New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines
- Autores
- Laiz Quiroga, Lucía Belén; Gimeno, Brenda Florencia; Martínez León, Eduardo Antonio; Von Meyeren, Micaela; Mutto, Adrián Angel; Bariani, Maria Victoria; Osycka Salut, Claudia Elena
- Año de publicación
- 2021
- Idioma
- inglés
- Tipo de recurso
- documento de conferencia
- Estado
- versión publicada
- Descripción
- Cryopreserved sperm (SPZ) are used widely in domestic animals for assisted reproduction techniques (ART). However, cryopreservation procedures negatively affect SPZ quality, causing changes at the structural and molecular levels. Therefore, using cryopreserved SPZ for ART can decrease the efficiency of these techniques as well as the quality of the embryos obtained, being necessary to optimize the handling of this type of sample. Particularly in equines, ICSI is the most used technique for low-quality equine semen samples where SPZ is selected based on their motility and morphology. Physiologically, the oviduct epithelial cells (OEC) are involved in the selection of a population of SPZ suitable for oocyte fertilization, which is released from the oviduct due to the capacitation process. This work aimed to evaluate different conditions for the in vitro manipulation and incubation of equine cryopreserved SPZ and to establish an in vitro OEC culture to select an SPZ population with fertilizing capacity suitable for its use in ART in this species.Regarding the handling and the effect on the motility of post-thawed equine cryopreserved SPZ samples, we used non-capacitating Whitten´s medium, less effect on motility was observed through the time (120 min) when SPZ were incubated at concentrations of 30 mill/ml compared to lower concentrated samples (CASA, p<0.05). Also, the motility decreased by half with centrifugations at 200g longer than 1 min (p<0.05). On the other hand, we have established a culture model of OECs in vitro, in which we demonstrated the expression of epithelial markers (E-cadherin and cytokeratin) by both RT-PCR and immunofluorescence (IF). When we performed SPZ-OEC cocultures, we observed that the attached SPZs were motile and presented intact acrosome (PSA-FITC, IF), suggesting a selection by the oviductal model. Then, the co-cultures were incubated in capacitating conditions (capacitating Whitten´s medium) and the released SPZ population was recovered. Under these conditions, a greater number of live sperm (Hoechst258 assessed by IF), capacitated (activation of PKA and phosphorylation on tyrosine residues by IF), with progressive motility (CASA) and with the intact acrosome (PSA-FITC, IF) compared to the control was observed (p<0.05). Moreover, the decrease in motility through the time was less in these SPZ than in the cryopreserved SPZ incubated in the absence of OECs. This sperm population could be recovered for its use in different ART.Improvements in handling and selection of cryopreserved SPZ not only generate tools to solve the problem that IVF presents in equines, but would also improve efficiency in other ART such as ICSI, allowing the use of a population of higher quality gametes which it would positively impact the quality of the embryos obtained.
Fil: Laiz Quiroga, Lucía Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Gimeno, Brenda Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; Argentina
Fil: Martínez León, Eduardo Antonio. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; Argentina
Fil: Von Meyeren, Micaela. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Mutto, Adrián Angel. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Bariani, Maria Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; Argentina
Fil: Osycka Salut, Claudia Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; Argentina
IV Joint Meeting of The Biology Societies of Argentina
Mendoza
Argentina
Sociedad de Biología de Cuyo
Sociedad Argentina de Biología
Sociedad de Biología de Córdoba
Sociedad de Biología de Rosario
Asociación de Biología de Tucumán - Materia
-
CRYOPRESERVED SPERM
HANDLING
OVIDUCT SELECTION
EQUINES - Nivel de accesibilidad
- acceso abierto
- Condiciones de uso
- https://creativecommons.org/licenses/by/2.5/ar/
- Repositorio
- Institución
- Consejo Nacional de Investigaciones Científicas y Técnicas
- OAI Identificador
- oai:ri.conicet.gov.ar:11336/153951
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New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in EquinesLaiz Quiroga, Lucía BelénGimeno, Brenda FlorenciaMartínez León, Eduardo AntonioVon Meyeren, MicaelaMutto, Adrián AngelBariani, Maria VictoriaOsycka Salut, Claudia ElenaCRYOPRESERVED SPERMHANDLINGOVIDUCT SELECTIONEQUINEShttps://purl.org/becyt/ford/4.3https://purl.org/becyt/ford/4Cryopreserved sperm (SPZ) are used widely in domestic animals for assisted reproduction techniques (ART). However, cryopreservation procedures negatively affect SPZ quality, causing changes at the structural and molecular levels. Therefore, using cryopreserved SPZ for ART can decrease the efficiency of these techniques as well as the quality of the embryos obtained, being necessary to optimize the handling of this type of sample. Particularly in equines, ICSI is the most used technique for low-quality equine semen samples where SPZ is selected based on their motility and morphology. Physiologically, the oviduct epithelial cells (OEC) are involved in the selection of a population of SPZ suitable for oocyte fertilization, which is released from the oviduct due to the capacitation process. This work aimed to evaluate different conditions for the in vitro manipulation and incubation of equine cryopreserved SPZ and to establish an in vitro OEC culture to select an SPZ population with fertilizing capacity suitable for its use in ART in this species.Regarding the handling and the effect on the motility of post-thawed equine cryopreserved SPZ samples, we used non-capacitating Whitten´s medium, less effect on motility was observed through the time (120 min) when SPZ were incubated at concentrations of 30 mill/ml compared to lower concentrated samples (CASA, p<0.05). Also, the motility decreased by half with centrifugations at 200g longer than 1 min (p<0.05). On the other hand, we have established a culture model of OECs in vitro, in which we demonstrated the expression of epithelial markers (E-cadherin and cytokeratin) by both RT-PCR and immunofluorescence (IF). When we performed SPZ-OEC cocultures, we observed that the attached SPZs were motile and presented intact acrosome (PSA-FITC, IF), suggesting a selection by the oviductal model. Then, the co-cultures were incubated in capacitating conditions (capacitating Whitten´s medium) and the released SPZ population was recovered. Under these conditions, a greater number of live sperm (Hoechst258 assessed by IF), capacitated (activation of PKA and phosphorylation on tyrosine residues by IF), with progressive motility (CASA) and with the intact acrosome (PSA-FITC, IF) compared to the control was observed (p<0.05). Moreover, the decrease in motility through the time was less in these SPZ than in the cryopreserved SPZ incubated in the absence of OECs. This sperm population could be recovered for its use in different ART.Improvements in handling and selection of cryopreserved SPZ not only generate tools to solve the problem that IVF presents in equines, but would also improve efficiency in other ART such as ICSI, allowing the use of a population of higher quality gametes which it would positively impact the quality of the embryos obtained.Fil: Laiz Quiroga, Lucía Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Gimeno, Brenda Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Martínez León, Eduardo Antonio. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Von Meyeren, Micaela. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Mutto, Adrián Angel. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Bariani, Maria Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaFil: Osycka Salut, Claudia Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaIV Joint Meeting of The Biology Societies of ArgentinaMendozaArgentinaSociedad de Biología de CuyoSociedad Argentina de BiologíaSociedad de Biología de CórdobaSociedad de Biología de RosarioAsociación de Biología de TucumánTech Science Press2021info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/conferenceObjectReuniónJournalhttp://purl.org/coar/resource_type/c_5794info:ar-repo/semantics/documentoDeConferenciaapplication/pdfapplication/vnd.openxmlformats-officedocument.wordprocessingml.documentapplication/pdfhttp://hdl.handle.net/11336/153951New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines; IV Joint Meeting of The Biology Societies of Argentina; Mendoza; Argentina; 2020; 47-480327-95451667-5746CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.techscience.com/biocell/v45nSuppl.3/44000Internacionalinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:36:23Zoai:ri.conicet.gov.ar:11336/153951instacron: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:34982025-09-29 09:36:23.325CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse |
dc.title.none.fl_str_mv |
New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines |
title |
New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines |
spellingShingle |
New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines Laiz Quiroga, Lucía Belén CRYOPRESERVED SPERM HANDLING OVIDUCT SELECTION EQUINES |
title_short |
New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines |
title_full |
New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines |
title_fullStr |
New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines |
title_full_unstemmed |
New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines |
title_sort |
New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines |
dc.creator.none.fl_str_mv |
Laiz Quiroga, Lucía Belén Gimeno, Brenda Florencia Martínez León, Eduardo Antonio Von Meyeren, Micaela Mutto, Adrián Angel Bariani, Maria Victoria Osycka Salut, Claudia Elena |
author |
Laiz Quiroga, Lucía Belén |
author_facet |
Laiz Quiroga, Lucía Belén Gimeno, Brenda Florencia Martínez León, Eduardo Antonio Von Meyeren, Micaela Mutto, Adrián Angel Bariani, Maria Victoria Osycka Salut, Claudia Elena |
author_role |
author |
author2 |
Gimeno, Brenda Florencia Martínez León, Eduardo Antonio Von Meyeren, Micaela Mutto, Adrián Angel Bariani, Maria Victoria Osycka Salut, Claudia Elena |
author2_role |
author author author author author author |
dc.subject.none.fl_str_mv |
CRYOPRESERVED SPERM HANDLING OVIDUCT SELECTION EQUINES |
topic |
CRYOPRESERVED SPERM HANDLING OVIDUCT SELECTION EQUINES |
purl_subject.fl_str_mv |
https://purl.org/becyt/ford/4.3 https://purl.org/becyt/ford/4 |
dc.description.none.fl_txt_mv |
Cryopreserved sperm (SPZ) are used widely in domestic animals for assisted reproduction techniques (ART). However, cryopreservation procedures negatively affect SPZ quality, causing changes at the structural and molecular levels. Therefore, using cryopreserved SPZ for ART can decrease the efficiency of these techniques as well as the quality of the embryos obtained, being necessary to optimize the handling of this type of sample. Particularly in equines, ICSI is the most used technique for low-quality equine semen samples where SPZ is selected based on their motility and morphology. Physiologically, the oviduct epithelial cells (OEC) are involved in the selection of a population of SPZ suitable for oocyte fertilization, which is released from the oviduct due to the capacitation process. This work aimed to evaluate different conditions for the in vitro manipulation and incubation of equine cryopreserved SPZ and to establish an in vitro OEC culture to select an SPZ population with fertilizing capacity suitable for its use in ART in this species.Regarding the handling and the effect on the motility of post-thawed equine cryopreserved SPZ samples, we used non-capacitating Whitten´s medium, less effect on motility was observed through the time (120 min) when SPZ were incubated at concentrations of 30 mill/ml compared to lower concentrated samples (CASA, p<0.05). Also, the motility decreased by half with centrifugations at 200g longer than 1 min (p<0.05). On the other hand, we have established a culture model of OECs in vitro, in which we demonstrated the expression of epithelial markers (E-cadherin and cytokeratin) by both RT-PCR and immunofluorescence (IF). When we performed SPZ-OEC cocultures, we observed that the attached SPZs were motile and presented intact acrosome (PSA-FITC, IF), suggesting a selection by the oviductal model. Then, the co-cultures were incubated in capacitating conditions (capacitating Whitten´s medium) and the released SPZ population was recovered. Under these conditions, a greater number of live sperm (Hoechst258 assessed by IF), capacitated (activation of PKA and phosphorylation on tyrosine residues by IF), with progressive motility (CASA) and with the intact acrosome (PSA-FITC, IF) compared to the control was observed (p<0.05). Moreover, the decrease in motility through the time was less in these SPZ than in the cryopreserved SPZ incubated in the absence of OECs. This sperm population could be recovered for its use in different ART.Improvements in handling and selection of cryopreserved SPZ not only generate tools to solve the problem that IVF presents in equines, but would also improve efficiency in other ART such as ICSI, allowing the use of a population of higher quality gametes which it would positively impact the quality of the embryos obtained. Fil: Laiz Quiroga, Lucía Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; Argentina Fil: Gimeno, Brenda Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; Argentina Fil: Martínez León, Eduardo Antonio. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; Argentina Fil: Von Meyeren, Micaela. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina Fil: Mutto, Adrián Angel. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina Fil: Bariani, Maria Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; Argentina Fil: Osycka Salut, Claudia Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; Argentina IV Joint Meeting of The Biology Societies of Argentina Mendoza Argentina Sociedad de Biología de Cuyo Sociedad Argentina de Biología Sociedad de Biología de Córdoba Sociedad de Biología de Rosario Asociación de Biología de Tucumán |
description |
Cryopreserved sperm (SPZ) are used widely in domestic animals for assisted reproduction techniques (ART). However, cryopreservation procedures negatively affect SPZ quality, causing changes at the structural and molecular levels. Therefore, using cryopreserved SPZ for ART can decrease the efficiency of these techniques as well as the quality of the embryos obtained, being necessary to optimize the handling of this type of sample. Particularly in equines, ICSI is the most used technique for low-quality equine semen samples where SPZ is selected based on their motility and morphology. Physiologically, the oviduct epithelial cells (OEC) are involved in the selection of a population of SPZ suitable for oocyte fertilization, which is released from the oviduct due to the capacitation process. This work aimed to evaluate different conditions for the in vitro manipulation and incubation of equine cryopreserved SPZ and to establish an in vitro OEC culture to select an SPZ population with fertilizing capacity suitable for its use in ART in this species.Regarding the handling and the effect on the motility of post-thawed equine cryopreserved SPZ samples, we used non-capacitating Whitten´s medium, less effect on motility was observed through the time (120 min) when SPZ were incubated at concentrations of 30 mill/ml compared to lower concentrated samples (CASA, p<0.05). Also, the motility decreased by half with centrifugations at 200g longer than 1 min (p<0.05). On the other hand, we have established a culture model of OECs in vitro, in which we demonstrated the expression of epithelial markers (E-cadherin and cytokeratin) by both RT-PCR and immunofluorescence (IF). When we performed SPZ-OEC cocultures, we observed that the attached SPZs were motile and presented intact acrosome (PSA-FITC, IF), suggesting a selection by the oviductal model. Then, the co-cultures were incubated in capacitating conditions (capacitating Whitten´s medium) and the released SPZ population was recovered. Under these conditions, a greater number of live sperm (Hoechst258 assessed by IF), capacitated (activation of PKA and phosphorylation on tyrosine residues by IF), with progressive motility (CASA) and with the intact acrosome (PSA-FITC, IF) compared to the control was observed (p<0.05). Moreover, the decrease in motility through the time was less in these SPZ than in the cryopreserved SPZ incubated in the absence of OECs. This sperm population could be recovered for its use in different ART.Improvements in handling and selection of cryopreserved SPZ not only generate tools to solve the problem that IVF presents in equines, but would also improve efficiency in other ART such as ICSI, allowing the use of a population of higher quality gametes which it would positively impact the quality of the embryos obtained. |
publishDate |
2021 |
dc.date.none.fl_str_mv |
2021 |
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info:eu-repo/semantics/publishedVersion info:eu-repo/semantics/conferenceObject Reunión Journal http://purl.org/coar/resource_type/c_5794 info:ar-repo/semantics/documentoDeConferencia |
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publishedVersion |
format |
conferenceObject |
dc.identifier.none.fl_str_mv |
http://hdl.handle.net/11336/153951 New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines; IV Joint Meeting of The Biology Societies of Argentina; Mendoza; Argentina; 2020; 47-48 0327-9545 1667-5746 CONICET Digital CONICET |
url |
http://hdl.handle.net/11336/153951 |
identifier_str_mv |
New Tools for in Vitro Handling and Selection of Cryopreserved Sperm Samples in Equines; IV Joint Meeting of The Biology Societies of Argentina; Mendoza; Argentina; 2020; 47-48 0327-9545 1667-5746 CONICET Digital CONICET |
dc.language.none.fl_str_mv |
eng |
language |
eng |
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info:eu-repo/semantics/altIdentifier/url/https://www.techscience.com/biocell/v45nSuppl.3/44000 |
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Tech Science Press |
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Tech Science Press |
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