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1
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Chan JCY, Chaban R, Chang SH, Angel LF, Montgomery RA, Pierson RN. Future of Lung Transplantation: Xenotransplantation and Bioengineering Lungs. Clin Chest Med 2023; 44:201-214. [PMID: 36774165 PMCID: PMC11078107 DOI: 10.1016/j.ccm.2022.11.003] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/11/2023]
Abstract
Xenotransplantation promises to alleviate the issue of donor organ shortages and to decrease waiting times for transplantation. Recent advances in genetic engineering have allowed for the creation of pigs with up to 16 genetic modifications. Several combinations of genetic modifications have been associated with extended graft survival and life-supporting function in experimental heart and kidney xenotransplants. Lung xenotransplantation carries specific challenges related to the large surface area of the lung vascular bed, its innate immune system's intrinsic hyperreactivity to perceived 'danger', and its anatomic vulnerability to airway flooding after even localized loss of alveolocapillary barrier function. This article discusses the current status of lung xenotransplantation, and challenges related to immunology, physiology, anatomy, and infection. Tissue engineering as a feasible alternative to develop a viable lung replacement solution is discussed.
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Affiliation(s)
- Justin C Y Chan
- NYU Transplant Institute, New York University, 530 1st Avenue, Suite 7R, New York, NY 10016, USA.
| | - Ryan Chaban
- Department of Surgery, Center for Transplantation Sciences, Massachusetts General Hospital and Harvard Medical School, 55 Fruit Street, Boston, MA 02114, USA; Department of Cardiovascular Surgery, University Hospital of Johannes Gutenberg University, Langenbeckstr. 1, Bau 505, 5. OG55131 Mainz, Germany
| | - Stephanie H Chang
- NYU Transplant Institute, New York University, 530 1st Avenue, Suite 7R, New York, NY 10016, USA
| | - Luis F Angel
- NYU Transplant Institute, New York University, 530 1st Avenue, Suite 7R, New York, NY 10016, USA
| | - Robert A Montgomery
- NYU Transplant Institute, New York University, 530 1st Avenue, Suite 7R, New York, NY 10016, USA
| | - Richard N Pierson
- Department of Surgery, Center for Transplantation Sciences, Massachusetts General Hospital and Harvard Medical School, 55 Fruit Street, Boston, MA 02114, USA
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2
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Denner J. What does the PERV copy number tell us? Xenotransplantation 2022; 29:e12732. [PMID: 35112403 DOI: 10.1111/xen.12732] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2021] [Revised: 01/05/2022] [Accepted: 01/12/2022] [Indexed: 11/27/2022]
Affiliation(s)
- Joachim Denner
- Institute of Virology, Free University Berlin, Berlin, Germany
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3
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High Prevalence of Recombinant Porcine Endogenous Retroviruses (PERV-A/Cs) in Minipigs: A Review on Origin and Presence. Viruses 2021; 13:v13091869. [PMID: 34578447 PMCID: PMC8473008 DOI: 10.3390/v13091869] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/15/2021] [Revised: 09/14/2021] [Accepted: 09/15/2021] [Indexed: 12/17/2022] Open
Abstract
Minipigs play an important role in biomedical research and they have also been used as donor animals for preclinical xenotransplantations. Since zoonotic microorganisms including viruses can be transmitted when pig cells, tissues or organs are transplanted, virus safety is an important feature in xenotransplantation. Whereas most porcine viruses can be eliminated from pig herds by different strategies, this is not possible for porcine endogenous retroviruses (PERVs). PERVs are integrated in the genome of pigs and some of them release infectious particles able to infect human cells. Whereas PERV-A and PERV-B are present in all pigs and can infect cells from humans and other species, PERV-C is present in most, but not all pigs and infects only pig cells. Recombinant viruses between PERV-A and PERV-C have been found in some pigs; these recombinants infect human cells and are characterized by high replication rates. PERV-A/C recombinants have been found mainly in minipigs of different origin. The possible reasons of this high prevalence of PERV-A/C in minipigs, including inbreeding and higher numbers and expression of replication-competent PERV-C in these animals, are discussed in this review. Based on these data, it is highly recommended to use only pig donors in clinical xenotransplantation that are negative for PERV-C.
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4
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Kosanke M, Osetek K, Haase A, Wiehlmann L, Davenport C, Schwarzer A, Adams F, Kleppa MJ, Schambach A, Merkert S, Wunderlich S, Menke S, Dorda M, Martin U. Reprogramming enriches for somatic cell clones with small-scale mutations in cancer-associated genes. Mol Ther 2021; 29:2535-2553. [PMID: 33831558 DOI: 10.1016/j.ymthe.2021.04.007] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/29/2020] [Revised: 03/03/2021] [Accepted: 04/02/2021] [Indexed: 02/06/2023] Open
Abstract
Cellular therapies based on induced pluripotent stem cells (iPSCs) come out of age and an increasing number of clinical trials applying iPSC-based transplants are ongoing or in preparation. Recent studies, however, demonstrated a high number of small-scale mutations in iPSCs. Although the mutational load in iPSCs seems to be largely derived from their parental cells, it is still unknown whether reprogramming may enrich for individual mutations that could lead to loss of functionality and tumor formation from iPSC derivatives. 30 hiPSC lines were analyzed by whole exome sequencing. High accuracy amplicon sequencing showed that all analyzed small-scale variants pre-existed in their parental cells and that individual mutations present in small subpopulations of parental cells become enriched among hiPSC clones during reprogramming. Among those, putatively actionable driver mutations affect genes related to cell-cycle control, cell death, and pluripotency and may confer a selective advantage during reprogramming. Finally, a short hairpin RNA (shRNA)-based experimental approach was applied to provide additional evidence for the individual impact of such genes on the reprogramming efficiency. In conclusion, we show that enriched mutations in curated onco- and tumor suppressor genes may account for an increased tumor risk and impact the clinical value of patient-derived hiPSCs.
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Affiliation(s)
- Maike Kosanke
- Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany, Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), 30625 Hannover, Germany
| | - Katarzyna Osetek
- Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany, Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), 30625 Hannover, Germany
| | - Alexandra Haase
- Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany, Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), 30625 Hannover, Germany
| | - Lutz Wiehlmann
- Research Core Unit Genomics, Hannover Medical School, 30625 Hannover, Germany
| | - Colin Davenport
- Research Core Unit Genomics, Hannover Medical School, 30625 Hannover, Germany
| | - Adrian Schwarzer
- Department of Hematology, Oncology and Stem Cell Transplantation, Institute of Experimental Hematology, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany
| | - Felix Adams
- Department of Hematology, Oncology and Stem Cell Transplantation, Institute of Experimental Hematology, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany
| | - Marc-Jens Kleppa
- Department of Hematology, Oncology and Stem Cell Transplantation, Institute of Experimental Hematology, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany
| | - Axel Schambach
- Department of Hematology, Oncology and Stem Cell Transplantation, Institute of Experimental Hematology, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany
| | - Sylvia Merkert
- Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany, Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), 30625 Hannover, Germany
| | - Stephanie Wunderlich
- Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany, Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), 30625 Hannover, Germany
| | - Sandra Menke
- Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany, Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), 30625 Hannover, Germany
| | - Marie Dorda
- Research Core Unit Genomics, Hannover Medical School, 30625 Hannover, Germany
| | - Ulrich Martin
- Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, REBIRTH - Research Center for Translational Regenerative Medicine, Hannover Medical School, 30625 Hannover, Germany, Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), 30625 Hannover, Germany.
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6
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Weiss RA. Infection hazards of xenotransplantation: Retrospect and prospect. Xenotransplantation 2018; 25:e12401. [PMID: 29756309 DOI: 10.1111/xen.12401] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/08/2018] [Accepted: 03/29/2018] [Indexed: 02/06/2023]
Affiliation(s)
- Robin A Weiss
- Division of Infection & Immunity, University College London, London, UK
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7
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Łopata K, Wojdas E, Nowak R, Łopata P, Mazurek U. Porcine Endogenous Retrovirus (PERV) - Molecular Structure and Replication Strategy in the Context of Retroviral Infection Risk of Human Cells. Front Microbiol 2018; 9:730. [PMID: 29755422 PMCID: PMC5932395 DOI: 10.3389/fmicb.2018.00730] [Citation(s) in RCA: 26] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/26/2017] [Accepted: 03/28/2018] [Indexed: 12/28/2022] Open
Abstract
The xenotransplantation of porcine tissues may help overcome the shortage of human organs for transplantation. However, there are some concerns about recipient safety because the risk of porcine endogenous retrovirus (PERV) transmission to human cells remains unknown. Although, to date, no PERV infections have been noted in vivo, the possibility of such infections has been confirmed in vitro. Better understanding of the structure and replication cycle of PERVs is a prerequisite for determining the risk of infection and planning PERV-detection strategies. This review presents the current state of knowledge about the structure and replication cycle of PERVs in the context of retroviral infection risk.
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Affiliation(s)
- Krzysztof Łopata
- Department of Molecular Biology, School of Pharmacy with the Division of Laboratory Medicine in Sosnowiec, Medical University of Silesia, Katowice, Poland
| | - Emilia Wojdas
- Department of Molecular Biology, School of Pharmacy with the Division of Laboratory Medicine in Sosnowiec, Medical University of Silesia, Katowice, Poland.,Department of Instrumental Analysis, School of Pharmacy with the Division of Laboratory Medicine in Sosnowiec, Medical University of Silesia, Katowice, Poland
| | - Roman Nowak
- Department of Molecular Biology, School of Pharmacy with the Division of Laboratory Medicine in Sosnowiec, Medical University of Silesia, Katowice, Poland
| | - Paweł Łopata
- Department of Molecular Biology, School of Pharmacy with the Division of Laboratory Medicine in Sosnowiec, Medical University of Silesia, Katowice, Poland
| | - Urszula Mazurek
- Department of Molecular Biology, School of Pharmacy with the Division of Laboratory Medicine in Sosnowiec, Medical University of Silesia, Katowice, Poland
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8
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Harris E, Liu Y, Cunniffe G, Morrissey D, Carroll S, Mulhall K, Kelly DJ. Biofabrication of soft tissue templates for engineering the bone-ligament interface. Biotechnol Bioeng 2017. [DOI: 10.1002/bit.26362] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022]
Affiliation(s)
- Ella Harris
- Trinity Centre for Bioengineering; Trinity Biomedical Sciences Institute; Trinity College Dublin Dublin Ireland
| | - Yurong Liu
- Trinity Centre for Bioengineering; Trinity Biomedical Sciences Institute; Trinity College Dublin Dublin Ireland
| | - Grainne Cunniffe
- Trinity Centre for Bioengineering; Trinity Biomedical Sciences Institute; Trinity College Dublin Dublin Ireland
- Department of Mechanical and Manufacturing Engineering; School of Engineering; Trinity College Dublin Dublin Ireland
| | | | - Simon Carroll
- Trinity Centre for Bioengineering; Trinity Biomedical Sciences Institute; Trinity College Dublin Dublin Ireland
- Department of Mechanical and Manufacturing Engineering; School of Engineering; Trinity College Dublin Dublin Ireland
| | - Kevin Mulhall
- Trinity Centre for Bioengineering; Trinity Biomedical Sciences Institute; Trinity College Dublin Dublin Ireland
- Royal College of Surgeons in Ireland; Dublin Ireland
| | - Daniel J. Kelly
- Trinity Centre for Bioengineering; Trinity Biomedical Sciences Institute; Trinity College Dublin Dublin Ireland
- Department of Mechanical and Manufacturing Engineering; School of Engineering; Trinity College Dublin Dublin Ireland
- Department of Anatomy; Royal College of Surgeons in Ireland; Dublin Ireland
- Advanced Materials and Bioengineering Research Centre (AMBER); Royal College of Surgeons in Ireland and Trinity College Dublin; Dublin Ireland
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9
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Pakhomov O, Martignat L, Honiger J, Clémenceau B, Saï P, Darquy S. AN69 Hollow Fiber Membrane will Reduce but Not Abolish the Risk of Transmission of Porcine Endogenous Retroviruses. Cell Transplant 2017; 14:749-56. [PMID: 16454349 DOI: 10.3727/000000005783982468] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022] Open
Abstract
As the risk of porcine endogenous retrovirus (PERV) infection is a major obstacle to the xenotransplantation of porcine tissue, we investigated whether an AN69 hollow fibre membrane, used for islets of Langerhans transplantation, could prevent the transfer of PERVs and thus reduce the risk of PERV infection. PK15 cells were used as a PERV source. A specific and highly sensitive RCR was used for detection of a PERV provirus DNA (gag region) and a porcine mtDNA. Human U293 cells were incubated in vitro with encapsulated PK15 cells, concentrated encapsulated PK15 supernatant, or concentrated PK15 supernatant as a control. CD1 mice were implanted in vivo with encapsulated PK15 cells or injected with PK15 supernatant. We found no infection in human cells incubated with either encapsulated PK15 supernatant or in 10 out of 11 samples after coincubation with encapsulated PK15 cells. Infection of human cells was, however, detected in 1 out of 11 samples after coincubation with encapsulated PK15 cells. The presence of PERV provirus DNA and porcine mtDNA was detected in all the investigated tissues of the mice injected with PK15 supernatant and in various tissues of the mice implanted with encapsulated PK15 cells. Four weeks after the last injection of PK15 supernatant or a fiber explantation, no mouse showed any presence of PERV provirus DNA or porcine mtDNA. Our results demonstrate that AN69 hollow fiber membrane will reduce but not abolish the risk of PERV infection. Because the real risk of PERV infection still remains unknown, it is necessary to investigate further the real protection that could be provided by hollow fibers to ensure the safety of clinical xenotransplantation.
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Affiliation(s)
- Oleg Pakhomov
- Biology of Nutrition, Paris 5 University Faculty of Pharmacy, France
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Edge AS, Gosse ME, Dinsmore J. Xenogeneic Cell Therapy: Current Progress and Future Developments in Porcine Cell Transplantation. Cell Transplant 2017; 7:525-39. [PMID: 9853581 DOI: 10.1177/096368979800700603] [Citation(s) in RCA: 35] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022] Open
Abstract
The multitude of distinct cell types present in mature and developing tissues display unique physiologic characteristics. Cellular therapy is a novel technology with the promise of utilizing this diversity to treat a wide range of human degenerative diseases. Intractable diseases, disorders, and injuries are characterized by cell death or aberrant cellular function. Cell transplantation can replace diseased or lost tissue to provide restorative therapy for these conditions. The limited use of cell transplants as a basis for current therapy can, in part, be attributed to the lack of available human cells suitable for transplantation. This has prevented further realization of the promise of cell transplantation as a platform technology. Accordingly, cell-based therapies such as blood transfusions, for which the cells are readily available, are a standard part of current medical practice. Despite numerous attempts to expand primary human cells in tissue culture, current technological limitations of this approach in regard to proliferative capacity and maintenance of the differentiated phenotype has prevented their use for transplantation. Further, use of human stem cells for the derivation of specific cell types for transplantation is an area of future application with great potential, but hurdles remain in regard to deriving and sufficiently expanding these multi-potential cells. Thus, it appears that primary cells are at present a superior source for transplantation. This review focuses on pigs as a source of a variety of primary cells to advance cell therapy to the clinic and implement achievement of its full potential. We outline the advantages and disadvantages of xenogeneic cell therapy while underscoring the utility of transplantable porcine cells for the treatment of human disease. © 1998 Elsevier Science Inc.
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Affiliation(s)
- A S Edge
- Diacrin Inc., Charlestown, MA 02129, USA
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11
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Sakiyama R, Blau BJ, Miki T. Clinical translation of bioartificial liver support systems with human pluripotent stem cell-derived hepatic cells. World J Gastroenterol 2017; 23:1974-1979. [PMID: 28373763 PMCID: PMC5360638 DOI: 10.3748/wjg.v23.i11.1974] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/04/2016] [Revised: 12/02/2016] [Accepted: 01/11/2017] [Indexed: 02/06/2023] Open
Abstract
There is currently a pressing need for alternative therapies to liver transplantation. The number of patients waiting for a liver transplant is substantially higher than the number of transplantable donor livers, resulting in a long waiting time and a high waiting list mortality. An extracorporeal liver support system is one possible approach to overcome this problem. However, the ideal cell source for developing bioartificial liver (BAL) support systems has yet to be determined. Recent advancements in stem cell technology allow researchers to generate highly functional hepatocyte-like cells from human pluripotent stem cells (hPSCs). In this mini-review, we summarize previous clinical trials with different BAL systems, and discuss advantages of and potential obstacles to utilizing hPSC-derived hepatic cells in clinical-scale BAL systems.
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12
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Kim N, Choi J, Kim S, Gwon YD, Cho Y, Yang JM, Oh YK, Kim YB. Transmission of Porcine Endogenous Retrovirus Produced from Different Recipient Cells In Vivo. PLoS One 2016; 11:e0165156. [PMID: 27832080 PMCID: PMC5104465 DOI: 10.1371/journal.pone.0165156] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2016] [Accepted: 09/29/2016] [Indexed: 12/04/2022] Open
Abstract
Humanized pigs have been developed to reduce the incidence of immune rejection in xenotransplantation, but significant concerns remain, such as transmission of viral zoonosis. Porcine endogenous retroviruses (PERV), which exist in the genome of pigs, are produced as infectious virions from all porcine cells and cause zoonosis. Here, we examined the possibility of zoonosis of hosts under conditions of immune suppression or xenotransplantation of cells producing host-adapted viruses. Upon transplantation of PERV-producing porcine cells into mice, no transmission of PERV was detected, whereas, transmission of PERV from mice transplanted with mouse-adapted PERV-producing cells was detected. In addition, the frequency of PERV transmission was increased in CsA treated mice transplanted with PERV-producing murine cells, compared with PERV-producing porcine cells. Transmission of PERV to host animals did not affect weight but immune responses, in particular, the number of T cells from PERV-transmitted mice, were notably reduced. The observed risk of PERV zoonosis highlights the requirement for thorough evaluation of viral zoonosis under particular host conditions, such as immunosuppressive treatment and transplantation with host-adapted virus-producing cells.
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Affiliation(s)
- Nayoung Kim
- Department of Life Sciences, Sogang University, Seoul, Republic of Korea
| | - Jiwon Choi
- Department of Bio-industrial Technologies, Konkuk University, Neungdong-ro, Gwangjin-gu, Seoul, Republic of Korea
| | - Sehyun Kim
- Department of Bio-industrial Technologies, Konkuk University, Neungdong-ro, Gwangjin-gu, Seoul, Republic of Korea
| | - Yong-Dae Gwon
- Department of Bio-industrial Technologies, Konkuk University, Neungdong-ro, Gwangjin-gu, Seoul, Republic of Korea
| | - Yeondong Cho
- Department of Bio-industrial Technologies, Konkuk University, Neungdong-ro, Gwangjin-gu, Seoul, Republic of Korea
| | - Jae Myung Yang
- Department of Life Sciences, Sogang University, Seoul, Republic of Korea
| | - Yu-Kyoung Oh
- College of Pharmacy and Research Institute of Pharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea
| | - Young bong Kim
- Department of Bio-industrial Technologies, Konkuk University, Neungdong-ro, Gwangjin-gu, Seoul, Republic of Korea
- * E-mail:
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13
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Shinoka T, Miyachi H. Current Status of Tissue Engineering Heart Valve. World J Pediatr Congenit Heart Surg 2016; 7:677-684. [DOI: 10.1177/2150135116664873] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2016] [Accepted: 07/26/2016] [Indexed: 12/31/2022]
Abstract
The development of surgically implantable heart valve prostheses has contributed to improved outcomes in patients with cardiovascular disease. However, there are drawbacks, such as risk of infection and lack of growth potential. Tissue-engineered heart valve (TEHV) holds great promise to address these drawbacks as the ideal TEHV is easily implanted, biocompatible, non-thrombogenic, durable, degradable, and ultimately remodels into native-like tissue. In general, three main components used in creating a tissue-engineered construct are (1) a scaffold material, (2) a cell type for seeding the scaffold, and (3) a subsequent remodeling process driven by cell accumulation and proliferation, and/or biochemical and mechanical signaling. Despite rapid progress in the field over the past decade, TEHVs have not been translated into clinical applications successfully. To successfully utilize TEHVs clinically, further elucidation of the mechanisms for TEHV remodeling and further translational research outcome evaluations will be required. Tissue engineering is a major breakthrough in cardiovascular medicine that holds amazing promise for the future of reconstructive surgical procedures. In this article, we review the history of regenerative medicine, advances in the field, and state-of-the-art in valvular tissue engineering.
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Affiliation(s)
- Toshiharu Shinoka
- Department of Cardiothoracic Surgery, Nationwide Children’s Hospital, Columbus, OH, USA
| | - Hideki Miyachi
- Department of Cardiothoracic Surgery, Nationwide Children’s Hospital, Columbus, OH, USA
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14
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Timsit MO, Branchereau J, Thuret R, Kleinclauss F. [Renal transplantation in 2046: Future and perspectives]. Prog Urol 2016; 26:1132-1142. [PMID: 27665406 DOI: 10.1016/j.purol.2016.08.023] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2016] [Revised: 08/24/2016] [Accepted: 08/25/2016] [Indexed: 11/25/2022]
Abstract
OBJECTIVES To report major findings that may build the future of kidney transplantation. MATERIAL AND METHODS Relevant publications were identified through Medline (http://www.ncbi.nlm.nih.gov) and Embase (http://www.embase.com) database from 1960 to 2016 using the following keywords, in association, "bio-engineering; heterotransplantation; immunomodulation; kidney; regenerative medicine; xenotransplantation". Articles were selected according to methods, language of publication and relevance. A total of 5621 articles were identified including 2264 for xenotransplantation, 1058 for regenerative medicine and 2299 for immunomodulation; after careful selection, 86 publications were eligible for our review. RESULTS Despite genetic constructs, xenotransplantation faces the inevitable obstacle of species barrier. Uncertainty regarding xenograft acceptance by recipients as well as ethical considerations due to the debatable utilization of animal lives, are major limits for its future. Regenerative medicine and tridimensional bioprinting allow successful implantation of organs. Bioengineering, using decellularized tissue matrices or synthetic scaffold, seeded with pluripotent cells and assembled using bioreactors, provide exciting results but remain far for reconstituting renal complexity and vascular patency. Immune tolerance may be achieved through a tough initial T-cell depletion or a combined haplo-identical bone marrow transplant leading to lymphohematopoietic chimerism. CONCLUSION Current researches aim to increase the pool of organs available for transplantation (xenotransplants and bio-artificial kidneys) and to increase allograft survival through the induction of immune tolerance. Reported results suggest the onset of a thrilling new era for renal transplantation providing end-stage renal disease-patients with an improved survival and quality of life.
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Affiliation(s)
- M-O Timsit
- Service d'urologie, hôpital européen Georges-Pompidou, AP-HP, 20, rue Leblanc, 75015 Paris, France; Université Paris-Descartes, 75006 Paris, France.
| | - J Branchereau
- Service d'urologie et transplantation, CHU de Nantes, 44000 Nantes, France
| | - R Thuret
- Service d'urologie et transplantation rénale, CHU de Montpellier, 34090 Montpellier, France; Université de Montpellier, 34090 Montpellier, France
| | - F Kleinclauss
- Service d'urologie et transplantation rénale, CHRU de Besançon, 25000 Besançon, France; Université de Franche-Comté, 25000 Besançon, France; Inserm UMR 1098, 25000 Besançon, France
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15
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Yoon JK, Choi J, Lee HJ, Cho Y, Gwon YD, Jang Y, Kim S, Choi H, Lee JH, Kim YB. Distribution of Porcine Endogenous Retrovirus in Different Organs of the Hybrid of a Landrace and a Jeju Domestic Pig in Korea. Transplant Proc 2016; 47:2067-71. [PMID: 26293098 DOI: 10.1016/j.transproceed.2015.05.023] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/16/2015] [Accepted: 05/22/2015] [Indexed: 01/30/2023]
Abstract
Xenotransplantation offers a solution to the shortage of available organs for transplantation, and the pig represents an ideal source of such organs. However, porcine endogenous retrovirus (PERV), whose genome is integrated in pigs, has been suggested to pose a potential risk of xenotransmission. Expression of PERVs in different organs of pigs was carefully measured at DNA, mRNA, and protein levels, providing information valuable for the application of pig organs in xenotransplantation. An analysis of PERV DNA showed that a very similar number of PERV copies was present in the genome of all organs, whereas mRNA and protein levels of PERV varied depending on the organ, with kidney, liver, and spleen expressing high levels of both mRNA and protein. In contrast, mRNA and protein levels were dissimilar in the lung and brain, where mRNA levels were low but protein levels were high. This discrepancy indicates that mRNA levels are not always reflected in protein expression. In addition, the difference between mRNA and protein highlights the importance of choosing the proper analysis method for diagnosing viral infection. In summary, this study provides insight into the distribution of PERV in various organs at the DNA, mRNA, and protein levels, and also informs the proper selection of tissues or organs for future clinical xenotransplantation.
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Affiliation(s)
- J K Yoon
- Department of Bio-industrial Technologies, Konkuk University, Seoul, Korea
| | - J Choi
- Department of Bio-industrial Technologies, Konkuk University, Seoul, Korea
| | - H J Lee
- Department of Bio-industrial Technologies, Konkuk University, Seoul, Korea
| | - Y Cho
- Department of Bio-industrial Technologies, Konkuk University, Seoul, Korea
| | - Y D Gwon
- Department of Bio-industrial Technologies, Konkuk University, Seoul, Korea
| | - Y Jang
- Department of Bio-industrial Technologies, Konkuk University, Seoul, Korea
| | - S Kim
- Department of Bio-industrial Technologies, Konkuk University, Seoul, Korea
| | - H Choi
- Department of Bio-industrial Technologies, Konkuk University, Seoul, Korea
| | - J H Lee
- College of Veterinary Medicine, Konkuk University, Seoul, Korea
| | - Y B Kim
- Department of Bio-industrial Technologies, Konkuk University, Seoul, Korea.
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Tang HB, Ouyang K, Rao GB, Ma L, Zhong H, Bai A, Qin S, Chen F, Lin J, Cao Y, Liao YJ, Zhang J, Wu J. Characterization of Complete Genome Sequences of a Porcine Endogenous Retrovirus Isolated From China Bama Minipig Reveals an Evolutionary Time Earlier Than That of Isolates From European Minipigs. Transplant Proc 2016; 48:222-8. [PMID: 26915872 DOI: 10.1016/j.transproceed.2015.12.005] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2015] [Accepted: 12/10/2015] [Indexed: 10/22/2022]
Abstract
BACKGROUND A porcine endogenous retroviruses (PERV) isolate, PERV-A-BM, was isolated from a Guangxi Bama minipig in China. METHODS To understand its genetic variation and evolution, the complete PERV-A-BM genome sequences were determined and compared with isolates from different Sus scrofa breeds and porcine cell lines. A total of 69 nucleotide substitutions were found in the full-length genome, including 26 non-synonymous mutations. RESULTS Phylogenetic trees based on the complete genome sequence as well as the gag, pol, and env gene sequences from 21 PERV isolates demonstrated that the PERV-A-BM was closely related to the EF133960 isolate from Chinese Wuzhishan miniature pigs inbred in Hainan, China, and distantly related to strains isolated from European-born pigs. CONCLUSIONS The estimation of age in the proviral PERV-A-BM integrating into the host genome reveals that the age of PERV-A-BM is at least 8.3 × 10(6) years, an evolutionary time earlier than that of isolates from European-born pigs.
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Affiliation(s)
- H-B Tang
- Guangxi Veterinary Research Institute, Nanning, Guangxi, China
| | - K Ouyang
- Guangxi Veterinary Research Institute, Nanning, Guangxi, China
| | - G-B Rao
- Guangxi Veterinary Research Institute, Nanning, Guangxi, China
| | - L Ma
- Guangxi Veterinary Research Institute, Nanning, Guangxi, China
| | - H Zhong
- School of Marine Sciences and Biotechnology, Guangxi University for Nationalities, Nanning, China
| | - A Bai
- Guangxi Veterinary Research Institute, Nanning, Guangxi, China
| | - S Qin
- Guangxi Veterinary Research Institute, Nanning, Guangxi, China
| | - F Chen
- Guangxi Veterinary Research Institute, Nanning, Guangxi, China
| | - J Lin
- Guangxi Veterinary Research Institute, Nanning, Guangxi, China
| | - Y Cao
- Guangxi Veterinary Research Institute, Nanning, Guangxi, China
| | - Y-J Liao
- School of Marine Sciences and Biotechnology, Guangxi University for Nationalities, Nanning, China
| | - J Zhang
- Laboratory for Viral Safety of National Centre of Biomedical Analysis, Institute of Transfusion Medicine, The Academy of Military Medical Sciences, Beijing, China
| | - J Wu
- Guangxi Veterinary Research Institute, Nanning, Guangxi, China.
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Choi J, Kim HM, Yoon JK, Cho Y, Lee HJ, Kim KC, Kim CK, Kim GW, Kim YB. Identification of Porcine Endogenous Retrovirus (PERV) packaging sequence and development of PERV packaging viral vector system. J Microbiol 2015; 53:348-53. [PMID: 25935307 DOI: 10.1007/s12275-015-5134-0] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/11/2015] [Revised: 04/09/2015] [Accepted: 04/13/2015] [Indexed: 12/11/2022]
Abstract
Studies of the retroviruses have focused on the specific interaction of the nucleocapsid protein with a packaging signal in the viral RNA as important for this selectivity, but the packaging signal in porcine endogenous retrovirus (PERV) has not been defined. Herein, we identified and analyzed this packaging signal in PERV and found hairpin structures with conserved tetranucleotides in their loops and nucleocapsid recognition sequences; both of which are key elements in the viral packaging signal of MLV. We evaluated packaging efficiency of sequence variants isolated from viral and proviral integrated genomes. All viral packaging sequences (Ψ) were identical, while five distinct packaging sequences were identified from proviral sources. One proviral sequence (Ψ1) was identical to that of the viral Ψ and had the highest packaging efficiency. Three variants (Ψ2, Ψ3, Ψ4) maintained key elements of the viral packaging signal, but had nucleotide replacements and consequently demonstrated reduced packaging efficiency. Despite of the same overall hairpin structure, the proviral variant (Ψ5) had only one GACG sequence in the hairpin loop and showed the lowest packaging efficiency other than ∆Ψ, in which the essential packaging sequence was removed. This result, thus, defined the packaging sequences in PERV and emphasized the importance of nucleotide sequence and RNA structure in the determination of packaging efficiency. In addition, we demonstrate efficient infection and gene expression from the PERV based viral vector, which may serve as a novel alternative to current retroviral expression systems.
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Affiliation(s)
- Jiwon Choi
- Department of Bioindustrial Technologies, Konkuk University, Seoul, 143-701, Korea
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Xenozoonoses. LABORATORY ANIMAL MEDICINE 2015. [PMCID: PMC7150069 DOI: 10.1016/b978-0-12-409527-4.00029-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
Immunological and technical advances have led to tremendous increases in the number of people potentially able to benefit from allotransplantation. Ironically, it is the success of the field that has led to a renewed interest in xenotransplantation during the past several decades. To a large part, this has occurred because of the great scarcity of human organ and tissue donors. However, it has expanded to include the use of cells from animals into humans such as porcine islet cells for diabetes or extracorporeal perfusion of human blood through animal organs or cells. Similar to allotransplantation, issues regarding transmission of infections from the graft to the human recipient were brought up for consideration with these procedures in the 1990s (Michaels and Simmons, 1994; Chapman et al., 1995; Hammel et al., 1998; Fishman et al., 1998). A risk for infection exists with the use of any biologic agent regardless of whether it is from a human or an animal source. Accordingly, transmission of infections from human organs, tissues, or cells is a well-recognized cause of disease after allotransplantation (Ison and Grossi, 2013; Green and Michaels, 2012). As the human graft shortage continues, newer cellular therapies are explored. Thus, attention continues to be given to the potential use of xenogeneic organs, tissues, or cells for human maladies through xenotransplantation. The potential for novel zoonotic infections to emerge because of xenotransplantation (xenozoonoses or xenosis) led to a debate on whether the field should be permitted to progress. This chapter reviews the issues of xenotransplantation related to infections from animals to humans. Lessons learned from infections with prior nonhuman primate xenotransplantation and human allotransplantation are used to help inform about risks with newer xenogeneic procedures. In addition, information on known zoonoses is reviewed to better develop constructs to decrease the hazard of infection with these novel procedures.
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Zoonosis as a Risk to the Xenograft Recipient and to Society: Theoretical Issues. Xenotransplantation 2014. [DOI: 10.1128/9781555818043.ch11] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
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Retroviruses and Xenotransplantation. Xenotransplantation 2014. [DOI: 10.1128/9781555818043.ch13] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
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Jung YD, Lee JR, Kim YJ, Ha HS, Oh KB, Im GS, Choi BH, Kim HS. Promoter activity analysis and methylation characterization of LTR elements of PERVs in NIH miniature pig. Genes Genet Syst 2014; 88:135-42. [PMID: 23832305 DOI: 10.1266/ggs.88.135] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/23/2022] Open
Abstract
The potential risk of porcine endogenous retrovirus (PERV) transmission is an important issue in xenotransplantation (pig-to-human transplantation). Long terminal repeats (LTRs) in PERV elements show promoter activity that could affect neighboring functional genes. The methylation status and promoter activities of 3 LTR structures (PERV-LTR1, LTR2, and LTR3 elements) belonging to the PERV-A family were examined using luciferase reporter genes in human liver cell lines (HepG2 and Hep3B). The PERV LTR3 element exhibited hypomethylation and stronger promoter activity than the other LTR elements in human liver cells. We also performed comparative sequences analysis of the PERV LTR elements by using bioinformatics tools. Our findings showed that several transcription factors such as Nkx2-2 and Elk-1 positively influenced the high transcriptional activity of the PERV LTR3 element.
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Affiliation(s)
- Yi-Deun Jung
- Department of Biological Sciences, College of Natural Sciences, Pusan National University, Busan 609-735, Republic of Korea
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Hasan J, Fisher J, Ingham E. Current strategies in meniscal regeneration. J Biomed Mater Res B Appl Biomater 2013; 102:619-34. [PMID: 24030973 DOI: 10.1002/jbm.b.33030] [Citation(s) in RCA: 30] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/09/2013] [Revised: 07/13/2013] [Accepted: 08/18/2013] [Indexed: 12/26/2022]
Abstract
The meniscus plays an important role in the biomechanics and tribology of the knee joint. Damage to or disease of the meniscus is now recognized to predispose to the development of osteoarthritis. Treatment of meniscal injury through arthroscopic surgery has become one of the most common orthopedic surgical procedures, and in the United States this can represent 10 to 20% of procedures related to the knee. The meniscus has a limited healing capacity constrained to the vascularized periphery and therefore, surgical repair of the avascular regions is not always feasible. Replacement and repair of the meniscus to treat injuries is being investigated using tissue engineering strategies. Promising as these approaches may be, there are, however, major barriers to overcome before translation to the clinic.
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Affiliation(s)
- Jahid Hasan
- Institute of Medical and Biological Engineering, Schools of Biomedical Sciences and Mechanical Engineering, University of Leeds, Leeds, LS2 9JT, UK
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Xiang S, Ma Y, Yan Q, Lv M, Zhao X, Yin H, Zhang N, Jia J, Yu R, Zhang J. Construction and characterization of an infectious replication competent clone of porcine endogenous retrovirus from Chinese miniature pigs. Virol J 2013; 10:228. [PMID: 23837947 PMCID: PMC3718662 DOI: 10.1186/1743-422x-10-228] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2012] [Accepted: 05/30/2013] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND Xenotransplantation from animals has been considered to be a preferable approach to alleviate the shortage of human allografts. Pigs are the most suitable candidate because of the anatomical and physiological similarities shared with humans as well as ethical concerns. However, it may be associated with the risk of transmission of infectious porcine pathogens. Porcine endogenous retroviruses (PERVs) are of particular concern because they have been shown to infect human cells in vitro. To date, researches on the molecular characteristics and potential pathogenicity of PERV are still tenuous. In this report, an infectious replication competent clone of PERV from Wuzhishan pigs (WZSPs) in China was generated and characterized. This infectious clone will contribute to studies on PERV virology and control of PERV in xenotransplantation using Chinese miniature pigs. METHODS The proviral DNA of PERV from WZSPs was amplified in two overlapping halves. Then the two fragments were isolated, subcloned and fused to generate pBluescriptαSK+-WZS-PERV recombinant clones. Screened with RT-PCR, a molecular clone of PERV designated as WZS-PERV(2) was selected. Its infectivity and replication competency were characterized in HEK293 cells by PCR, real-time fluorescent quantitative RT-PCR, western blot, indirect immunofluorescence assay as well as sequence analysis. RESULTS The ability of WZS-PERV(2) to infect human cells and produce infectious virions were shown after transfection of the clone into HEK293 cells and infection of PERV derived from this recombinant clone. The expression of Gag proteins were detected in HEK293 cells infected with the virus derived from the clone by the indirect immunofluorescence assay and western blot. The results of sequences analysis and comparison combined with the PCR based genotyping result demonstrated that the WZS-PERV(2) belonged to PERV-A subgroup. Compared with a previous proviral DNA clone of PERV (PERV-WZSP), G to A hypermutation occurred in the env gene of WZS-PERV(2) was found, whereas APOBEC proteins have the potential to inhibit the replication of a variety of retroviruses through a cDNA cytosine deamination mechanism, so we presumed these G to A hypermutation might be the contribution of porcine APOBEC3F. CONCLUSIONS Altogether, an infectious replication competent clone of PERV from Chinese miniature pigs (WZSPs) termed WZS-PERV(2) was generated, characterized and sequenced.
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Affiliation(s)
- Silong Xiang
- Key Laboratory of Drug Targeting and Drug Delivery System, Ministry of Education, West China School of Pharmacy, Sichuan University, Chengdu 610041, China
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Jung YD, Ha HS, Park SJ, Oh KB, Im GS, Kim TH, Seong HH, Kim HS. Identification and promoter analysis of PERV LTR subtypes in NIH-miniature pig. Mol Cells 2013; 35:99-105. [PMID: 23456331 PMCID: PMC3887905 DOI: 10.1007/s10059-013-2289-6] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2011] [Revised: 11/20/2012] [Accepted: 12/11/2012] [Indexed: 11/28/2022] Open
Abstract
Porcine endogenous retroviruses (PERVs) are integrated into the genomes of all pigs. Since some PERVs can also infect human cells, they represent a potential risk for xenotransplantation involving pig cells or organs. The long terminal repeat (LTR) elements of PERVs show promoter activity that can affect human functional genes; therefore, we examined these elements in this study. We detected several expressed LTRs in the NIH-miniature pig liver, among which we identified 9 different subtypes. When these LTRs were compared, distinct structures that contained several insertion and deletion (INDEL) events and tandem repeats were identified in the U3 region. The transcriptional activity of the 9 LTR subtypes was analyzed in the PK15 porcine cell line and in the HepG2 and Hep3B human liver cell lines, and transcriptional regulation was found to be different in the 3 cell lines. The D LTR subtype was found to have stronger promoter activity than all other types in 4 different human cell lines (HepG2, Hep3B, U251, and 293). Using computational approaches, the D type was shown to contain 4 transcription factor-binding sites distinct from those in the U3 regions of the other subtypes. Therefore, deletion mutants were constructed and examined by a transient transfection luciferase assay. The results of this analysis indicated that the binding site for the Hand1:E47 transcription factor might play a positive role in the transcriptional regulation of PERV LTR subtype D in human liver cell lines.
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Affiliation(s)
- Yi-Deun Jung
- Department of Biological Sciences, College of Natural Sciences, Pusan National University, Busan 609-735,
Korea
| | | | - Sang-Je Park
- Department of Biological Sciences, College of Natural Sciences, Pusan National University, Busan 609-735,
Korea
| | | | | | | | | | - Heui-Soo Kim
- Department of Biological Sciences, College of Natural Sciences, Pusan National University, Busan 609-735,
Korea
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Abstract
BACKGROUND Orthotopic liver transplantation (OLT) is the most effective therapy for liver failure. However, OLT is severely limited by the shortage of liver donors. Bioartificial liver (BAL) shows great potential as an alternative therapy for liver failure. In recent years, progress has been made in BAL regarding genetically engineered cell lines, immortalized human hepatocytes, methods for preserving the phenotype of primary human hepatocytes, and other functional hepatocytes derived from stem cells. DATA SOURCES A systematic search of PubMed and ISI Web of Science was performed to identify relevant studies in English language literature using the key words such as liver failure, bioartificial liver, hepatocyte, stem cells, differentiation, and immortalization. More than 200 articles related to the cell sources of hepatocyte in BAL were systematically reviewed. RESULTS Methods for preserving the phenotype of primary human hepatocytes have been successfully developed. Many genetically engineered cell lines and immortalized human hepatocytes have also been established. Among these cell lines, the incorporation of BAL with GS-HepG2 cells or alginate-encapsulated HepG2 cells could prolong the survival time and improve pathophysiological parameters in an animal model of liver failure. The cBAL111 cells were evaluated using the AMC-BAL bioreactor, which could eliminate ammonia and lidocaine, and produce albumin. Importantly, BAL loading with HepLi-4 cells could significantly improve the blood biochemical parameters, and prolong the survival time in pigs with liver failure. Other functional hepatocytes differentiated from stem cells, such as human liver progenitor cells, have been successfully achieved. CONCLUSIONS Aside from genetically modified liver cell lines and immortalized human hepatocytes, other functional hepatocytes derived from stem cells show great potential as cell sources for BAL. BAL with safe and effective liver cells may be achieved for clinical liver failure in the near future.
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Affiliation(s)
- Xiao-Ping Pan
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310003, China
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Fang X, Mou Y, Huang Z, Li Y, Han L, Zhang Y, Feng Y, Chen Y, Jiang X, Zhao W, Sun X, Xiong Z, Yang L, Liu H, Fan D, Mao L, Ren L, Liu C, Wang J, Li K, Wang G, Yang S, Lai L, Zhang G, Li Y, Wang J, Bolund L, Yang H, Wang J, Feng S, Li S, Du Y. The sequence and analysis of a Chinese pig genome. Gigascience 2012; 1:16. [PMID: 23587058 PMCID: PMC3626506 DOI: 10.1186/2047-217x-1-16] [Citation(s) in RCA: 104] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2012] [Accepted: 08/08/2012] [Indexed: 12/21/2022] Open
Abstract
Background The pig is an economically important food source, amounting to approximately 40% of all meat consumed worldwide. Pigs also serve as an important model organism because of their similarity to humans at the anatomical, physiological and genetic level, making them very useful for studying a variety of human diseases. A pig strain of particular interest is the miniature pig, specifically the Wuzhishan pig (WZSP), as it has been extensively inbred. Its high level of homozygosity offers increased ease for selective breeding for specific traits and a more straightforward understanding of the genetic changes that underlie its biological characteristics. WZSP also serves as a promising means for applications in surgery, tissue engineering, and xenotransplantation. Here, we report the sequencing and analysis of an inbreeding WZSP genome. Results Our results reveal some unique genomic features, including a relatively high level of homozygosity in the diploid genome, an unusual distribution of heterozygosity, an over-representation of tRNA-derived transposable elements, a small amount of porcine endogenous retrovirus, and a lack of type C retroviruses. In addition, we carried out systematic research on gene evolution, together with a detailed investigation of the counterparts of human drug target genes. Conclusion Our results provide the opportunity to more clearly define the genomic character of pig, which could enhance our ability to create more useful pig models.
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Affiliation(s)
- Xiaodong Fang
- BGI-Shenzhen, Bei Shan Road, Yantian, Shenzhen, 518083, China.
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Bittmann I, Mihica D, Plesker R, Denner J. Expression of porcine endogenous retroviruses (PERV) in different organs of a pig. Virology 2012; 433:329-36. [DOI: 10.1016/j.virol.2012.08.030] [Citation(s) in RCA: 44] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2012] [Revised: 06/21/2012] [Accepted: 08/18/2012] [Indexed: 10/27/2022]
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Denner J, Tönjes RR. Infection barriers to successful xenotransplantation focusing on porcine endogenous retroviruses. Clin Microbiol Rev 2012; 25:318-43. [PMID: 22491774 PMCID: PMC3346299 DOI: 10.1128/cmr.05011-11] [Citation(s) in RCA: 148] [Impact Index Per Article: 11.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2023] Open
Abstract
Xenotransplantation may be a solution to overcome the shortage of organs for the treatment of patients with organ failure, but it may be associated with the transmission of porcine microorganisms and the development of xenozoonoses. Whereas most microorganisms may be eliminated by pathogen-free breeding of the donor animals, porcine endogenous retroviruses (PERVs) cannot be eliminated, since these are integrated into the genomes of all pigs. Human-tropic PERV-A and -B are present in all pigs and are able to infect human cells. Infection of ecotropic PERV-C is limited to pig cells. PERVs may adapt to host cells by varying the number of LTR-binding transcription factor binding sites. Like all retroviruses, they may induce tumors and/or immunodeficiencies. To date, all experimental, preclinical, and clinical xenotransplantations using pig cells, tissues, and organs have not shown transmission of PERV. Highly sensitive and specific methods have been developed to analyze the PERV status of donor pigs and to monitor recipients for PERV infection. Strategies have been developed to prevent PERV transmission, including selection of PERV-C-negative, low-producer pigs, generation of an effective vaccine, selection of effective antiretrovirals, and generation of animals transgenic for a PERV-specific short hairpin RNA inhibiting PERV expression by RNA interference.
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Genetic prevalence of porcine endogenous retrovirus in chinese experimental miniature pigs. Transplant Proc 2012; 43:2762-9. [PMID: 21911159 DOI: 10.1016/j.transproceed.2011.06.061] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2011] [Revised: 06/13/2011] [Accepted: 06/16/2011] [Indexed: 11/21/2022]
Abstract
Pig-to-human xenotransplantation poses the potential risk of interspecies transmission of porcine endogenous retrovirus (PERV). The Chinese experimental miniature pig may be used as a pig-to-human xenograft donor. However, data for the distribution of PERV provirus in genomic DNA and PERV expression at the RNA level for the Chinese experimental miniature pig population are lacking. In this study, PERV was investigated in this regard using polymerase chain reaction (PCR), real-time quantitative PCR, and real-time quantitative reverse transcription PCR. The results showed that the genotype distribution was PERV-A subtype 100%, PERV-B subtype 100%, and PERV-C subtype 30% among 20 pig genomic DNA samples. Both PERV copy number in genomic DNA and PERV expression at the RNA level varied significantly among individuals, ranging from 3.95 ± 0.14 to 95.52 ± 2.20 and 3.66 ± 0.13 to 43.03 ± 2.50, respectively. For some individuals, the PERV copy number (eg, 3.95 ± 0.14) in genomic DNA and PERV expression (eg, 3.66 ± 0.13) at the RNA level were low. These results suggested that the Chinese experimental miniature pig is a possible donor for xenotransplantation. Our results provide reference information for selective breeding, which will benefit the application of these animals for the study of xenotransplantation.
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Abstract
Xenotransplantation, the transplantation of cells, tissues, or organs between different species, has the potential to overcome the current shortage of human organs and tissues for transplantation. In the last decade, the progress made in the field is remarkable, suggesting that clinical xenotransplantation procedures, particularly those involving cells, may become a reality in the not-too-distant future. However, several hurdles remain, mainly immunological barriers, physiological discrepancies, and safety issues, making xenotransplantion a complex and multidisciplinary discipline.
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Han B, Shi XL, Xiao JQ, Zhang Y, Chu XH, Gu JY, Tan JJ, Gu ZZ, Ding YT. Influence of chitosan nanofiber scaffold on porcine endogenous retroviral expression and infectivity in pig hepatocytes. World J Gastroenterol 2011; 17:2774-80. [PMID: 21734784 PMCID: PMC3122264 DOI: 10.3748/wjg.v17.i22.2774] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/15/2010] [Revised: 11/15/2010] [Accepted: 11/22/2010] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the influence of chitosan nanofiber scaffold on the production and infectivity of porcine endogenous retrovirus (PERV) expressed by porcine hepatocytes.
METHODS: Freshly isolated porcine hepatocytes were cultured with or without chitosan nanofiber scaffold (defined as Nano group and Hep group) for 7 d. The daily collection of culture medium was used to detect reverse transcriptase (RT) activity with RT activity assay kits and PERV RNA by reverse transcription-polymerase chain reaction (PCR) and real time PCR with the PERV specific primers. And Western blotting was performed with the lysates of daily retrieved cells to determine the PERV protein gag p30. Besides, the in-vitro infectivity of the supernatant was tested by incubating the human embryo kidney 293 (HEK293) cells.
RESULTS: The similar changing trends between two groups were observed in real time PCR, RT activity assay and Western blotting. Two peaks of PERV expression at 10H and Day 2 were found and followed by a regular decline. No significant difference was found between two groups except the significantly high level of PERV RNA at Day 6 and PERV protein at Day 5 in Nano group than that in Hep group. And in the in-vitro infection experiment, no HEK293 cell was infected by the supernatant.
CONCLUSION: Chitosan nanofiber scaffold might prolong the PERV secreting time in pig hepatocytes but would not obviously influence its productive amount and infectivity, so it could be applied in the bioartificial liver without the increased risk of the virus transmission.
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Nascimento FF, Gongora J, Charleston M, Tristem M, Lowden S, Moran C. Evolution of endogenous retroviruses in the Suidae: evidence for different viral subpopulations in African and Eurasian host species. BMC Evol Biol 2011; 11:139. [PMID: 21609472 PMCID: PMC3128044 DOI: 10.1186/1471-2148-11-139] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2010] [Accepted: 05/24/2011] [Indexed: 12/03/2022] Open
Abstract
Background Porcine endogenous retroviruses (PERVs) represent remnants of an exogenous form that have become integrated in the domestic pig (Sus scrofa) genome. Although they are usually inactive, the capacity of γ1 ERVs to infect human cells in vitro has raised concerns about xenotransplantation because the viruses could cross the species barrier to humans. Here we have analyzed the evolution of γ1 ERVs in ten species of Suidae (suids, pigs and hogs) from Eurasia and Africa using DNA sequences for their coding domains (gag, pro/pol and env genes). For comparison with γ1 PERVs, we have also analysed γ2 ERVs which in domestic pigs are known to be inactive and do not pose a risk to xenotransplantation. Results Phylogenetic analysis using Bayesian inference showed that γ1 and γ2 ERVs have distinctive evolutionary histories. Firstly, two different viral lineages of γ1 ERVs were found and a coevolutionary analysis demonstrated that they correspond broadly to their host phylogeny, one of Eurasian and another of African species, and show no evidence of horizontal transmission. γ2 ERVs, however, show a bush-like evolution, suggesting a rapid viral radiation from a single common ancestor with no correspondence between host and viral evolutionary trees. Furthermore, though γ1 ERV env genes do not possess frequent stop codons, γ2 env genes do. To understand whether γ1 suid ERVs may be still replicating, we have also evaluated their likely mechanism of proliferation by statistically testing internal to terminal branches using nonsynonymous versus synonymous substitution ratios. Our results suggest that γ1 ERVs are increasing in copy number by reinfection, which requires the translocation of the virus from one cell to another. Conclusions Evidence of at least two viral subpopulations was observed in γ1 ERVs from Eurasian and African host species. These results should be taken into account in xenotransplantation since γ1 ERVs appear to be codiverging with their host and maintaining ongoing capacity to infect somatic and germ cells.
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Di Nicuolo G, D'Alessandro A, Andria B, Scuderi V, Scognamiglio M, Tammaro A, Mancini A, Cozzolino S, Di Florio E, Bracco A, Calise F, Chamuleau RAFM. Long-term absence of porcine endogenous retrovirus infection in chronically immunosuppressed patients after treatment with the porcine cell-based Academic Medical Center bioartificial liver. Xenotransplantation 2011; 17:431-9. [PMID: 21158944 DOI: 10.1111/j.1399-3089.2010.00617.x] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
BACKGROUND Clinical use of porcine cell-based bioartificial liver (BAL) support in acute liver failure as bridging therapy for liver transplantation exposes the patient to the risk of transmission of porcine endogenous retroviruses (PERVs) to human. This risk may be enhanced when patients receive liver transplant and are subsequently immunosuppressed. As further follow-up of previously reported patients (Di Nicuolo et al. 2005), an assessment of PERV infection was made in the same patient population pharmacologically immunosuppressed for several years after BAL treatment and in healthcare workers (HCWs) involved in the clinical trial at that time. METHODS Plasma and peripheral blood mononuclear cells (PBMCs) from eight patients treated with the Academic Medical Center-BAL (AMC-BAL), who survived to transplant, and 13 HCWs, who were involved in the trial, were assessed to detect PERV infection. A novel quantitative real-time polymerase chain reaction assay has been used. RESULTS Eight patients who received a liver transplant after AMC-BAL treatment are still alive under long-term pharmacological immunosuppression. The current clinical follow-up ranges from 5.6 to 8.7 yr after BAL treatment. A new q-real-time PCR assay has been developed and validated to detect PERV infection. The limit of quantification of PERV DNA was ≥ 5 copies per 1 × 10(5) PBMCs. The linear dynamic range was from 5 × 10(0) to 5 × 10(6) copies. In both patients and HCWs, neither PERV DNA in PBMCs nor PERV RNA in plasma and PBMC samples have been found. CONCLUSION Up to 8.7 yr after exposure to treatment with porcine liver cell-based BAL, no PERV infection has been found in long-term immunosuppressed patients and in HCWs by a new highly sensitive and specific q-real-time PCR assay.
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Generation of neutralising antibodies against porcine endogenous retroviruses (PERVs). Virology 2011; 411:78-86. [DOI: 10.1016/j.virol.2010.12.032] [Citation(s) in RCA: 47] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2010] [Revised: 09/20/2010] [Accepted: 12/17/2010] [Indexed: 11/19/2022]
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Park SJ, Huh JW, Kim DS, Ha HS, Jung YD, Ahn K, Oh KB, Park EW, Chang KT, Kim HS. Analysis of the molecular and regulatory properties of active porcine endogenous retrovirus gamma-1 long terminal repeats in kidney tissues of the NIH-Miniature pig. Mol Cells 2010; 30:319-25. [PMID: 20811814 DOI: 10.1007/s10059-010-0121-0] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2010] [Revised: 06/10/2010] [Accepted: 06/28/2010] [Indexed: 11/26/2022] Open
Abstract
The pig genome contains the gamma 1 family of porcine endogenous retroviruses (PERVs), which are a major obstacle to the development of successful xenotransplantation from pig to human. Long terminal repeats (LTRs) found in PERVs are known to be essential elements for the control of the transcriptional activity of single virus by different transcription factors (TFs). To identify transcribed PERV LTR elements, RT-PCR and DNA sequencing analyses were performed. Twenty-nine actively transcribed LTR elements were identified in the kidney tissues of the NIH-Miniature pig. These elements were divided into two major groups (I and II), and four minor groups (I-1, I-2, I-3, and II-1), by the presence of insertion and deletion (INDEL) sequences. Group I elements showed strong transcriptional activity compared to group II elements. Four different LTR elements (PL1, PL2, PL3, and PL4) as representative of the groups were analyzed by using a transient transfection assay. The regulation of their promoter activity was investigated by treatment with M.SssI (CpG DNA methyltransferase) and garcinol (histone acetyltransferase inhibitor). The transcriptional activity of PERV LTR elements was significantly reduced by treatment with M.SssI. These data indicate that transcribed PERV LTR elements harbor sufficient promoter activity to regulate the transcription of a single virus, and the transcriptional activity of PERV LTRs may be controlled by DNA methylation events.
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Affiliation(s)
- Sang-Je Park
- Department of Biological Sciences, College of Natural Sciences, Pusan National University, Busan, 609-735, Korea
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Frühauf JH, Mertsching H, Giri S, Frühauf NR, Bader A. Porcine endogenous retrovirus released by a bioartificial liver infects primary human cells. Liver Int 2009; 29:1553-61. [PMID: 19686312 DOI: 10.1111/j.1478-3231.2009.02087.x] [Citation(s) in RCA: 29] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/13/2023]
Abstract
BACKGROUND Porcine endogenous retrovirus (PERV) remains a safety risk in pig-to-human xenotransplantation. There is no evidence of in vivo productive infection in humans because PERV is inactivated by human serum. However, PERV can infect human cell lines and human primary cells in vitro and inhibit human immune functions. AIMS We investigated the potential of primary porcine liver cells to transmit PERV to primary human cells in a bioreactor-based bioartificial liver (BAL). METHODS Primary human hepatocytes, endothelial cells and the human cell line HEK 293 were exposed to supernatants from BAL or from the porcine cell line PK-15. PERV polymerase-specific reverse-transcriptase polymerase chain reaction (RT-PCR) and PCR were used to investigate PERV transmission to human cells. An assay of RT activity was used to detect the presence of retrovirus in the supernatants of BAL, primary human hepatocytes and endothelial cells. RESULTS Primary human hepatocytes (hHep), endothelial cells and HEK 293 cells were reproducibly infected by PERV, originating from primary porcine liver cells within the BAL and from PK-15 cells. Infected cells were positive for PERV-specific DNA and RNA after 8-10 days on an average, and RT activity was detectable in the supernatants of infected hHep and HEK 293 cells. CONCLUSION A risk of PERV infection in human cells is documented in this study, indicating that short-term contact of primary porcine liver cell supernatants with primary human cells could result in PERV transmission.
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Affiliation(s)
- Jan-Henning Frühauf
- Department of Cell Techniques and Applied Stem Cell Biology, Biomedical-Biotechnological Center (BBZ), Leipzig, Germany
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39
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Inhibition of porcine endogenous retrovirus (PERV) replication by HIV-1 gene expression inhibitors. Antiviral Res 2009; 83:201-4. [DOI: 10.1016/j.antiviral.2009.04.011] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/11/2009] [Revised: 04/14/2009] [Accepted: 04/28/2009] [Indexed: 10/20/2022]
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40
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Garkavenko O, Wynyard S, Nathu D, Simond D, Muzina M, Muzina Z, Scobie L, Hector RD, Croxson MC, Tan P, Elliott BR. Porcine endogenous retrovirus (PERV) and its transmission characteristics: a study of the New Zealand designated pathogen-free herd. Cell Transplant 2009; 17:1381-8. [PMID: 19364075 DOI: 10.3727/096368908787648056] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022] Open
Abstract
Previously a strategy for monitoring of pigs intended for cell transplantation was developed and successfully applied to several representative herds in New Zealand. A designated pathogen-free (DPF) herd has been chosen as a good candidate for xenotransplantation. This herd has previously tested free of infectious agents relevant to xenotransplantation and we present here an in depth study of porcine endogenous retrovirus (PERV) transmission. A panel of assays that describes the constraints for the transmission of PERV has been suggested. It includes a) infectivity test in coculture of DPF pig primary cells with both human and pig target cell lines; b) RT activity in supernatant of stimulated primary cells from DPF pigs; c) viral load in donor's blood plasma; d) PERV proviral copy number in DPF pig genome; e) PERV class C prevalence in the herd and its recombination potential. There was no evidence of PERV transmission from DPF pig tissue to either pig or human cells. Additionally, there was no evidence of PERV RNA present in pig blood plasma. PERV copy number differs in individual pigs from as low as 3 copies to 30 copies and the presence of PERV-C varied between animals and breeds. In all DPF pigs tested, a specific locus for PERV-C potentially associated with the recombination of PERV in miniature swine was absent. Presented data on the PERV transmission allows us to classify the DPF potential donors as "null" or noninfectious pigs.
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Affiliation(s)
- O Garkavenko
- Living Cell Technologies Ltd, Manukau 2025, Auckland, New Zealand.
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41
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Kim NY, Lee D, Lee J, Park EW, Jung WW, Yang JM, Kim YB. Characterization of the replication-competent porcine endogenous retrovirus class B molecular clone originated from Korean domestic pig. Virus Genes 2009; 39:210-6. [DOI: 10.1007/s11262-009-0377-7] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2008] [Accepted: 06/06/2009] [Indexed: 10/20/2022]
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42
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Ramsoondar J, Vaught T, Ball S, Mendicino M, Monahan J, Jobst P, Vance A, Duncan J, Wells K, Ayares D. Production of transgenic pigs that express porcine endogenous retrovirus small interfering RNAs. Xenotransplantation 2009; 16:164-80. [DOI: 10.1111/j.1399-3089.2009.00525.x] [Citation(s) in RCA: 113] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
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Polymerase chain reaction in detection of porcine endogenous retrovirus (PERV) from porcine tissues. Indian J Microbiol 2009; 49:68-71. [PMID: 23100752 DOI: 10.1007/s12088-009-0002-4] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/21/2008] [Accepted: 03/13/2008] [Indexed: 10/21/2022] Open
Abstract
Pigs offer an unlimited source of xenografts for humans. The use of transplants from animal origin offers a potential solution to the limited supply of human organs and tissues. However, like many other mammalian species, pigs harbor porcine endogenous retrovirus (PERV), which are encoded in their genomic DNA and are assumed to have been integrated into the porcine germline. The ability of PERV to infect human cells in vitro has heightened safety concerns regarding the transmission of PERV to pig xenograft recipients. Porcine tissues were analyzed using validated assays specifi c for PERV: polymerase chain reaction (PCR) (for PERV DNA) and reverse transcriptase (RT)-PCR (for PERV RNA). PERV-specifi c gag sequences were found in the porcine heart tissue samples using DNA-PCR and RT-PCR. PCR is a rapid and specifi c test for the detection of PERV from xenografts. These fi ndings have demonstrated that the presence of both DNA and RNA forms of PERV in porcine tissues needs to be carefully considered when the infectious disease potential of xenotransplantation is being assessed.
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44
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Dieckhoff B, Kessler B, Jobst D, Kues W, Petersen B, Pfeifer A, Kurth R, Niemann H, Wolf E, Denner J. Distribution and expression of porcine endogenous retroviruses in multi-transgenic pigs generated for xenotransplantation. Xenotransplantation 2009; 16:64-73. [DOI: 10.1111/j.1399-3089.2009.00515.x] [Citation(s) in RCA: 68] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
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Lee D, Kim NY, Bae GE, Lee HJ, Kwon M, Kim SS, Lee HT, Yang JM, Kim YB. Transmissible infection of human 293T cells with porcine endogenous retroviruses subgroup a from NIH-miniature pig. Transplant Proc 2009; 40:3742-5. [PMID: 19100479 DOI: 10.1016/j.transproceed.2008.09.035] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2007] [Revised: 04/30/2008] [Accepted: 09/03/2008] [Indexed: 11/28/2022]
Abstract
In pig-to-human xenotransplantation, zoonotic infections have been an important barrier. The risk of zoonosis has been emphasized in xenotransplantation after finding that porcine endogenous retroviruses (PERVs) can infect human cells in vitro. Until now, transmissions of PERVs from PK15 cells have been studied in vitro and in vivo, but transmission of PERVs originating from miniature pigs have not been extensively reported. Peripheral blood mononuclear cells from miniature swine showed PERV transmission to human cells. In contrast, specific pathogen-free (SPF) pig islet cells showed no PERV transmission when co-incubated with 293T cells. To evaluate the risk of zoonosis with our experimental mini pigs, we tested the infectivity of PERVs from NIH-miniature pig primary ear cells for human 293T cells. As a result, all subgroups of infectious PERV virion (PERV-A, -B, and -C) were detected in the primary cell culture media. Unlike PERV-C, PERV-A and -B infected human 293T cells. Interestingly, only proviral PERV-A replicated in 293T cells to produce virions after infection. Our results suggested that a prevention study of PERV xenotransmission from experimental miniature pigs should concentrate on PERV-A control.
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Affiliation(s)
- D Lee
- Department of Animal Biotechnology, College of Animal Bioscience and Technology, Konkuk University, Korea
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46
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Huh JW, Kim DS, Ha HS, Ahn K, Chang KT, Cho BW, Kim HS. Identification and molecular characterization of PERV gamma1 long terminal repeats. Mol Cells 2009; 27:119-23. [PMID: 19214442 DOI: 10.1007/s10059-009-0013-3] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2008] [Revised: 11/06/2008] [Accepted: 11/11/2008] [Indexed: 10/21/2022] Open
Abstract
Porcine endogenous retroviruses (PERVs) gamma1 in the pig genome have the potential to act as harmful factors in xenotransplantation (pig-to-human). Long terminal repeats (LTRs) are known to be strong promoter elements that could control the transcription activity of PERV elements and the adjacent functional genes. To investigate the transcribed PERV gamma1 LTR elements in pig tissues, bioin-formatic and experimental approaches were conducted. Using RT-PCR amplification and sequencing approaches, 69 different transcribed LTR elements were identified. And 69 LTR elements could be divided into six groups (15 subgroups) by internal variation including tandem repeated sequences, insertion and deletion (INDEL). Remarkably, all internal variations were indentified in U3 region of LTR elements. Taken together, the identification and characterization of various PERV LTR transcripts allow us to extend our knowledge of PERV and its transcriptional study.
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Affiliation(s)
- Jae-Won Huh
- Division of Biological Sciences, College of Natural Sciences, Pusan National University, Busan, 609-735, Korea
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47
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Liver Substitution. Artif Organs 2009. [DOI: 10.1007/978-1-84882-283-2_4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
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Ma Y, Lv M, Xu S, Wu J, Tian K, Zhang J. Identification of full-length proviral DNA of porcine endogenous retrovirus from Chinese Wuzhishan miniature pigs inbred. Comp Immunol Microbiol Infect Dis 2008; 33:323-31. [PMID: 19070900 DOI: 10.1016/j.cimid.2008.10.007] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 10/25/2008] [Indexed: 10/21/2022]
Abstract
Existence of porcine endogenous retrovirus (PERV) hinders pigs to be used in clinical xenotransplantation to alleviate the shortage of human transplants. Chinese miniature pigs are potential organ donors for xenotransplantation in China. However, so far, an adequate level of information on the molecular characteristics of PERV from Chinese miniature pigs has not been available. We described here the cloning and characterization of full-length proviral DNA of PERV from Chinese Wuzhishan miniature pigs inbred (WZSP). Full-length nucleotide sequences of PERV-WZSP and other PERVs were aligned and phylogenetic tree was constructed from deduced amino-acid sequences of env. The results demonstrated that the full-length proviral DNA of PERV-WZSP belongs to gammaretrovirus and shares high similarity with other PERVs. Sequence analysis also suggested that different patterns of LTR existed in the same porcine germ line and partial PERV-C sequence may recombine with PERV-A sequence in LTR.
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Affiliation(s)
- Yuyuan Ma
- Laboratory for Viral Safety of National Centre of Biomedical Analysis, Institute of Transfusion Medicine, The Academy of Military Medical Sciences, Beijing 100850, China
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49
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Garkavenko O, Dieckhoff B, Wynyard S, Denner J, Elliott RB, Tan PL, Croxson MC. Absence of transmission of potentially xenotic viruses in a prospective pig to primate islet xenotransplantation study. J Med Virol 2008; 80:2046-52. [PMID: 18814261 DOI: 10.1002/jmv.21272] [Citation(s) in RCA: 55] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Shortage of human donor organs for transplantation has prompted usage of animals as an alternative donor source. Pigs are the most acceptable candidate animals but issues of xenozoonoses remain. Despite careful monitoring of designated pathogen free pigs there is still a risk that their tissues may carry infectious agents. Thus xenotransplantation requires extensive pre-clinical study on safety of the graft especially for those viruses that are either potentially oncogenic and/or immunosuppressive, or can establish persistent infection. A prospective pig-to-primate islet xenotransplantation study was performed which includes monitoring for potentially xenotic viruses namely porcine endogenous retrovirus (PERV), porcine cytomegalovirus (PCMV), porcine lymphotropic herpesvirus (PLHV), and porcine circovirus (PCV) using both molecular diagnostic-PCR and RT-PCR and serology methods. There was no evidence of pig virus transmission into primate recipients. This preclinical study underlines the information concerning viral safety of islet cell xenograft in pig-to-primate xenotransplantation.
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Affiliation(s)
- Olga Garkavenko
- Molecular Diagnostic Laboratory, Living Cell Technologies NZ Ltd, Auckland, New Zealand.
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50
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Gu C, Wei X, Wang Y, Chen Y, Liu J, Wang H, Sun G, Yi D. No infection with porcine endogenous retrovirus in recipients of acellular porcine aortic valves: a two-year study. Xenotransplantation 2008; 15:121-8. [PMID: 18447885 DOI: 10.1111/j.1399-3089.2008.00447.x] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
BACKGROUND Engineered tissue heart valves may become a promising therapeutics for heart valve disease. Compared with synthetic materials, acellular porcine scaffolds are considered as suitable matrices for tissue-engineered heart valves for the mechanical and structural properties of native tissue. Whether acellular porcine scaffolds can cause infection in recipients with porcine endogenous retrovirus (PERV) is critical for evaluating the safety of transplantation of tissue-engineered heart valves based on acellular porcine scaffolds. This study was completed to evaluate the risk of PERV transmission for application of acellular porcine aortic valves (PAVs). METHODS Native aortic valves obtained from Chinese pigs of different species were acellularized by our modified detergent-enzymatic acellularization procedure. Polymerase chain reaction and reverse transcriptase polymerase chain reaction for pol sequences were used to detect PERV infection. In vitro, ovine endothelial cells (ECs) were inoculated and cocultured with supernatants of porcine aortic endothelial cells (PAECs) and acellualr PAVs, respectively. On 7th day, DNA and RNA of ovine ECs were isolated and tested for PERV. In vivo, acellular PAVs were implanted in the descending thoracic aorta in 40 sheep. Blood samples from the sheep and implanted acellular PAVs were collected 24 months after operation and tested for PERV. RESULTS All cells were removed from the PAVs. Acellularized PAVs were repopulated by autologous cells of the host 24 months after implantation. PERV was detected in all native PAVs, porcine blood samples, acellularized PAVs and ovine ECs inoculated with supernatants of PAECs; no PERV was detected in ovine ECs cocultured with acellularized PAVs, ovine blood samples, and implanted acellular PAVs. CONCLUSIONS Acellularized PAVs processed by modified detergent-enzymatic acellularization procedure can be used for cardiovascular tissue-engineered grafts as matrix scaffolds without risk of PERV transmission.
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Affiliation(s)
- Chunhu Gu
- Department of Cardiovascular Surgery, Xijing Hospital, Fourth Military Medical University, Xi'an, Shaanxi Province, China
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