Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro

Autores
Mellisho, Edwin A.; Velásquez, Alejandra E.; Nuñez, María J.; Cabezas, Joel G.; Cueto, Juan Agustin; Fader Kaiser, Claudio Marcelo; Castro, Fidel O.; Rodriguez Alvarez, Lleretny
Año de publicación
2017
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Extracellular vesicles (EVs) have been identified within different body fluids and cell culture media. However, there is very little information on the secretion of these vesicles during early embryonic development. The aims of this work were first to demonstrate the secretion of extracellular vesicles by pre-implantation bovine embryos and second to identify and characterize the population of EVs secreted by bovine blastocysts during the period from day seven to nine of embryo culture and its correlation with further embryo development up to day 11. Bovine embryos were produced by in vitro fertilization (IVF) or parthenogenetic activation (PA) and cultured until blastocyst stage. Blastocyst selection was performed at day 7 post IVF/PA considering two variables: stage of development and quality of embryos. Selected blastocysts were cultured in vitro for 48 hours in groups (exp. 1) or individually (exp. 2) in SOF media depleted of exosomes. At day 9 post IVF/PA the media was collected and EVs isolated by ultracentrifugation. Transmission electron microscopy revealed the presence of heterogeneous vesicles of different sizes and population: microvesicles (MVs) and exosomes (EXs) of rounded shape, enclosed by a lipid bi-layer and ranging from 30 to 385 nm of diameter. Flow cytometry analysis allowed identifying CD63 and CD9 proteins as exosome markers. Nanoparticle tracking analysis generated a large number of variables, which required the use of multivariate statistics. The results indicated that the concentration of vesicles is higher in those blastocysts with arrested development from day 9 up to day 11 of in vitro development (6.7 x 108 particles/ml) derived from IVF (p <0.05), compared to PA blastocysts (4.7 x 108 particles/ml). Likewise, the profile (concentration and diameter) of particles secreted by embryos derived from IVF were different from those secreted by PA embryos. In conclusion, we demonstrated that bovine blastocysts secrete MVs/EXs to the culture media. Data suggest that characteristics of the population of EVs vary depending on embryo competence.
Fil: Mellisho, Edwin A.. Universidad de Concepción; Chile
Fil: Velásquez, Alejandra E.. Universidad de Concepción; Chile
Fil: Nuñez, María J.. Universidad de Concepción; Chile
Fil: Cabezas, Joel G.. Universidad de Concepción; Chile
Fil: Cueto, Juan Agustin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentina
Fil: Fader Kaiser, Claudio Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentina
Fil: Castro, Fidel O.. Universidad de Concepción; Chile
Fil: Rodriguez Alvarez, Lleretny. Universidad de Concepción; Chile
Materia
EXOSOMES
MVBs
ELECTRON MICROSCOPY
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/49431
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/49431
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitroMellisho, Edwin A.Velásquez, Alejandra E.Nuñez, María J.Cabezas, Joel G.Cueto, Juan AgustinFader Kaiser, Claudio MarceloCastro, Fidel O.Rodriguez Alvarez, LleretnyEXOSOMESMVBsELECTRON MICROSCOPYhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Extracellular vesicles (EVs) have been identified within different body fluids and cell culture media. However, there is very little information on the secretion of these vesicles during early embryonic development. The aims of this work were first to demonstrate the secretion of extracellular vesicles by pre-implantation bovine embryos and second to identify and characterize the population of EVs secreted by bovine blastocysts during the period from day seven to nine of embryo culture and its correlation with further embryo development up to day 11. Bovine embryos were produced by in vitro fertilization (IVF) or parthenogenetic activation (PA) and cultured until blastocyst stage. Blastocyst selection was performed at day 7 post IVF/PA considering two variables: stage of development and quality of embryos. Selected blastocysts were cultured in vitro for 48 hours in groups (exp. 1) or individually (exp. 2) in SOF media depleted of exosomes. At day 9 post IVF/PA the media was collected and EVs isolated by ultracentrifugation. Transmission electron microscopy revealed the presence of heterogeneous vesicles of different sizes and population: microvesicles (MVs) and exosomes (EXs) of rounded shape, enclosed by a lipid bi-layer and ranging from 30 to 385 nm of diameter. Flow cytometry analysis allowed identifying CD63 and CD9 proteins as exosome markers. Nanoparticle tracking analysis generated a large number of variables, which required the use of multivariate statistics. The results indicated that the concentration of vesicles is higher in those blastocysts with arrested development from day 9 up to day 11 of in vitro development (6.7 x 108 particles/ml) derived from IVF (p <0.05), compared to PA blastocysts (4.7 x 108 particles/ml). Likewise, the profile (concentration and diameter) of particles secreted by embryos derived from IVF were different from those secreted by PA embryos. In conclusion, we demonstrated that bovine blastocysts secrete MVs/EXs to the culture media. Data suggest that characteristics of the population of EVs vary depending on embryo competence.Fil: Mellisho, Edwin A.. Universidad de Concepción; ChileFil: Velásquez, Alejandra E.. Universidad de Concepción; ChileFil: Nuñez, María J.. Universidad de Concepción; ChileFil: Cabezas, Joel G.. Universidad de Concepción; ChileFil: Cueto, Juan Agustin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Fader Kaiser, Claudio Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Castro, Fidel O.. Universidad de Concepción; ChileFil: Rodriguez Alvarez, Lleretny. Universidad de Concepción; ChilePublic Library of Science2017-05info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/49431Mellisho, Edwin A.; Velásquez, Alejandra E.; Nuñez, María J.; Cabezas, Joel G.; Cueto, Juan Agustin; et al.; Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro; Public Library of Science; Plos One; 12; 5; 5-2017; 1-151932-6203CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1371/journal.pone.0178306info:eu-repo/semantics/altIdentifier/url/http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0178306info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-10-22T11:58:37Zoai:ri.conicet.gov.ar:11336/49431instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-10-22 11:58:37.948CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro
title Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro
spellingShingle Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro
Mellisho, Edwin A.
EXOSOMES
MVBs
ELECTRON MICROSCOPY
title_short Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro
title_full Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro
title_fullStr Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro
title_full_unstemmed Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro
title_sort Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro
dc.creator.none.fl_str_mv Mellisho, Edwin A.
Velásquez, Alejandra E.
Nuñez, María J.
Cabezas, Joel G.
Cueto, Juan Agustin
Fader Kaiser, Claudio Marcelo
Castro, Fidel O.
Rodriguez Alvarez, Lleretny
author Mellisho, Edwin A.
author_facet Mellisho, Edwin A.
Velásquez, Alejandra E.
Nuñez, María J.
Cabezas, Joel G.
Cueto, Juan Agustin
Fader Kaiser, Claudio Marcelo
Castro, Fidel O.
Rodriguez Alvarez, Lleretny
author_role author
author2 Velásquez, Alejandra E.
Nuñez, María J.
Cabezas, Joel G.
Cueto, Juan Agustin
Fader Kaiser, Claudio Marcelo
Castro, Fidel O.
Rodriguez Alvarez, Lleretny
author2_role author
author
author
author
author
author
author
dc.subject.none.fl_str_mv EXOSOMES
MVBs
ELECTRON MICROSCOPY
topic EXOSOMES
MVBs
ELECTRON MICROSCOPY
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Extracellular vesicles (EVs) have been identified within different body fluids and cell culture media. However, there is very little information on the secretion of these vesicles during early embryonic development. The aims of this work were first to demonstrate the secretion of extracellular vesicles by pre-implantation bovine embryos and second to identify and characterize the population of EVs secreted by bovine blastocysts during the period from day seven to nine of embryo culture and its correlation with further embryo development up to day 11. Bovine embryos were produced by in vitro fertilization (IVF) or parthenogenetic activation (PA) and cultured until blastocyst stage. Blastocyst selection was performed at day 7 post IVF/PA considering two variables: stage of development and quality of embryos. Selected blastocysts were cultured in vitro for 48 hours in groups (exp. 1) or individually (exp. 2) in SOF media depleted of exosomes. At day 9 post IVF/PA the media was collected and EVs isolated by ultracentrifugation. Transmission electron microscopy revealed the presence of heterogeneous vesicles of different sizes and population: microvesicles (MVs) and exosomes (EXs) of rounded shape, enclosed by a lipid bi-layer and ranging from 30 to 385 nm of diameter. Flow cytometry analysis allowed identifying CD63 and CD9 proteins as exosome markers. Nanoparticle tracking analysis generated a large number of variables, which required the use of multivariate statistics. The results indicated that the concentration of vesicles is higher in those blastocysts with arrested development from day 9 up to day 11 of in vitro development (6.7 x 108 particles/ml) derived from IVF (p <0.05), compared to PA blastocysts (4.7 x 108 particles/ml). Likewise, the profile (concentration and diameter) of particles secreted by embryos derived from IVF were different from those secreted by PA embryos. In conclusion, we demonstrated that bovine blastocysts secrete MVs/EXs to the culture media. Data suggest that characteristics of the population of EVs vary depending on embryo competence.
Fil: Mellisho, Edwin A.. Universidad de Concepción; Chile
Fil: Velásquez, Alejandra E.. Universidad de Concepción; Chile
Fil: Nuñez, María J.. Universidad de Concepción; Chile
Fil: Cabezas, Joel G.. Universidad de Concepción; Chile
Fil: Cueto, Juan Agustin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentina
Fil: Fader Kaiser, Claudio Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentina
Fil: Castro, Fidel O.. Universidad de Concepción; Chile
Fil: Rodriguez Alvarez, Lleretny. Universidad de Concepción; Chile
description Extracellular vesicles (EVs) have been identified within different body fluids and cell culture media. However, there is very little information on the secretion of these vesicles during early embryonic development. The aims of this work were first to demonstrate the secretion of extracellular vesicles by pre-implantation bovine embryos and second to identify and characterize the population of EVs secreted by bovine blastocysts during the period from day seven to nine of embryo culture and its correlation with further embryo development up to day 11. Bovine embryos were produced by in vitro fertilization (IVF) or parthenogenetic activation (PA) and cultured until blastocyst stage. Blastocyst selection was performed at day 7 post IVF/PA considering two variables: stage of development and quality of embryos. Selected blastocysts were cultured in vitro for 48 hours in groups (exp. 1) or individually (exp. 2) in SOF media depleted of exosomes. At day 9 post IVF/PA the media was collected and EVs isolated by ultracentrifugation. Transmission electron microscopy revealed the presence of heterogeneous vesicles of different sizes and population: microvesicles (MVs) and exosomes (EXs) of rounded shape, enclosed by a lipid bi-layer and ranging from 30 to 385 nm of diameter. Flow cytometry analysis allowed identifying CD63 and CD9 proteins as exosome markers. Nanoparticle tracking analysis generated a large number of variables, which required the use of multivariate statistics. The results indicated that the concentration of vesicles is higher in those blastocysts with arrested development from day 9 up to day 11 of in vitro development (6.7 x 108 particles/ml) derived from IVF (p <0.05), compared to PA blastocysts (4.7 x 108 particles/ml). Likewise, the profile (concentration and diameter) of particles secreted by embryos derived from IVF were different from those secreted by PA embryos. In conclusion, we demonstrated that bovine blastocysts secrete MVs/EXs to the culture media. Data suggest that characteristics of the population of EVs vary depending on embryo competence.
publishDate 2017
dc.date.none.fl_str_mv 2017-05
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/49431
Mellisho, Edwin A.; Velásquez, Alejandra E.; Nuñez, María J.; Cabezas, Joel G.; Cueto, Juan Agustin; et al.; Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro; Public Library of Science; Plos One; 12; 5; 5-2017; 1-15
1932-6203
CONICET Digital
CONICET
url http://hdl.handle.net/11336/49431
identifier_str_mv Mellisho, Edwin A.; Velásquez, Alejandra E.; Nuñez, María J.; Cabezas, Joel G.; Cueto, Juan Agustin; et al.; Identification and characteristics of Extracellular vesicles from bovine blastocysts produced in vitro; Public Library of Science; Plos One; 12; 5; 5-2017; 1-15
1932-6203
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1371/journal.pone.0178306
info:eu-repo/semantics/altIdentifier/url/http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0178306
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Public Library of Science
publisher.none.fl_str_mv Public Library of Science
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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