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dc.contributor.author
Locatelli, Paola
dc.contributor.author
Olea, Fernanda Daniela
dc.contributor.author
Hnatiuk, Anna
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Sepúlveda, Diana Elizabeth
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Pérez Sáez, Juan Manuel
dc.contributor.author
Argüello, Rafael
dc.contributor.author
Crottogini, Alberto Jose
dc.date.available
2017-08-23T22:09:11Z
dc.date.issued
2013-01
dc.identifier.citation
Locatelli, Paola; Olea, Fernanda Daniela; Hnatiuk, Anna; Sepúlveda, Diana Elizabeth; Pérez Sáez, Juan Manuel; et al.; Efficient plasmid-mediated gene transfection of ovine bone marrow mesenchymal stromal cells; Elsevier; Cytotherapy; 15; 2; 1-2013; 163-170
dc.identifier.issn
1465-3249
dc.identifier.uri
http://hdl.handle.net/11336/22913
dc.description.abstract
BACKGROUND AIMS: Given the close similarity between ovine and human cardiomyocytes, sheep models of myocardial infarction and heart failure are increasingly used in studies of stem cell-mediated heart regeneration. In these studies, mesenchymal stromal cells (MSCs) are frequently employed. To enhance the paracrine effects of these MSCs, ex vivo transfection with genes encoding growth factors has been proposed. Although viral vectors exhibit higher transfection efficiency than plasmids, they entail the risks of uncontrolled transgene expression and immune reactions that preclude repeated administration. Our aim was to optimize the efficiency of plasmid-mediated transfection of ovine MSCs, while preserving cell viability. METHODS: Varying amounts of diverse cationic lipids were used to obtain the reagent-to-DNA mass ratio showing highest luciferase activity. Transfection efficiency (flow cytometry) was tested on plasmid-green fluorescent protein-transfected MSCs at increasing DNA mass. RESULTS: Lipofectamine LTX 5 μL and Plus reagent 4 μL with 2 μg of DNA yielded 42.3 ± 4.7% transfection efficiency, while preserving cell viability. Using these transfection conditions, we transfected MSCs with a plasmid encoding human vascular endothelial growth factor (VEGF) and found high VEGF protein concentrations in the culture supernatant from day 2 (1968 ± 324 pg/mL per μg DNA) through at least day 12 (888 ± 386 pg/mL per μg DNA) after transfection. CONCLUSIONS: Plasmid-mediated transfection of ovine MSCs to over-express paracrine heart-regenerative growth factors is feasible and efficient and overcomes the risks and limitations associated with the use of viral vectors.
dc.format
application/pdf
dc.language.iso
eng
dc.publisher
Elsevier
dc.rights
info:eu-repo/semantics/openAccess
dc.rights.uri
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.subject
Mesenchymal Stromal Cell
dc.subject
Plasmids
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Transfection Efficiency
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Sheep
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Vegf
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Bioquímica y Biología Molecular
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Ciencias Biológicas
dc.subject.classification
CIENCIAS NATURALES Y EXACTAS
dc.title
Efficient plasmid-mediated gene transfection of ovine bone marrow mesenchymal stromal cells
dc.type
info:eu-repo/semantics/article
dc.type
info:ar-repo/semantics/artículo
dc.type
info:eu-repo/semantics/publishedVersion
dc.date.updated
2017-08-22T21:24:29Z
dc.journal.volume
15
dc.journal.number
2
dc.journal.pagination
163-170
dc.journal.pais
Países Bajos
dc.journal.ciudad
Ámsterdam
dc.description.fil
Fil: Locatelli, Paola. Universidad Favaloro; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
dc.description.fil
Fil: Olea, Fernanda Daniela. Universidad Favaloro; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
dc.description.fil
Fil: Hnatiuk, Anna. Universidad Favaloro; Argentina
dc.description.fil
Fil: Sepúlveda, Diana Elizabeth. Universidad Favaloro; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
dc.description.fil
Fil: Pérez Sáez, Juan Manuel. Bio Sidus S.A; Argentina
dc.description.fil
Fil: Argüello, Rafael. Universidad Favaloro; Argentina
dc.description.fil
Fil: Crottogini, Alberto Jose. Universidad Favaloro; Argentina
dc.journal.title
Cytotherapy
dc.relation.alternativeid
info:eu-repo/semantics/altIdentifier/url/http://www.sciencedirect.com/science/article/pii/S1465324912000333
dc.relation.alternativeid
info:eu-repo/semantics/altIdentifier/doi/http://dx.doi.org/10.1016/j.jcyt.2012.11.004
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