected by histopathological examination and liver function. Pretreatment with Curcumin significantly alleviated liver injury while improving liver function and overall post-transplantation survival compared with the control groups. The Western blotting showed that Curcumin inhibited the function of KCs via down-regulating the nuclear factor κb (NF-κb) signaling pathway by activating peroxisome proliferator-activated receptor γ (PPARγ) and flow cytometry revealed that Curcumin suppressed pro-inflammatory phenotype (M1) of KCs while promoting its anti-inflammatory phenotype (M2) polarization. These results showed that Curcumin may exert positive effects on I/R injury after OLT through activating PPARγ by inhibiting the activation of NF-κb pathway and remodeling the polarization of KCs. This may reveal a potential therapy for I/R injury after liver transplantation.
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Affiliation(s)
- Yan Liu
- Department of Gastroenterology, The Fifth People's Hospital of Chengdu, Chengdu, 611130, PR China
| | - Wenfeng Zhang
- Department of Hepatobiliary Surgery and Chongqing Key Laboratory of Hepatobiliary Surgery, Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, PR China
| | - Yao Cheng
- Department of Hepatobiliary Surgery and Chongqing Key Laboratory of Hepatobiliary Surgery, Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, PR China
| | - Chunmu Miao
- Department of Hepatobiliary Surgery and Chongqing Key Laboratory of Hepatobiliary Surgery, Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, PR China
| | - Jianping Gong
- Department of Hepatobiliary Surgery and Chongqing Key Laboratory of Hepatobiliary Surgery, Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, PR China
| | - Menghao Wang
- Department of Hepatobiliary Surgery and Chongqing Key Laboratory of Hepatobiliary Surgery, Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, PR China.
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6
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Oldani G, Lacotte S, Orci LA, Delaune V, Slits F, Gex Q, Morel P, Rubbia-Brandt L, Toso C. Efficient nonarterialized mouse liver transplantation using 3-dimensional-printed instruments. Liver Transpl 2016; 22:1688-1696. [PMID: 27616447 DOI: 10.1002/lt.24637] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/10/2016] [Accepted: 07/27/2016] [Indexed: 01/13/2023]
Abstract
Because of the wide availability of genetically modified animals, mouse orthotopic liver transplantation is often preferred over rat liver transplantation. We present a simplified mouse liver transplantation technique and compare transplantation outcomes with versus without hepatic artery anastomosis. Instruments for liver implantation were designed and printed with a 3-dimensional (3D) printer. The suprahepatic vena cava anastomosis was performed with a 10-0 running suture. The vena porta and infrahepatic vena cava were joined on extraluminal cuffs, using the 3D-printed device for spatial alignment and stabilization. The hepatic artery was reconstructed in half of the recipients using intraluminal stents. Liver function tests (3, 7, and 28 days) and histology (7 and 28 days) were assessed after transplantation. We performed 22 consecutive syngeneic C57BL/6 mouse orthotopic liver transplantations. The median portal clamping time was 12.5 ± 1.5 minutes. The survival rate at 4 weeks was 100% for both arterialized and nonarterialized recipients (n = 7, 4 recipients of each group being killed for early histology at day 7). Liver function tests at 3, 7, and 28 days were similar between arterialized versus nonarterialized groups. Liver parenchyma demonstrated only irrelevant abnormalities in both groups. The proposed device allows for a shorter clamping time compared with the published literature. Using this technique, the artery does not need to be anastomosed, with no impact on graft and recipient outcomes. The device is available for 3D printing. Liver Transplantation 22 1688-1696 2016 AASLD.
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Affiliation(s)
- Graziano Oldani
- Division of Abdominal Surgery, Department of Surgery, Faculty of Medicine, University of Geneva, Geneva, Switzerland.,Hepato-Pancreato-Biliary Centre, Geneva University Hospitals, Geneva, Switzerland
| | - Stéphanie Lacotte
- Division of Abdominal Surgery, Department of Surgery, Faculty of Medicine, University of Geneva, Geneva, Switzerland
| | - Lorenzo A Orci
- Division of Abdominal Surgery, Department of Surgery, Faculty of Medicine, University of Geneva, Geneva, Switzerland.,Hepato-Pancreato-Biliary Centre, Geneva University Hospitals, Geneva, Switzerland
| | - Vaihere Delaune
- Division of Abdominal Surgery, Department of Surgery, Faculty of Medicine, University of Geneva, Geneva, Switzerland.,Hepato-Pancreato-Biliary Centre, Geneva University Hospitals, Geneva, Switzerland
| | - Florence Slits
- Division of Abdominal Surgery, Department of Surgery, Faculty of Medicine, University of Geneva, Geneva, Switzerland
| | - Quentin Gex
- Division of Abdominal Surgery, Department of Surgery, Faculty of Medicine, University of Geneva, Geneva, Switzerland
| | - Philippe Morel
- Division of Abdominal Surgery, Department of Surgery, Faculty of Medicine, University of Geneva, Geneva, Switzerland.,Hepato-Pancreato-Biliary Centre, Geneva University Hospitals, Geneva, Switzerland
| | - Laura Rubbia-Brandt
- Hepato-Pancreato-Biliary Centre, Geneva University Hospitals, Geneva, Switzerland.,Division of Clinical Pathology, Department of Pathology and Immunology, Geneva University Hospitals and Faculty of Medicine, University of Geneva, Geneva, Switzerland
| | - Christian Toso
- Division of Abdominal Surgery, Department of Surgery, Faculty of Medicine, University of Geneva, Geneva, Switzerland.,Hepato-Pancreato-Biliary Centre, Geneva University Hospitals, Geneva, Switzerland
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7
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Li DY, Shi XJ, Li W, Du XH, Wang GY. Key Points in Establishing a Model of Mouse Liver Transplantation. Transplant Proc 2016; 47:2683-9. [PMID: 26680072 DOI: 10.1016/j.transproceed.2015.07.031] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2015] [Accepted: 07/14/2015] [Indexed: 12/14/2022]
Abstract
The explosion of interest in research into the mouse genome and immune system has meant that the mouse orthotopic liver transplantation (MOLT) model has become a popular means of studying transplantation immunity, organ preservation, ischemia-reperfusion injury, and surgical techniques, among others. Although numerous modifications and refinements of surgical techniques have simplified the operation, the relatively short duration of postoperative survival after MOLT remains an obstacle to longer-term follow-up studies. Here, we summarize the scientific basis of MOLT and our experience improving and refining the model in six key areas: anesthesia, operative technique, perfusion and preservation of the liver, cuff technique, anhepatic time, and the value of rearterialization for the liver graft. We also compare the characteristics of different surgical techniques, and give recommendations for the best means of tailoring technique to the objectives of a study. In doing so, we aim to assist other investigators in establishing and perfecting the MOLT model in their routine research practice.
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Affiliation(s)
- D-Y Li
- Department of Hepatobiliary & Pancreatic Surgery, the First Norman Bethune Hospital Affiliated to Jilin University, Jilin Province, China
| | - X-J Shi
- Department of Hepatobiliary & Pancreatic Surgery, the First Norman Bethune Hospital Affiliated to Jilin University, Jilin Province, China
| | - W Li
- Department of Hepatobiliary & Pancreatic Surgery, Third Hospital (China-Japan Union Hospital) of Jilin University, Jilin Province, China
| | - X-H Du
- Department of Hepatobiliary & Pancreatic Surgery, the First Norman Bethune Hospital Affiliated to Jilin University, Jilin Province, China
| | - G-Y Wang
- Department of Hepatobiliary & Pancreatic Surgery, the First Norman Bethune Hospital Affiliated to Jilin University, Jilin Province, China.
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8
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Orthotopic mouse liver transplantation to study liver biology and allograft tolerance. Nat Protoc 2016; 11:1163-74. [PMID: 27254462 DOI: 10.1038/nprot.2016.073] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/29/2022]
Abstract
Orthotopic liver transplantation in the mouse is a powerful research tool that has led to important mechanistic insights into the regulation of hepatic injury, liver immunopathology, and transplant tolerance. However, it is a technically demanding surgical procedure. Setup of the orthotopic liver transplantation model comprises three main stages: surgery on the donor mouse; back-table preparation of the liver graft; and transplant of the liver into the recipient mouse. In this protocol, we describe our procedure in stepwise detail to allow efficient completion of both the donor and recipient operations. The protocol can result in consistently high technical success rates when performed by personnel experienced in the protocol. The technique can be completed in ∼2-3 h when performed by an individual who is well practiced in performing mouse transplantation in accordance with this protocol. We have achieved a perioperative survival rate close to 100%.
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Tian Y, Lesurtel M, Ungethuem U, Song Z, Maurizio E, Clavien PA. A novel technique in mouse liver transplantation. Transpl Int 2016; 29:742-3. [PMID: 27037719 DOI: 10.1111/tri.12780] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Affiliation(s)
- Yinghua Tian
- Department of Surgery and Transplantation, University Hospital of Zurich, Zurich, Switzerland.
| | - Mickael Lesurtel
- Department of Surgery and Transplantation, University Hospital of Zurich, Zurich, Switzerland
| | - Udo Ungethuem
- Department of Surgery and Transplantation, University Hospital of Zurich, Zurich, Switzerland
| | - Zhuolun Song
- Department of Surgery and Transplantation, University Hospital of Zurich, Zurich, Switzerland
| | - Eleonora Maurizio
- Department of Surgery and Transplantation, University Hospital of Zurich, Zurich, Switzerland
| | - Pierre-Alain Clavien
- Department of Surgery and Transplantation, University Hospital of Zurich, Zurich, Switzerland
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10
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Pan N, Liu Z, He J, Li S, Lv X, Wang L, Liu Q. Comparison of Methods for the Reconstruction of the Hepatic Artery in Mouse Orthotopic Liver Transplantation. PLoS One 2015. [PMID: 26207367 PMCID: PMC4514862 DOI: 10.1371/journal.pone.0133030] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022] Open
Abstract
Background The mouse model of arterialized orthotopic liver transplantation (AOLT) has played an important role in biomedical research. The available methods of sutured anastomosis for reconstruction of the hepatic artery are complicated, resulting in a high incidence of complications and failure. Therefore, we developed and evaluated a new model of AOLT in mice. Materials and methods Male inbred C57BL/6 mice were used in this study. A continuous suture approach was applied to connect the suprahepatic inferior vena cava (SHVC). The portal vein and infrahepatic inferior vena cava (IHVC) were connected according to the "two-cuff" method. The common bile duct was connected by a biliary stent. We used the stent (G3 group) or aortic trunk (G2 group) to reconstruct the hepatic artery. The patency of the hepatic artery was verified by transecting the artery near the graft after one week. The survival rate of the recipients and serum alanine aminotransferase (ALT) levels, hepatic pathologic alterations, apoptosis and necrosis were observed at one week postoperatively. Results The patency of the hepatic artery was verified in eight of ten mice in G3 and in six of ten mice in G2. The 7-day survival rate, extents of necrosis and apoptosis, and TGF-β levels were not significantly different among the three groups (P>0.05). However, the serum ALT levels and operation time were markedly lower in G3 compared with G2 or G1 (both P<0.05). Conclusions Reconstruction of the hepatic artery using a stent can be performed quickly with a high rate of patency. This model simplifies hepatic artery anastomosis and should be promoted in the field of biomedical research.
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Affiliation(s)
- Ning Pan
- Department of Anesthesiology, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
| | - Zhenzhen Liu
- Department of General Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
- Dalian Medical University, Dalian, Liaoning Province, China
| | - Jinjing He
- Department of General Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
- Dalian Medical University, Dalian, Liaoning Province, China
| | - Song Li
- Department of General Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
- Dalian Medical University, Dalian, Liaoning Province, China
| | - Xiangwei Lv
- Department of General Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
- Dalian Medical University, Dalian, Liaoning Province, China
| | - Liming Wang
- Department of General Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
| | - Qinlong Liu
- Department of General Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
- * E-mail:
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11
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Chen J, Gong W, Ge F, Huang T, Wu D, Liang T. A review of various techniques of mouse liver transplantation. Transplant Proc 2014; 45:2517-21. [PMID: 23953573 DOI: 10.1016/j.transproceed.2013.03.035] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2012] [Revised: 12/10/2012] [Accepted: 03/06/2013] [Indexed: 12/15/2022]
Abstract
OBJECTIVES Liver transplantation in a mouse model is a valuable tool for studying transplantation immunobiology and clinic-relevant issues. However, the successful establishment is highly technical and demanding, impeding its widespread use. Herein, the aims of this study were to review and analyze the various techniques of liver transplantation in mice to circumvent pitfalls and minimize the incidence of complications. MATERIALS AND METHODS A search of PubMed was made by using the key words "mouse liver transplantation" for articles published between January 1973 and July 2012. Of the 473 publications identified, 14 were shown to be closely associated with mouse liver transplantation and 4 articles discussed specific microsurgical techniques. Through reviewing these articles, a series of potential factors were collected and analyzed in combination with other murine transplantation models, which might influence successfully establishing a mouse model for liver transplantation. RESULTS A mouse liver transplantation model is feasible and practical for experimental studies. Mouse strain, type of anesthesia, type of perfusion and storage solution, and reconstruction of bile duct are relevant factors but not determinants for a successful transplantation. Cold and warm ischemia time should be less than 4.0 hours and 20 minutes, respectively. CONCLUSIONS The cuff preparation, reconstruction of the hepatic artery, and length of the anhepatic phase play critical roles in successfully establishing a liver transplantation model in mice.
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Affiliation(s)
- J Chen
- Department of Surgery, Transplant International Research Centre, Second Affiliated Hospital of School of Medicine, Zhejiang University, Hangzhou City, People's Republic of China
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12
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Liu H, Guan L, Li Z, Wang Z, Li W. Mechanisms of murine spontaneous liver transplant tolerance. EXP CLIN TRANSPLANT 2014; 12:1-8. [PMID: 24471716 DOI: 10.6002/ect.2013.0154] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/05/2022]
Abstract
Liver transplant is associated with the induction of peripheral immune tolerance. Liver allografts are accepted spontaneously in most combinations of mismatch in major histocompatibility complex, without any requirements for immunosuppression. Liver nonparenchymal cells (especially dendritic cells and Kupffer cells), costimulatory pathways, and activated T-cell apoptosis may contribute to the induction of liver tolerance. Therefore, liver tolerance is an active process that includes T-cell activation, proliferation, infiltration of the allograft, and T-cell apoptosis. Liver dendritic cells may modulate the amount of alloreactive T cells in liver graft recipients by expressing the coinhibitory molecule programmed death-ligand 1 and the immunosuppressive enzyme indoleamine 2,3-dioxygenase. Liver dendritic cells also may induce activated T-cell apoptosis and Foxp3+ regulatory T cells. Future studies may clarify the precise function of liver nonparenchymal cells, the interactions between programmed death-ligand 1 and other costimulatory signals, and the contribution of the liver microenvironment to the induction and expansion of Foxp 3 regulatory T cells.
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Affiliation(s)
- Hongyu Liu
- Department of Hepatobiliary-Pancreatic Surgery, The Third Hospital of Jilin University (China-Japan Union Hospital), Changchun, China
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13
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Zhou S, Palanisamy AP, McGillicuddy JW, Theruvath TP, Emre SH, Chavin KD. New method of stent-facilitated arterial reconstruction for orthotopic mouse liver transplantation. J Surg Res 2013; 187:297-301. [PMID: 24252852 DOI: 10.1016/j.jss.2013.10.024] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2013] [Revised: 09/26/2013] [Accepted: 10/15/2013] [Indexed: 12/15/2022]
Abstract
BACKGROUND Arterialized orthotopic liver transplantation (OLT) in the mouse mimics human liver transplantation physiologically and clinically. The present method of sutured anastomosis for reconstruction of the hepatic artery is complex and is associated with high incidence of complications and failure. This makes the endpoint assessment of using this complex model difficult because of the many variables of the technical aspect. METHODS A total of 14 pairs of donors and recipients from syngeneic male mice were used for arterialized OLT. The grafts were stored in University of Wisconsin solution at 4°C for less than 4 h, and the recipients underwent OLT using a two-cuff technique. The arterial reconstruction was facilitated by the use of a single stent connecting the donor liver artery segment to the recipient common hepatic artery. RESULTS All 14 recipients survived with the time for arterial reconstruction ranging from 4-10 min. Patency of the artery was confirmed by transecting the artery near the graft 2 and 14 d after transplantation. At day 2, five of the six arteries transected were patent and at day 14, seven of the remaining eight were patent for an overall patency rate of 85.7%. CONCLUSIONS The stent-facilitated arterial reconstruction can be done quickly with a high patency rate. This model expands the translational research efforts to address marginal livers such as steatotic livers.
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Affiliation(s)
- Shaotang Zhou
- Division of Transplantation Surgery, Department of Surgery, Medical University of South Carolina, Charleston, South Carolina
| | - Arun P Palanisamy
- Division of Transplantation Surgery, Department of Surgery, Medical University of South Carolina, Charleston, South Carolina
| | - John W McGillicuddy
- Division of Transplantation Surgery, Department of Surgery, Medical University of South Carolina, Charleston, South Carolina
| | - Tom P Theruvath
- Division of Transplantation Surgery, Department of Surgery, Medical University of South Carolina, Charleston, South Carolina
| | - Sukru H Emre
- Department of Surgery, Yale School of Medicine, New Haven, Connecticut
| | - Kenneth D Chavin
- Division of Transplantation Surgery, Department of Surgery, Medical University of South Carolina, Charleston, South Carolina.
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14
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Wang G, Hu B, Li Z. Cold ischemia/reperfusion injury in a mouse model of partial liver transplantation. J Surg Res 2013; 181:337-41. [DOI: 10.1016/j.jss.2012.06.068] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2012] [Revised: 05/20/2012] [Accepted: 06/26/2012] [Indexed: 10/28/2022]
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Humar B, Raptis DA, Weber A, Graf R, Clavien PA, Tian Y. Sewed revascularization for arterialized liver transplantation in mice. J Surg Res 2013; 184:e1-7. [PMID: 23587457 DOI: 10.1016/j.jss.2013.03.022] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2012] [Revised: 02/08/2013] [Accepted: 03/07/2013] [Indexed: 01/27/2023]
Abstract
BACKGROUND Mouse models of liver transplantation are powerful tools for biomedical research. The cuff method is currently the most popular approach for revascularization of mouse liver grafts, as it is relatively easy to perform hence reducing the anhepatic time. However, the use of cuffs may induce a tissue reaction, causing chronic obstruction of anastomosed vessels, leading to portal hypertension. Here, we applied the suture technique for arterialized liver transplantation in mice. MATERIALS AND METHODS Liver transplantation was performed on 14 pairs of C57BL/6 mice. All hepatic vessels were anastomosed by sewing. The bile duct was connected with a stent. The liver grafts were harvested for histology on day 30 after surgery. Serum aspartate transaminase, alkaline phosphatase and bilirubin were measured at d 3, 7, and 30 after implantation. RESULTS With a mean anhepatic time of 25.78 ± 3 min, the survival rate was 86% (n = 14) at 30 d following surgery. During this period, no significant liver injury was observed as assessed by serum markers and histology. Survival remained stable when grafts were exposed to 6 h cold ischemia prior to implantation. Vessel examination at the end of the studied period revealed an intact patency and a lack of collateral vessel growth. CONCLUSION Arterialized liver transplantation with sewed revascularization in mice is technically feasible. Both sewing and arterialization seem to be important factors promoting the survival of mouse recipients. The mouse model of suture arterialized orthotopic liver transplantation provides a novel tool for modern transplantation research and might be particularly suited for studies requiring longer-term survival of recipients.
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Affiliation(s)
- Bostjan Humar
- Laboratory of Hepatobiliary and Liver Transplantation, Department of Surgery, University Hospital of Zurich, Zurich, Switzerland
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16
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Liu Q, Rehman H, Krishnasamy Y, Haque K, Schnellmann R, Lemasters J, Zhong Z. Amphiregulin stimulates liver regeneration after small-for-size mouse liver transplantation. Am J Transplant 2012; 12:2052-61. [PMID: 22694592 PMCID: PMC3409348 DOI: 10.1111/j.1600-6143.2012.04069.x] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023]
Abstract
This study investigated whether amphiregulin (AR), a ligand of the epidermal growth factor receptor (EGFR), improves liver regeneration after small-for-size liver transplantation. Livers of male C57BL/6 mice were reduced to ~50% and ~30% of original sizes and transplanted. After transplantation, AR and AR mRNA increased in 50% but not in 30% grafts. 5-Bromodeoxyuridine (BrdU) labeling, proliferating cell nuclear antigen (PCNA) expression and mitotic index increased substantially in 50% but not 30% grafts. Hyperbilirubinemia and hypoalbuminemia occurred and survival decreased after transplantation of 30% but not 50% grafts. AR neutralizing antibody blunted regeneration in 50% grafts whereas AR injection (5 μg/mouse, iv) stimulated liver regeneration, improved liver function and increased survival after transplantation of 30% grafts. Phosphorylation of EGFR and its downstream signaling molecules Akt, mTOR, p70S6K, ERK and JNK increased markedly in 50% but not 30% grafts. AR stimulated EGFR phosphorylation and its downstream signaling pathways. EGFR inhibitor PD153035 suppressed regeneration of 50% grafts and largely abrogated stimulation of regeneration of 30% grafts by AR. AR also increased cyclin D1 and cyclin E expression in 30% grafts. Together, liver regeneration is suppressed in small-for-size grafts, as least in part, due to decreased AR formation. AR supplementation could be a promising therapy to stimulate regeneration of partial liver grafts.
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Affiliation(s)
- Q. Liu
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425,Department of General Surgery, the Second Affiliated Hospital of Dalian Medical University, Dalian, China
| | - H. Rehman
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425
| | - Y. Krishnasamy
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425
| | - K. Haque
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425
| | - R.G. Schnellmann
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425,Ralph H. Johnson VA Medical Center, Charleston, SC 29403
| | - J.J. Lemasters
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425,Department of Biochemistry & Molecular Biology, Medical University of South Carolina, Charleston, SC 29425,Hollings Cancer Center, Medical University of South Carolina, Charleston, SC 29425
| | - Z. Zhong
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425,Hollings Cancer Center, Medical University of South Carolina, Charleston, SC 29425
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17
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Ohashi N, Hori T, Chen F, Jermanus S, Eckman CB, Nakao A, Uemoto S, Nguyen JH. Matrix metalloproteinase-9 contributes to parenchymal hemorrhage and necrosis in the remnant liver after extended hepatectomy in mice. World J Gastroenterol 2012; 18:2320-2333. [PMID: 22654423 PMCID: PMC3353366 DOI: 10.3748/wjg.v18.i19.2320] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/26/2011] [Revised: 10/27/2011] [Accepted: 02/27/2012] [Indexed: 02/06/2023] Open
Abstract
AIM To investigate the effect of matrix metalloproteinase-9 (MMP-9) on the remnant liver after massive hepatectomy in the mouse. METHODS Age-matched, C57BL/6 wild-type (WT), MMP-9(-/-), and tissue inhibitors of metalloproteinases (TIMP)-1(-/-) mice were used. The mice received 80%-partial hepatectomy (PH). Samples were obtained at 6 h after 80%-PH, and we used histology, immunohistochemical staining, western blotting analysis and zymography to investigate the effect of PH on MMP-9. The role of MMP-9 after PH was investigated using a monoclonal antibody and MMP inhibitor. RESULTS We examined the remnant liver 6 h after 80%-PH and found that MMP-9 deficiency attenuated the formation of hemorrhage and necrosis. There were significantly fewer and smaller hemorrhagic and necrotic lesions in MMP-9(-/-) remnant livers compared with WT and TIMP-1(-/-) livers (P < 0.01), with no difference between WT and TIMP-1(-/-) mice. Serum alanine aminotransaminase levels were significantly lower in MMP-9(-/-) mice compared with those in TIMP-1(-/-) mice (WT: 476 ± 83 IU/L, MMP-9(-/-): 392 ± 30 IU/L, TIMP-1(-/-): 673 ± 73 IU/L, P < 0.01). Western blotting and gelatin zymography demonstrated a lack of MMP-9 expression and activity in MMP-9(-/-) mice, which was in contrast to WT and TIMP-1(-/-) mice. No change in MMP-2 expression was observed in any of the study groups. Similar to MMP-9(-/-) mice, when WT mice were treated with MMP-9 monoclonal antibody or the synthetic inhibitor GM6001, hemorrhagic and necrotic lesions were significantly smaller and fewer than in control mice (P < 0.05). These results suggest that MMP-9 plays an important role in the development of parenchymal hemorrhage and necrosis in the small remnant liver. CONCLUSION Successful MMP-9 inhibition attenuates the formation of hemorrhage and necrosis and might be a potential therapy to ameliorate liver injury after massive hepatectomy.
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18
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Clavien PA, Oberkofler CE, Raptis DA, Lehmann K, Rickenbacher A, El-Badry AM. What is critical for liver surgery and partial liver transplantation: size or quality? Hepatology 2010; 52:715-29. [PMID: 20683967 DOI: 10.1002/hep.23713] [Citation(s) in RCA: 85] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Affiliation(s)
- Pierre-Alain Clavien
- Swiss Hepato-Pancreatico-Biliary and Transplantation Center, Department of Surgery, University of Zurich, Zurich, Switzerland.
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19
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Tian Y, Chen J, Gaspert A, Segerer S, Clavien PA, Wüthrich RP, Fehr T. Kidney transplantation in mice using left and right kidney grafts. J Surg Res 2010; 163:e91-7. [PMID: 20691991 DOI: 10.1016/j.jss.2010.04.019] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2009] [Revised: 03/31/2010] [Accepted: 04/13/2010] [Indexed: 10/19/2022]
Abstract
BACKGROUND Mouse kidney transplantation is a powerful tool for scientific research. The conventional method uses only the left donor kidney for grafting because of shorter renal vessels on the right side. MATERIALS AND METHODS We developed a new technique of harvesting both left and right kidneys from one donor mouse, and separately transplanted them into two recipients. Forty-six kidney grafts (23 left, 23 right kidneys) were transplanted to 46 recipient mice. Life-supporting kidney transplantation (in which both recipient kidneys were removed) was performed in 12 recipients (six of each group). RESULTS Cold ischemia times were considerably longer for the second kidney graft (2.5-3 versus 1 h), which resulted in reduced graft function at early time points. However, the 14 d survival rate was comparable with 80% for right and 70% for left kidney grafts. Recipient animals were sacrificed between 1 and 6 wk after transplantation. Histologic examination of surviving grafts showed intact renal parenchyma, whereas total necrosis was usually seen in failed grafts. The causes of graft failure were thrombosis of the renal artery, narrow outflow of the renal vein, and fistula of the ureter. In a subgroup of animals, specific staining for apoptosis was performed. A tendency for a higher rate of apoptosis was seen at 1 wk compared with 6 wk post-transplant, but no correlation with cold ischemia time was found. CONCLUSION We report a new microsurgical technique of mouse kidney transplantation using both right and left donor kidneys as grafts for two recipient mice. Right kidney grafts showed equal survival compared with left kidney grafts. Thus, this technique reduces overall operating time and costs for microsurgery experiments.
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Affiliation(s)
- Yinghua Tian
- Division of Visceral and Transplant Surgery, University Hospital Zürich, Zürich, Switzerland.
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20
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Bolinger B, Krebs P, Tian Y, Engeler D, Scandella E, Miller S, Palmer DC, Restifo NP, Clavien PA, Ludewig B. Immunologic ignorance of vascular endothelial cells expressing minor histocompatibility antigen. Blood 2008; 111:4588-95. [PMID: 18195091 PMCID: PMC3403502 DOI: 10.1182/blood-2007-09-114769] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/24/2007] [Accepted: 01/05/2008] [Indexed: 12/20/2022] Open
Abstract
Endothelial cells (ECs) presenting minor histocompatibility antigen (mhAg) are major target cells for alloreactive effector CD8(+) T cells during chronic transplant rejection and graft-versus-host disease (GVHD). The contribution of ECs to T-cell activation, however, is still a controversial issue. In this study, we have assessed the antigen-presenting capacity of ECs in vivo using a transgenic mouse model with beta-galactosidase (beta-gal) expression confined to the vascular endothelium (Tie2-LacZ mice). In a GVHD-like setting with adoptive transfer of beta-gal-specific T-cell receptor-transgenic T cells, beta-gal expression by ECs was not sufficient to either activate or tolerize CD8(+) T cells. Likewise, transplantation of fully vascularized heart or liver grafts from Tie2-LacZ mice into nontransgenic recipients did not suffice to activate beta-gal-specific CD8(+) T cells, indicating that CD8(+) T-cell responses against mhAg cannot be initiated by ECs. Moreover, we could show that spontaneous activation of beta-gal-specific CD8(+) T cells in Tie2-LacZ mice was exclusively dependent on CD11c(+) dendritic cells (DCs), demonstrating that mhAgs presented by ECs remain immunologically ignored unless presentation by DCs is granted.
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21
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Hoekstra H, Tian Y, Jochum W, Stieger B, Graf R, Porte RJ, Clavien PA. Dearterialization of the liver causes intrahepatic cholestasis due to reduced bile transporter expression. Transplantation 2008; 85:1159-66. [PMID: 18431237 DOI: 10.1097/tp.0b013e31816b2465] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
BACKGROUND Bile duct injury after hepatic artery thrombosis (HAT) in liver transplantation is believed to be caused by ischemia predominantly. We aimed to define the involvement of bile secretory dysfunction in the pathogenesis of liver injury after HAT. METHODS In a murine model, the main hepatic artery, the extrahepatic peribiliary plexus, or both arterial connections to the liver were interrupted (n=5 for each group). After 1, 14, or 28 days, hepatobiliary function was assessed by analysis of bile transporter expression, serum bile acids and bilirubin, and hepatic ATP content. In addition, cellular injury was assessed by light microscopy and biochemical markers. RESULTS There were no signs of hepatobiliary dysfunction or injury in sham-operated animals or in mice with interruption of the hepatic artery or the extrahepatic peribiliary plexus alone. However, as early as 24 hr after complete dearterialization, bile transporter expression was significantly reduced and intrahepatic cholestasis started to progress the following weeks. Histologic studies at 28 days after complete dearterialization showed severe hepatobiliary injury. CONCLUSIONS This study indicates that arterial blood supply is critical for normal bile secretion. Bile duct injury after complete arterial deprivation is preceded by a loss of bile secretory function and subsequent intrahepatic cholestasis.
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Affiliation(s)
- Harm Hoekstra
- Department of Visceral and Transplant Surgery, University Hospital Zurich, Zurich, Switzerland
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22
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Shen XD, Ke B, Zhai Y, Gao F, Tsuchihashi SI, Lassman CR, Busuttil RW, Kupiec-Weglinski JW. Absence of toll-like receptor 4 (TLR4) signaling in the donor organ reduces ischemia and reperfusion injury in a murine liver transplantation model. Liver Transpl 2007; 13:1435-43. [PMID: 17902130 DOI: 10.1002/lt.21251] [Citation(s) in RCA: 94] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
This study analyzes how toll-like receptor 4 (TLR4) signaling in the donor organ affects the ischemia and reperfusion injury (IRI) sequel following liver transplantation. Isogenic orthotopic liver transplantations (OLTs) with rearterialization were performed in groups of wild-type (WT) and TLR4 knockout (KO) mice after donor liver preservation in University of Wisconsin solution at 4 degrees C for 24 hours. Unlike WT OLTs, TLR4-deficient OLTs transplanted to either WT or TLR4 KO recipients suffered significantly less hepatocellular damage, as evidenced by serum alanine aminotransferase levels, and histological Suzuki's grading of liver IRI. Disruption of TLR4 signaling in OLTs decreased local neutrophil sequestration, CD4+ T cell infiltration, interferon (IFN)-gamma-inducible protein 10 (CXCL10) and an intercellular adhesion molecule (ICAM-1), as well as tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-2, and IFN-gamma, yet increased IL-4 and IL-10 expression. The well-functioning OLTs from TLR4 KO donors revealed attenuated activity of capase-3, and enhanced heme oygenase-1 (HO-1) expression, along with decreased levels of apoptotic endothelial cells/hepatocytes, as compared with WT OLTs with intact TLR4 signaling. Thus, the functional sentinel TLR4 complex in the donor organ plays a key role in the mechanism of hepatic IRI after OLT. Disruption of TLR4 pathway downregulated the early proinflammatory responses and ameliorated hepatic IRI. These results provide the rationale to locally modify innate TLR4 signaling in the donor organ to more efficiently control the adaptive posttransplantation IRI-dependent responses.
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Affiliation(s)
- Xiu-Da Shen
- Dumont-University of California, Los Angeles (UCLA) Transplant Center, Division of Liver and Pancreas Transplantation, Department of Surgery, David Geffen School of Medicine at University of California, Los Angeles (UCLA), Los Angeles, CA 90095, USA
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23
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Xie JF, Wang G, Debonera F, Han R, Dorf ME, Hancock W, Olthoff KM. Selective neutralization of the chemokine TCA3 reduces the increased injury of partial versus whole liver transplants induced by cold preservation. Transplantation 2007; 82:1501-9. [PMID: 17164723 DOI: 10.1097/01.tp.0000243167.11566.eb] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023]
Abstract
BACKGROUND Given the shortage of liver donors and the development of techniques for partial liver transplantation, we compared chemokine expression and inflammatory cell infiltration of partial versus whole grafts in a mouse syngeneic liver transplant model. METHODS Orthotopic liver transplantation, using whole or partial murine liver grafts, was performed following cold preservation in ViaSpan solution for periods of one to eight hours. RESULTS Partial grafts showed more severe cold ischemia/reperfusion injury and greater inflammatory cell infiltration than whole grafts, and was accompanied by the marked intrahepatic upregulation of multiple chemokines. Quantitative analysis showed that compared with expression in whole grafts harvested after the same period of cold ischemia, partial grafts had eightfold more T-cell activation gene (TCA)-3 (CCL1) chemokine messenger RNA (mRNA) expression (P<0.01) and sixfold more inducible protein (IP)-10 chemokine (CCL10) mRNA expression (P<0.01), as well as increased expression of the chemokine receptors CCR8 (receptor for TCA3) and CXCR3 (receptor for IP-10; P<0.01). Blockade of TCA3 by neutralizing monoclonal antibody significantly decreased intragraft IP-10 expression (P<0.05) but not tumor necrosis factor-alpha or interleukin-6 expression in partial grafts, and significantly decreased cold ischemia/reperfusion injury (P<0.05) and associated neutrophil and T-cell infiltration (P<0.01). CONCLUSIONS These data demonstrate that the chemokine TCA3/CCL1 is important to the pathogenesis of ischemic injury of experimental partial liver grafts, and that its therapeutic targeting within such grafts can overcome the deleterious effects of prolonged cold preservation and restore liver function to the level achieved using whole liver grafts.
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MESH Headings
- Animals
- Antibodies, Monoclonal/pharmacology
- Chemokine CCL1
- Chemokines, CC/antagonists & inhibitors
- Chemokines, CC/genetics
- Chemokines, CC/metabolism
- Cold Ischemia
- Cryopreservation
- Liver Transplantation
- Male
- Mice
- Neutrophil Infiltration/drug effects
- Organ Preservation
- RNA, Messenger/analysis
- RNA, Messenger/metabolism
- Receptors, CCR8
- Receptors, CXCR3
- Receptors, Chemokine/genetics
- Receptors, Chemokine/metabolism
- Receptors, Cytokine/genetics
- Receptors, Cytokine/metabolism
- Reperfusion Injury/metabolism
- Reperfusion Injury/pathology
- Reperfusion Injury/prevention & control
- T-Lymphocytes/drug effects
- Up-Regulation/drug effects
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Affiliation(s)
- Jin-Fu Xie
- Department of Surgery, University of Pennsylvania, Philadelphia, PA 19104, USA
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24
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Hoekstra H, Porte RJ, Tian Y, Jochum W, Stieger B, Moritz W, Slooff MJH, Graf R, Clavien PA. Bile salt toxicity aggravates cold ischemic injury of bile ducts after liver transplantation in Mdr2+/- mice. Hepatology 2006; 43:1022-31. [PMID: 16628673 DOI: 10.1002/hep.21169] [Citation(s) in RCA: 44] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Intrahepatic bile duct strictures are a serious complication after orthotopic liver transplantation (OLT). We examined the role of endogenous bile salt toxicity in the pathogenesis of bile duct injury after OLT. Livers from wild-type mice and mice heterozygous for disruption of the multidrug resistance 2 Mdr2 gene (Mdr2+/-) were transplanted into wild-type recipient mice. Mdr2+/- mice secrete only 50% of the normal amount of phospholipids into their bile, leading to an abnormally high bile salt/phospholipid ratio. In contrast to homozygous Mdr2-/- mice, the Mdr2+/- mice have normal liver histology and function under normal conditions. Two weeks after OLT, bile duct injury and cholestasis were assessed by light and electron microscopy, as well as through molecular and biochemical markers. There were no signs of bile duct injury or intrahepatic cholestasis in liver grafts from wild-type donors. Liver grafts from Mdr2+/- donors, however, had enlarged portal tracts with cellular damage, ductular proliferation, biliostasis, and a dense inflammatory infiltrate after OLT. Parallel to this observation, recipients of Mdr2+/- livers had significantly higher serum transaminases, alkaline phosphatase, total bilirubin, and bile salt levels, as compared with recipients of wild-type livers. In addition, hepatic bile transporter expression was compatible with the biochemical and histological cholestatic profile found in Mdr2+/- grafts after OLT. In conclusion, toxic bile composition, due to a high biliary bile salt/phospholipid ratio, acted synergistically with cold ischemia in the pathogenesis of bile duct injury after transplantation.
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Affiliation(s)
- Harm Hoekstra
- Swiss HPB Center, Department of Visceral and Transplant Surgery, University Hospital Zurich, Switzerland
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25
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Tian Y, Jochum W, Georgiev P, Moritz W, Graf R, Clavien PA. Kupffer cell-dependent TNF-alpha signaling mediates injury in the arterialized small-for-size liver transplantation in the mouse. Proc Natl Acad Sci U S A 2006; 103:4598-603. [PMID: 16537374 PMCID: PMC1400589 DOI: 10.1073/pnas.0600499103] [Citation(s) in RCA: 72] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
Implantation of small liver grafts causes liver injury and defective regeneration leading to graft failure. We investigated whether Kupffer cell-dependent TNF-alpha signaling contributes to this poor outcome. Partial 30% liver transplantation was performed in C57BL/6 wild-type mice (control group), and in three groups with down-regulation of the TNF-alpha pathway: (i) TNF receptor 1 knockout [TNFR-1(-/-)] mice, and mice pretreated with (ii) gadolinium chloride or (iii) pentoxifylline (PTX). Fifty-percent partial liver transplantation, a model associated with full recovery, and transplantation in IL-6 knockout [IL-6(-/-)] mice were performed in some experiments. Graft injury, regeneration, portal flow, liver microcirculation, leukocyte adhesion, and animal survival were assessed. Animal survival rates were 14% in the control group vs. 43% in the gadolinium chloride group, 57% for the TNFR-1(-/-) group, and 86% in the PTX group (P < 0.001). Markers of liver injury were reduced in all treated groups when compared with controls. Each treated group disclosed better portal flow and sinusoid perfusion, decreased leukocyte adherence, particularly in the PTX group. Liver regeneration occurred only in the treated groups. IL-6 and IL-10 levels were dramatically up-regulated (50x) in the PTX group, and at lower levels in other experimental groups. The protective effect of PTX was lost in IL-6(-/-) mice and protection was restored by a single dose of r-IL-6. In conclusion, interruption of TNF-alpha signaling or depletion of Kupffer cells improves survival after 30% liver transplantation, reduces liver injury, and enhances regeneration. The superior effects of PTX are mediated by IL-6.
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Affiliation(s)
- Yinghua Tian
- Swiss Hepato-Pancreatico Biliary Center, Departments of *Visceral and Transplantation Surgery and
| | - Wolfram Jochum
- Pathology, University Hospital of Zurich, Raemistrasse 100, CH-8091 Zurich, Switzerland
| | - Panco Georgiev
- Swiss Hepato-Pancreatico Biliary Center, Departments of *Visceral and Transplantation Surgery and
| | - Wolfgang Moritz
- Swiss Hepato-Pancreatico Biliary Center, Departments of *Visceral and Transplantation Surgery and
| | - Rolf Graf
- Swiss Hepato-Pancreatico Biliary Center, Departments of *Visceral and Transplantation Surgery and
| | - Pierre-Alain Clavien
- Swiss Hepato-Pancreatico Biliary Center, Departments of *Visceral and Transplantation Surgery and
- To whom correspondence should be addressed. E-mail:
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Dahm F, Georgiev P, Clavien PA. Small-for-size syndrome after partial liver transplantation: definition, mechanisms of disease and clinical implications. Am J Transplant 2005; 5:2605-10. [PMID: 16212618 DOI: 10.1111/j.1600-6143.2005.01081.x] [Citation(s) in RCA: 470] [Impact Index Per Article: 23.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023]
Abstract
Widespread application of cadaveric split or living donor liver transplantation bears considerable potential to increase the pool of available organs and thus alleviate the problem of organ shortage. Although splitting of a cadaveric liver into two grafts for adult recipients can be performed successfully, sufficient function of undersized grafts is a major concern. To minimize the risk for living donors, transplant surgeons aim at procuring the least necessary liver volume, also leading to potentially small grafts. When small partial grafts are unable to meet the functional demands, the recipients can develop a so-called small-for-size syndrome (SFSS). There is currently limited data on the pathogenesis of SFSS, with clinical studies mainly focusing on portal hyperperfusion. Additional aspects include graft-related factors such as functional and regenerative capacity, as well as recipient-related factors, such as overall health status and severity of cirrhosis. However, there is currently no consensus on the definition of SFSS. We propose a novel definition, based on simple clinical criteria, which divides the syndrome into either nonfunction or dysfunction of a small graft after the exclusion of other causes. This definition should ease comparability of future clinical trials, and thus improve understanding of the pathogenesis of SFSS.
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Affiliation(s)
- Felix Dahm
- Department of Visceral and Transplantation Surgery, University Hospital Zurich, Zurich, Switzerland
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27
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Shen XD, Gao F, Ke B, Zhai Y, Lassman CR, Tsuchihashi SI, Farmer DG, Busuttil RW, Kupiec-Weglinski JW. Inflammatory responses in a new mouse model of prolonged hepatic cold ischemia followed by arterialized orthotopic liver transplantation. Liver Transpl 2005; 11:1273-81. [PMID: 16184555 DOI: 10.1002/lt.20489] [Citation(s) in RCA: 37] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
The current models of liver ischemia/reperfusion injury (IRI) in mice are largely limited to a warm ischemic component. To investigate the mechanism of hepatic "cold" IRI, we developed and validated a new mouse model of prolonged cold preservation followed by syngeneic orthotopic liver transplantation (OLT). Two hundred and forty-three OLTs with or without rearterialization and preservation in University of Wisconsin solution at 4 degrees C were performed in Balb/c mice. The 14-day survivals in the nonarterialized OLT groups were 92% (11/12), 82% (9/11), and 8% (1/12) after 1-hour, 6-hour and 24-hour preservation, respectively. In contrast, hepatic artery reconstruction after 1-hour, 6-hour, and 24-hour preservation improved the outcome as evidenced by 2-week survival of 100% (12/12), 100% (10/10), and 33% (4/12), respectively, and diminished hepatocellular damage (serum alanine aminotransferase /histology). Moreover, 24-hour (but not 1-h) cold preservation of rearterialized OLTs increased hepatic CD4+ T-cell infiltration and proinflammatory cytokine (tumor necrosis factor-alpha, interleukin 2, interferon-gamma) production, as well as enhanced local apoptosis, and Toll-like receptor 4/caspase 3 expression. These cardinal features of hepatic IRI validate the model. In conclusion, we have developed and validated a new mouse model of IRI in which hepatic artery reconstruction was mandatory for long-term animal survival after prolonged (24-h) OLT preservation. With the availability of genetically manipulated mouse strains, this model should provide important insights into the mechanism of antigen-independent hepatic IRI and help design much needed refined therapeutic means to combat hepatic IRI in the clinics.
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Affiliation(s)
- Xiu-Da Shen
- Dumont-UCLA Transplant Center, Division of Liver and Pancreas Transplantation, Department of Surgery, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095, USA
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Birsner JH, Wan C, Cheng G, Evans ZP, Polito CC, Fiorini RN, Gilbert G, Haines JK, Schmidt MG, Chavin KD. Steatotic liver transplantation in the mouse: a model of primary nonfunction. J Surg Res 2004; 120:97-101. [PMID: 15172195 DOI: 10.1016/j.jss.2003.11.022] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2003] [Indexed: 12/16/2022]
Abstract
BACKGROUND The number of potential donor organs deemed suboptimal for transplantation because of hepatic steatosis is rising as the obesity rate increases. However, no mouse transplant model has been described within the framework of hepatic steatosis. We describe the development of and our initial experience with a steatotic mouse orthotopic liver transplant model using the ob/ob mouse. This model is technically achievable and functionally mimics primary nonfunction. MATERIALS AND METHODS Adapting techniques of a nonarterialized murine transplant model, C57BL6 ob/ob mice aged 5-7 weeks (26-35 g) and lean controls served as liver donors and recipients. Orthotopic liver transplantation (OLT) was performed using a two-cuff technique at the infrahepatic cava and portal vein. The suprahepatic cava was anastomosed end to end, and the bile duct was stented. The hepatic artery was not reconstructed. RESULTS Lean-to-lean OLT was performed with 70% (n = 10) long-term survival. ob/ob-to-age-matched lean recipients had 0% (n = 10) survival because of size discrepancy. ob/ob livers were transplanted to size-matched lean recipients (>3 months old) with short-term survival of 30% (n = 10). These mice survived the operation, awakened, but expired within 24 h. Serum transaminases revealed a significantly higher injury profile in the recipients of the steatotic livers, and histology showed massive centrilobular coagulative necrosis with hemorrhage, the overall picture being that of primary nonfunction. CONCLUSIONS This novel use of the ob/ob mouse for OLT provides us with a model for steatotic transplantation with primary nonfunction as the end point and may help to better understand the response of the steatotic liver to the insult of transplantation.
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Affiliation(s)
- Jennifer H Birsner
- Department of Surgery, Medical University of South Carolina, Charleston, South Carolina, USA
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Tian Y, Graf R, Clavien PA. Reconstruction of the hepatic artery in liver transplantation in mice: mandatory or useless? Transplantation 2004; 77:1310; author reply: 1310-1, 1311-2. [PMID: 15114108 DOI: 10.1097/01.tp.0000122226.92721.25] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/26/2022]
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RESPONSE TO “RECONSTRUCTION OF THE HEPATIC ARTERY IN LIVER TRANSPLANTATION IN MICE: MANDATORY OR USELESS?”. Transplantation 2004. [DOI: 10.1097/01.tp.0000122228.79061.50] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
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RESPONSE TO “RECONSTRUCTION OF THE HEPATIC ARTERY IN LIVER TRANSPLANTATION IN MICE: MANDATORY OR USELESS?”. Transplantation 2004. [DOI: 10.1097/01.tp.0000122227.10862.ab] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
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Que X, Debonera F, Xie J, Furth EE, Aldeguer X, Gelman AE, Olthoff KM. Pattern of ischemia reperfusion injury in a mouse orthotopic liver transplant model. J Surg Res 2004; 116:262-8. [PMID: 15013365 DOI: 10.1016/j.jss.2003.07.005] [Citation(s) in RCA: 32] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2003] [Indexed: 12/14/2022]
Abstract
BACKGROUND The molecular pathways of ischemic injury after liver transplantation are complex and difficult to dissect because of the presence of many variables. Transgenic and genetically deficient strains of mice provide ideal models for the study of the contribution of a single gene product in biological processes in vivo. Although well described in rats, prolonged preservation has not been studied in a mouse model of orthotopic liver transplantation (mOLT). The aim of this study was to establish a model of cold ischemia and reperfusion injury in mOLT and describe the pattern of the regenerative response to various lengths of cold storage. MATERIALS AND METHODS mOLT was performed using a syngeneic combination. Grafts were preserved at 4 degrees C in University of Wisconsin (Viaspan) solution for increasing periods of cold preservation. After cold storage, the liver grafts were transplanted and recipient survival was monitored. Hepatocellular injury was determined by histology, and the regenerative response was quantitated by interleukin 6 upregulation and DNA replication. RESULTS Long-term survival was 100%, 100%, 88%, and 0% for cold preservation of 1, 4, 8, and 16 h, respectively. Grafts with short preservation times (1 and 4 h) demonstrated limited injury and a weak regenerative response, with slight IL-6 early upregulation and minimal cell division. Eight hours of cold ischemia resulted in prominent injury and an intense regenerative response accompanied by significant IL-6 upregulation and DNA synthesis. Sixteen hours of storage resulted in all recipients succumbing to liver failure, with histology showing extensive hepatic necrosis. CONCLUSIONS This study demonstrated the feasibility of using the mOLT model for the study of molecular mechanisms associated with recovery from cold ischemia and reperfusion injury. Increasing lengths of cold ischemia correlate with progressive tissue damage whereas recovery is associated with a regenerative response that correlates with the severity of injury.
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Affiliation(s)
- Xingyi Que
- Department of Surgery, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA
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Tian Y, Graf R, Jochum W, Clavien PA. Arterialized partial orthotopic liver transplantation in the mouse: a new model and evaluation of the critical liver mass. Liver Transpl 2003; 9:789-95. [PMID: 12884190 DOI: 10.1053/jlts.2003.50170] [Citation(s) in RCA: 38] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
The availability of a model of partial orthotopic liver transplantation (OLT) in the mouse would be an important tool for studying injuries associated with transplantation. The goals of this study were three-fold: (1). to develop a model of partial OLT in the mouse, (2). to determine the minimal graft volume in this model, and (3). to define the injury associated with small volume incompatible with animal survival. Putative grafts of 30% and 50% were prepared. Their weight was 30 +/- 5% and 45 +/- 10%, respectively. Subsequently, 30% and 45% syngeneic partial liver grafts were orthotopically transplanted into C57BL/6 mice. Each recipient receiving a 45% graft survived permanently, whereas those receiving only a 30% graft volume died within 2 to 4 days of surgery. Serum transaminase levels normalized in the 45% graft group within 14 days after surgery. In this group, small foci of necrosis and mild steatosis were noted on histology at postoperative day 2, but no abnormalities were noted after 14 days and 100 days. In contrast, recipients who underwent transplantation with a 30% graft volume showed a comparable amount of necrosis and significant microvesicular steatosis in most hepatocytes 2 days after surgery. Hepatocyte proliferation was reduced in this group when compared with animals who underwent transplantation with a 45% graft volume. In conclusion, partial liver transplantation is feasible in the mouse with a critical graft volume ranging between 30% and 45%. Small liver grafts develop massive microvesicular steatosis and impaired regeneration rapidly leading to animal death.
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Affiliation(s)
- Yinghua Tian
- Division of Visceral and Transplantation Surgery, University Hospital of Zurich, Switzerland
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