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1
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Henning HHW, Batz-Schott J, Grünther B, Le Thi X, Waberski D. Fluorescent labelling of boar spermatozoa for quantitative studies on competitive sperm-oviduct binding. Reprod Fertil Dev 2020; 31:1520-1532. [PMID: 31072452 DOI: 10.1071/rd19081] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2018] [Accepted: 04/08/2019] [Indexed: 01/13/2023] Open
Abstract
Invitro sperm-oviduct binding assays enable assessment of the capacity of spermatozoa to form a 'reservoir' in the oviduct. Competitive approaches, such as experimental set-ups that test multiple males or semen samples simultaneously on the same tissue explants, are desirable because they reduce the likelihood of bias when using material from different females. Therefore, we established a fluorescent labelling technique that allows tagging and storage of spermatozoa before competitive studies of sperm-oviduct binding invitro. Fluorescent markers were tested for reliability and compatibility with parameters of boar spermatozoa viability. The addition of seminal plasma after density gradient centrifugation was essential to counteract centrifugation stress during the labelling procedure. It was demonstrated that sperm tagged with MitoTracker Green FM or MitoTracker Red FM can be successfully used in competitive sperm-oviduct binding studies. The assay was sensitive enough to indicate subtle effects of semen storage temperature on the ability of the spermatozoa to contribute to the female sperm reservoir.
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Affiliation(s)
- Heiko H W Henning
- Unit for Reproductive Medicine, Clinic for Pigs and Small Ruminants, University of Veterinary Medicine, Bünteweg 15, 30559 Hannover, Germany; and Department of Equine Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 112, 3584CM Utrecht, Netherlands
| | - Julia Batz-Schott
- Unit for Reproductive Medicine, Clinic for Pigs and Small Ruminants, University of Veterinary Medicine, Bünteweg 15, 30559 Hannover, Germany
| | - Benita Grünther
- Unit for Reproductive Medicine, Clinic for Pigs and Small Ruminants, University of Veterinary Medicine, Bünteweg 15, 30559 Hannover, Germany
| | - Xuyen Le Thi
- Unit for Reproductive Medicine, Clinic for Pigs and Small Ruminants, University of Veterinary Medicine, Bünteweg 15, 30559 Hannover, Germany
| | - Dagmar Waberski
- Unit for Reproductive Medicine, Clinic for Pigs and Small Ruminants, University of Veterinary Medicine, Bünteweg 15, 30559 Hannover, Germany; and Corresponding author.
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2
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Kumar D, Anand T, Talluri TR, Kues WA. Potential of transposon-mediated cellular reprogramming towards cell-based therapies. World J Stem Cells 2020; 12:527-544. [PMID: 32843912 PMCID: PMC7415244 DOI: 10.4252/wjsc.v12.i7.527] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/26/2020] [Revised: 05/09/2020] [Accepted: 05/28/2020] [Indexed: 02/07/2023] Open
Abstract
Induced pluripotent stem (iPS) cells present a seminal discovery in cell biology and promise to support innovative treatments of so far incurable diseases. To translate iPS technology into clinical trials, the safety and stability of these reprogrammed cells needs to be shown. In recent years, different non-viral transposon systems have been developed for the induction of cellular pluripotency, and for the directed differentiation into desired cell types. In this review, we summarize the current state of the art of different transposon systems in iPS-based cell therapies.
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Affiliation(s)
- Dharmendra Kumar
- Animal Physiology and Reproduction Division, ICAR-Central Institute for Research on Buffaloes, Hisar 125001, India.
| | - Taruna Anand
- NCVTC, ICAR-National Research Centre on Equines, Hisar 125001, India
| | - Thirumala R Talluri
- Equine Production Campus, ICAR-National Research Centre on Equines, Bikaner 334001, India
| | - Wilfried A Kues
- Friedrich-Loeffler-Institut, Institute of Farm Animal Genetics, Department of Biotechnology, Mariensee 31535, Germany
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3
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Lipták N, Bősze Z, Hiripi L. GFP transgenic animals in biomedical research: a review of potential disadvantages. Physiol Res 2019; 68:525-530. [PMID: 31342754 DOI: 10.33549/physiolres.934227] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023] Open
Abstract
Green Fluorescent protein (GFP) transgenic animals are accepted tools for studying various physiological processes, including organ development and cell migration. However, several in vivo studies claimed that GFP may impair transgenic animals' health. Glomerulosclerosis was observed in transgenic mice and rabbits with ubiquitous reporter protein expression. Heart-specific GFP expression evoked dilated cardiomyopathy and altered cardiac function in transgenic mouse and zebrafish lines, respectively. Moreover, growth retardation and increased axon swelling were observed in GFP and yellow fluorescent protein (YFP) transgenic mice, respectively. This review will focus on the potential drawbacks of the applications of GFP transgenic animals in biomedical research.
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Affiliation(s)
- N Lipták
- NARIC-Agricultural Biotechnology Institute, Animal Biotechnology Department, Gödöllő, Hungary.
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4
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Buermann A, Petkov S, Petersen B, Hein R, Lucas-Hahn A, Baars W, Brinkmann A, Niemann H, Schwinzer R. Pigs expressing the human inhibitory ligand PD-L1 (CD 274) provide a new source of xenogeneic cells and tissues with low immunogenic properties. Xenotransplantation 2018; 25:e12387. [DOI: 10.1111/xen.12387] [Citation(s) in RCA: 30] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/03/2017] [Revised: 12/27/2017] [Accepted: 01/12/2018] [Indexed: 01/01/2023]
Affiliation(s)
- Anna Buermann
- Transplant Laboratory; Department of General-, Visceral-, and Transplantation Surgery; Hannover Medical School; Hannover Germany
| | - Stoyan Petkov
- Institute of Farm Animal Genetics; Friedrich-Loeffler-Institut; Mariensee Germany
| | - Björn Petersen
- Institute of Farm Animal Genetics; Friedrich-Loeffler-Institut; Mariensee Germany
| | - Rabea Hein
- Transplant Laboratory; Department of General-, Visceral-, and Transplantation Surgery; Hannover Medical School; Hannover Germany
| | - Andrea Lucas-Hahn
- Institute of Farm Animal Genetics; Friedrich-Loeffler-Institut; Mariensee Germany
| | - Wiebke Baars
- Transplant Laboratory; Department of General-, Visceral-, and Transplantation Surgery; Hannover Medical School; Hannover Germany
| | - Antje Brinkmann
- Transplant Laboratory; Department of General-, Visceral-, and Transplantation Surgery; Hannover Medical School; Hannover Germany
| | - Heiner Niemann
- Institute of Farm Animal Genetics; Friedrich-Loeffler-Institut; Mariensee Germany
| | - Reinhard Schwinzer
- Transplant Laboratory; Department of General-, Visceral-, and Transplantation Surgery; Hannover Medical School; Hannover Germany
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5
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Secher JO, Ceylan A, Mazzoni G, Mashayekhi K, Li T, Muenthaisong S, Nielsen TT, Li D, Li S, Petkov S, Cirera S, Luo Y, Thombs L, Kadarmideen HN, Dinnyes A, Bolund L, Roelen BAJ, Schmidt M, Callesen H, Hyttel P, Freude KK. Systematic in vitro and in vivo characterization of Leukemia-inhibiting factor- and Fibroblast growth factor-derived porcine induced pluripotent stem cells. Mol Reprod Dev 2017; 84:229-245. [PMID: 28044390 PMCID: PMC6221014 DOI: 10.1002/mrd.22771] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2016] [Accepted: 12/16/2016] [Indexed: 12/14/2022]
Abstract
Derivation and stable maintenance of porcine induced pluripotent stem cells (piPSCs) is challenging. We herein systematically analyzed two piPSC lines, derived by lentiviral transduction and cultured under either leukemia inhibitory factor (LIF) or fibroblast growth factor (FGF) conditions, to shed more light on the underlying biological mechanisms of porcine pluripotency. LIF‐derived piPSCs were more successful than their FGF‐derived counterparts in the generation of in vitro chimeras and in teratoma formation. When LIF piPSCs chimeras were transferred into surrogate sows and allowed to develop, only their prescence within the embryonic membranes could be detected. Whole‐transcriptome analysis of the piPSCs and porcine neonatal fibroblasts showed that they clustered together, but apart from the two pluripotent cell populations of early porcine embryos, indicating incomplete reprogramming. Indeed, bioinformatic analysis of the pluripotency‐related gene network of the LIF‐ versus FGF‐derived piPSCs revealed that ZFP42 (REX1) expression was absent in both piPSC‐like cells, whereas it was expressed in the porcine inner cell mass at Day 7/8. A second striking difference was the expression of ATOH1 in piPSC‐like cells, which was absent in the inner cell mass. Moreover, our gene expression analyses plus correlation analyses of known pluripotency genes identified unique relationships between pluripotency genes in the inner cell mass, which are to some extent, in the piPSC‐like cells. This deficiency in downstream gene activation and divergent gene expression may be underlie the inability to derive germ line‐transmitting piPSCs, and provides unique insight into which genes are necessary to achieve fully reprogrammed piPSCs. 84: 229–245, 2017. © 2016 Wiley Periodicals, Inc.
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Affiliation(s)
- Jan O Secher
- Veterinary Reproduction and Obstetrics, Faculty of Health and Medical Sciences, Department of Large Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark
| | - Ahmet Ceylan
- Faculty of Veterinary Medicine Ankara University, Department of Histology and Embryology, Diskapi, Ankara, Turkey
| | - Gianluca Mazzoni
- Animal Breeding, Quantitative Genetics and Systems Biology Group, Faculty of Health and Medical Sciences, Department of Large Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark
| | - Kaveh Mashayekhi
- Faculty of Health and Medical Sciences, Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark.,BioTalentum Ltd., Gödöllő, Hungary.,Faculty of Veterinary Medicine, Departments of Equine Sciences and Farm Animal Health, Utrecht University, Utrecht, Netherlands
| | - Tong Li
- Faculty of Health and Medical Sciences, Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark.,BioTalentum Ltd., Gödöllő, Hungary.,Faculty of Veterinary Medicine, Departments of Equine Sciences and Farm Animal Health, Utrecht University, Utrecht, Netherlands
| | - Suchitra Muenthaisong
- BioTalentum Ltd., Gödöllő, Hungary.,Faculty of Veterinary Medicine, Department of Farm Animal Health, Utrecht University, Utrecht, Netherlands
| | - Troels T Nielsen
- Danish Dementia Research Centre, Rigshospitalet, University of Copenhagen, Copenhagen, Denmark
| | - Dong Li
- Faculty of Health and Medical Sciences, Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark
| | - Shengting Li
- Department of Biomedicine, Aarhus University, Aarhus C, Denmark
| | - Stoyan Petkov
- Institute for Farm Animal Genetics (FLI), Neustadt, Germany
| | - Susanna Cirera
- Faculty of Health and Medical Sciences, Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark
| | - Yonglun Luo
- Department of Biomedicine, Aarhus University, Aarhus C, Denmark
| | - Lori Thombs
- Department of Statistics, University of Missouri, Columbia, Missouri
| | - Haja N Kadarmideen
- Animal Breeding, Quantitative Genetics and Systems Biology Group, Faculty of Health and Medical Sciences, Department of Large Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark
| | - Andras Dinnyes
- BioTalentum Ltd., Gödöllő, Hungary.,Faculty of Veterinary Medicine, Departments of Equine Sciences and Farm Animal Health, Utrecht University, Utrecht, Netherlands.,Molecular Animal Biotechnology Laboratory, Szent István University, Gödöllő, Hungary
| | - Lars Bolund
- Department of Biomedicine, Aarhus University, Aarhus C, Denmark
| | - Bernard A J Roelen
- Faculty of Veterinary Medicine, Department of Farm Animal Health, Utrecht University, Utrecht, Netherlands
| | - Mette Schmidt
- Veterinary Reproduction and Obstetrics, Faculty of Health and Medical Sciences, Department of Large Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark
| | - Henrik Callesen
- Department of Animal Science, Aarhus University, Tjele, Denmark
| | - Poul Hyttel
- Faculty of Health and Medical Sciences, Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark
| | - Kristine K Freude
- Faculty of Health and Medical Sciences, Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark
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6
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Zeng F, Li Z, Cai G, Gao W, Jiang G, Liu D, Urschitz J, Moisyadi S, Wu Z. Characterization of Growth and Reproduction Performance, Transgene Integration, Expression, and Transmission Patterns in Transgenic Pigs Produced by piggyBac Transposition-Mediated Gene Transfer. Anim Biotechnol 2017; 27:245-55. [PMID: 27565868 DOI: 10.1080/10495398.2016.1178140] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/21/2022]
Abstract
Previously we successfully produced a group of EGFP-expressing founder transgenic pigs by a newly developed efficient and simple pig transgenesis method based on cytoplasmic injection of piggyBac plasmids. In this study, we investigated the growth and reproduction performance and characterized the transgene insertion, transmission, and expression patterns in transgenic pigs generated by piggyBac transposition. Results showed that transgene has no injurious effect on the growth and reproduction of transgenic pigs. Multiple copies of monogenic EGFP transgene were inserted at noncoding sequences of host genome, and passed from founder transgenic pigs to their transgenic offspring in segregation or linkage manner. The EGFP transgene was ubiquitously expressed in transgenic pigs, and its expression intensity was associated with transgene copy number but not related to its promoter DNA methylation level. To the best of our knowledge, this is first study that fully described the growth and reproduction performance, transgene insertion, expression, and transmission profiles in transgenic pigs produced by piggyBac system. It not only demonstrates that piggyBac transposition-mediated gene transfer is an effective and favorable approach for pig transgenesis, but also provides scientific information for understanding the transgene insertion, expression and transmission patterns in transgenic animals produced by piggyBac transposition.
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Affiliation(s)
- Fang Zeng
- a National Engineering Research Center for Breeding Swine Industry, College of Animal Science , South China Agricultural University , Guangzhou , China.,b Guangdong Provincial Key Laboratory of Agro-animal Genomics and Molecular Breeding, College of Animal Science , South China Agricultural University , Guangzhou , China
| | - Zicong Li
- a National Engineering Research Center for Breeding Swine Industry, College of Animal Science , South China Agricultural University , Guangzhou , China.,b Guangdong Provincial Key Laboratory of Agro-animal Genomics and Molecular Breeding, College of Animal Science , South China Agricultural University , Guangzhou , China
| | - Gengyuan Cai
- c Institute of Animal Science , Guangdong Academy of Agricultural Sciences , Guangzhou , China
| | - Wenchao Gao
- a National Engineering Research Center for Breeding Swine Industry, College of Animal Science , South China Agricultural University , Guangzhou , China.,b Guangdong Provincial Key Laboratory of Agro-animal Genomics and Molecular Breeding, College of Animal Science , South China Agricultural University , Guangzhou , China
| | - Gelong Jiang
- a National Engineering Research Center for Breeding Swine Industry, College of Animal Science , South China Agricultural University , Guangzhou , China.,b Guangdong Provincial Key Laboratory of Agro-animal Genomics and Molecular Breeding, College of Animal Science , South China Agricultural University , Guangzhou , China
| | - Dewu Liu
- a National Engineering Research Center for Breeding Swine Industry, College of Animal Science , South China Agricultural University , Guangzhou , China.,b Guangdong Provincial Key Laboratory of Agro-animal Genomics and Molecular Breeding, College of Animal Science , South China Agricultural University , Guangzhou , China
| | - Johann Urschitz
- d Institute for Biogenesis Research, Department of Anatomy, Biochemistry and Physiology, John A. Burns School of Medicine , University of Hawaii at Manoa , Honolulu , Hawaii , USA
| | - Stefan Moisyadi
- d Institute for Biogenesis Research, Department of Anatomy, Biochemistry and Physiology, John A. Burns School of Medicine , University of Hawaii at Manoa , Honolulu , Hawaii , USA.,e Manoa BioSciences , Honolulu , Hawaii , USA
| | - Zhenfang Wu
- a National Engineering Research Center for Breeding Swine Industry, College of Animal Science , South China Agricultural University , Guangzhou , China.,b Guangdong Provincial Key Laboratory of Agro-animal Genomics and Molecular Breeding, College of Animal Science , South China Agricultural University , Guangzhou , China
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7
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Mukherjee A, Garrels W, Talluri TR, Tiedemann D, Bősze Z, Ivics Z, Kues WA. Expression of Active Fluorophore Proteins in the Milk of Transgenic Pigs Bypassing the Secretory Pathway. Sci Rep 2016; 6:24464. [PMID: 27086548 PMCID: PMC4834472 DOI: 10.1038/srep24464] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/26/2015] [Accepted: 03/30/2016] [Indexed: 12/12/2022] Open
Abstract
We describe the expression of recombinant fluorescent proteins in the milk of two lines of transgenic pigs generated by Sleeping Beauty transposon-mediated genetic engineering. The Sleeping Beauty transposon consisted of an ubiquitously active CAGGS promoter driving a fluorophore cDNA, encoding either Venus or mCherry. Importantly, the fluorophore cDNAs did not encode for a signal peptide for the secretory pathway, and in previous studies of the transgenic animals a cytoplasmic localization of the fluorophore proteins was found. Unexpectedly, milk samples from lactating sows contained high levels of bioactive Venus or mCherry fluorophores. A detailed analysis suggested that exfoliated cells of the mammary epithelium carried the recombinant proteins passively into the milk. This is the first description of reporter fluorophore expression in the milk of livestock, and the findings may contribute to the development of an alternative concept for the production of bioactive recombinant proteins in the udder.
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Affiliation(s)
- Ayan Mukherjee
- Friedrich-Loeffler-Institut, Institut für Nutztiergenetik, Mariensee, Germany
| | - Wiebke Garrels
- Medical School Hannover, Institute of Laboratory Animal Sciences, Hannover, Germany
| | | | - Daniela Tiedemann
- Friedrich-Loeffler-Institut, Institut für Nutztiergenetik, Mariensee, Germany
| | - Zsuzsanna Bősze
- NARIC- Agricultural Biotechnology Institute, Gödöllö, Hungary
| | | | - Wilfried A. Kues
- Friedrich-Loeffler-Institut, Institut für Nutztiergenetik, Mariensee, Germany
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8
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Garrels W, Mukherjee A, Holler S, Cleve N, Talluri TR, Barg-Kues B, Diederich M, Köhler P, Petersen B, Lucas-Hahn A, Niemann H, Izsvák Z, Ivics Z, Kues WA. Identification and re-addressing of a transcriptionally permissive locus in the porcine genome. Transgenic Res 2015; 25:63-70. [DOI: 10.1007/s11248-015-9914-4] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/17/2015] [Accepted: 10/23/2015] [Indexed: 01/21/2023]
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9
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Talluri TR, Kumar D, Glage S, Garrels W, Ivics Z, Debowski K, Behr R, Niemann H, Kues WA. Derivation and characterization of bovine induced pluripotent stem cells by transposon-mediated reprogramming. Cell Reprogram 2015; 17:131-40. [PMID: 25826726 DOI: 10.1089/cell.2014.0080] [Citation(s) in RCA: 56] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
Induced pluripotent stem cells (iPSCs) are a seminal breakthrough in stem cell research and are promising tools for advanced regenerative therapies in humans and reproductive biotechnology in farm animals. iPSCs are particularly valuable in species in which authentic embryonic stem cell (ESC) lines are yet not available. Here, we describe a nonviral method for the derivation of bovine iPSCs employing Sleeping Beauty (SB) and piggyBac (PB) transposon systems encoding different combinations of reprogramming factors, each separated by self-cleaving peptide sequences and driven by the chimeric CAGGS promoter. One bovine iPSC line (biPS-1) generated by a PB vector containing six reprogramming genes was analyzed in detail, including morphology, alkaline phosphatase expression, and typical hallmarks of pluripotency, such as expression of pluripotency markers and formation of mature teratomas in immunodeficient mice. Moreover, the biPS-1 line allowed a second round of SB transposon-mediated gene transfer. These results are promising for derivation of germ line-competent bovine iPSCs and will facilitate genetic modification of the bovine genome.
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Affiliation(s)
- Thirumala R Talluri
- 1 Institut für Nutztiergenetik, Friedrich-Loeffler-Institut , Mariensee, 31535 Neustadt, Germany
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10
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Abstract
The first transgenic pigs were generated for agricultural purposes about three decades ago. Since then, the micromanipulation techniques of pig oocytes and embryos expanded from pronuclear injection of foreign DNA to somatic cell nuclear transfer, intracytoplasmic sperm injection-mediated gene transfer, lentiviral transduction, and cytoplasmic injection. Mechanistically, the passive transgenesis approach based on random integration of foreign DNA was developed to active genetic engineering techniques based on the transient activity of ectopic enzymes, such as transposases, recombinases, and programmable nucleases. Whole-genome sequencing and annotation of advanced genome maps of the pig complemented these developments. The full implementation of these tools promises to immensely increase the efficiency and, in parallel, to reduce the costs for the generation of genetically engineered pigs. Today, the major application of genetically engineered pigs is found in the field of biomedical disease modeling. It is anticipated that genetically engineered pigs will increasingly be used in biomedical research, since this model shows several similarities to humans with regard to physiology, metabolism, genome organization, pathology, and aging.
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Affiliation(s)
- Gökhan Gün
- Department of Biotechnology, Friedrich-Loeffler-Institut, Institut für Nutztiergenetik, Mariensee, Neustadt, Germany
- Molecular Biology & Genetics, Istanbul Technical University, Istanbul, Turkey
- Histology and Embryology Department, Faculty of Veterinary Medicine, Istanbul University, Istanbul, Turkey
| | - Wilfried A. Kues
- Department of Biotechnology, Friedrich-Loeffler-Institut, Institut für Nutztiergenetik, Mariensee, Neustadt, Germany
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11
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Garrels W, Holler S, Taylor U, Herrmann D, Niemann H, Ivics Z, Kues WA. Assessment of fetal cell chimerism in transgenic pig lines generated by Sleeping beauty transposition. PLoS One 2014; 9:e96673. [PMID: 24811124 PMCID: PMC4014516 DOI: 10.1371/journal.pone.0096673] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/14/2013] [Accepted: 04/10/2014] [Indexed: 12/22/2022] Open
Abstract
Human cells migrate between mother and fetus during pregnancy and persist in the respective host for long-term after birth. Fetal microchimerism occurs also in twins sharing a common placenta or chorion. Whether microchimerism occurs in multiparous mammals such as the domestic pig, where fetuses have separate placentas and chorions, is not well understood. Here, we assessed cell chimerism in litters of wild-type sows inseminated with semen of transposon transgenic boars. Segregation of three independent monomeric transposons ensured an excess of transgenic over non-transgenic offspring in every litter. Transgenic siblings (n = 35) showed robust ubiquitous expression of the reporter transposon encoding a fluorescent protein, and provided an unique resource to assess a potential cell trafficking to non-transgenic littermates (n = 7) or mothers (n = 4). Sensitive flow cytometry, fluorescence microscopy, and real-time PCR provided no evidence for microchimerism in porcine littermates, or piglets and their mothers in both blood and solid organs. These data indicate that the epitheliochorial structure of the porcine placenta effectively prevents cellular exchange during gestation.
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Affiliation(s)
- Wiebke Garrels
- Institut für Nutztiergenetik, Friedrich-Loeffler-Institut, Mariensee, Germany
| | - Stephanie Holler
- Institut für Nutztiergenetik, Friedrich-Loeffler-Institut, Mariensee, Germany
| | - Ulrike Taylor
- Institut für Nutztiergenetik, Friedrich-Loeffler-Institut, Mariensee, Germany
| | - Doris Herrmann
- Institut für Nutztiergenetik, Friedrich-Loeffler-Institut, Mariensee, Germany
| | - Heiner Niemann
- Institut für Nutztiergenetik, Friedrich-Loeffler-Institut, Mariensee, Germany
| | | | - Wilfried A Kues
- Institut für Nutztiergenetik, Friedrich-Loeffler-Institut, Mariensee, Germany
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