Desmosterol Incorporation Into Ram Sperm Membrane Before Cryopreservation Improves in vitro and in vivo Fertility

Autores
Carro, María de las Mercedes; Ramírez Vasquez, Rafael R.; Peñalva, Daniel A.; Buschiazzo, Jorgelina; Hozbor, Federico Andres
Año de publicación
2021
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Pregnancy rates in ewes are markedly low after cervical insemination with frozen-thawed sperm. Sensitivity of ram sperm to freeze-thawing is related to the lipid composition of the membrane, particularly to its low sterol content. Recently, we proved that sterol content of ram sperm can be increased by treatment with methyl-β-cyclodextrin-sterol complexes and we provided mechanistic based evidence on the differential behavior of cholesterol and desmosterol in the ram sperm membrane. In the present study, we evaluated the role of increasing cholesterol and desmosterol content of ram sperm before cryopreservation, on the extent and distribution of sterols, cryocapacitation status, acrosome integrity, DNA damage associated with apoptosis and fertility competence in vitro and in vivo of post-thawed sperm. After freeze-thawing, similar levels of sterol content were evidenced in control sperm cells and in those pre-incubated with either cholesterol or desmosterol. Still, moderately higher levels of sterols were registered in treated sperm compared to the control, indicating no physiological excess of sterols after thawing or sterol losses that exceed the control. Live cell imaging of fluorescent cholesterol evidenced the presence of sperm sub-populations differentially affected by freeze-thawing. Similar unimodal frequency profiles were observed between sterol-enriched groups, while the control exhibited a sub-population of sperm compatible with low sterol content. Tyrosine phosphorylation was significantly lower when ram sperm incorporated cholesterol compared to the control. No difference in this capacitation parameter was found between the latter and desmosterol-enriched sperm. The percentage of sperm with damaged acrosomes post-thawing, assessed by a fluorescent lectin, was reduced in sperm that incorporated sterols before freezing, irrespective of the sterol class. These results suggest that sterols exert a stabilizing effect on the acrosome. No differences were found in levels of apoptotic DNA fragmentation among experimental groups. As to fertility trials, desmosterol-enriched sperm gave rise to higher rates of in vitro activated oocytes by heterologous fertilization and to significantly lower pregnancy loss in vivo. Our research provides new insights on sterol incorporation into ram sperm prior to cryopreservation, in particular on the additional benefit of incorporating desmosterol as a strategy to improve fertility outcome.
EEA Balcarce
Fil: Carro, María de las Mercedes. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.
Fil: Ramírez Vasquez, Rafael R. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.
Fil: Peñalva, Daniel A. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Sur; Argentina. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina.
Fil: Buschiazzo, Jorgelina. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.
Fil: Hozbor, Federico Andrés. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.
Fuente
Frontiers in Cell and Developmental Biology 9:660165.
Materia
Criopreservación
Ovinos
Colesterol
Inseminación Artificial
Cryopreservation
Semen
Sheep
Cholesterol
Artificial Insemination
Nivel de accesibilidad
acceso abierto
Condiciones de uso
http://creativecommons.org/licenses/by-nc-sa/4.0/
Repositorio
INTA Digital (INTA)
Institución
Instituto Nacional de Tecnología Agropecuaria
OAI Identificador
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spelling Desmosterol Incorporation Into Ram Sperm Membrane Before Cryopreservation Improves in vitro and in vivo FertilityCarro, María de las MercedesRamírez Vasquez, Rafael R.Peñalva, Daniel A.Buschiazzo, JorgelinaHozbor, Federico AndresCriopreservaciónOvinosColesterolInseminación ArtificialCryopreservationSemenSheepCholesterolArtificial InseminationPregnancy rates in ewes are markedly low after cervical insemination with frozen-thawed sperm. Sensitivity of ram sperm to freeze-thawing is related to the lipid composition of the membrane, particularly to its low sterol content. Recently, we proved that sterol content of ram sperm can be increased by treatment with methyl-β-cyclodextrin-sterol complexes and we provided mechanistic based evidence on the differential behavior of cholesterol and desmosterol in the ram sperm membrane. In the present study, we evaluated the role of increasing cholesterol and desmosterol content of ram sperm before cryopreservation, on the extent and distribution of sterols, cryocapacitation status, acrosome integrity, DNA damage associated with apoptosis and fertility competence in vitro and in vivo of post-thawed sperm. After freeze-thawing, similar levels of sterol content were evidenced in control sperm cells and in those pre-incubated with either cholesterol or desmosterol. Still, moderately higher levels of sterols were registered in treated sperm compared to the control, indicating no physiological excess of sterols after thawing or sterol losses that exceed the control. Live cell imaging of fluorescent cholesterol evidenced the presence of sperm sub-populations differentially affected by freeze-thawing. Similar unimodal frequency profiles were observed between sterol-enriched groups, while the control exhibited a sub-population of sperm compatible with low sterol content. Tyrosine phosphorylation was significantly lower when ram sperm incorporated cholesterol compared to the control. No difference in this capacitation parameter was found between the latter and desmosterol-enriched sperm. The percentage of sperm with damaged acrosomes post-thawing, assessed by a fluorescent lectin, was reduced in sperm that incorporated sterols before freezing, irrespective of the sterol class. These results suggest that sterols exert a stabilizing effect on the acrosome. No differences were found in levels of apoptotic DNA fragmentation among experimental groups. As to fertility trials, desmosterol-enriched sperm gave rise to higher rates of in vitro activated oocytes by heterologous fertilization and to significantly lower pregnancy loss in vivo. Our research provides new insights on sterol incorporation into ram sperm prior to cryopreservation, in particular on the additional benefit of incorporating desmosterol as a strategy to improve fertility outcome.EEA BalcarceFil: Carro, María de las Mercedes. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.Fil: Ramírez Vasquez, Rafael R. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.Fil: Peñalva, Daniel A. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Sur; Argentina. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina.Fil: Buschiazzo, Jorgelina. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.Fil: Hozbor, Federico Andrés. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.Frontiers Media2022-11-28T10:26:14Z2022-11-28T10:26:14Z2021-06info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfhttp://hdl.handle.net/20.500.12123/13454https://www.frontiersin.org/articles/10.3389/fcell.2021.660165/full2296-634Xhttps://doi.org/10.3389/fcell.2021.660165Frontiers in Cell and Developmental Biology 9:660165.reponame:INTA Digital (INTA)instname:Instituto Nacional de Tecnología Agropecuariaenginfo:eu-repograntAgreement/INTA/PNSA-1115053/AR./Biotecnologías reproductivas y desarrollo de metodologías de diagnóstico, control y prevención de las enfermedades infecciosas y parasitarias que afectan la concepción, gestación y período neonatal en especies de interés zootécnico.info:eu-repograntAgreement/INTA/PNSA-1115054/AR./Enfermedades parasitarias, infecciosas y tóxico metabólicas que afectan la productividad de los bóvidos para producción de carne y leche.info:eu-repo/semantics/openAccesshttp://creativecommons.org/licenses/by-nc-sa/4.0/Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)2025-09-04T09:49:38Zoai:localhost:20.500.12123/13454instacron:INTAInstitucionalhttp://repositorio.inta.gob.ar/Organismo científico-tecnológicoNo correspondehttp://repositorio.inta.gob.ar/oai/requesttripaldi.nicolas@inta.gob.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:l2025-09-04 09:49:38.98INTA Digital (INTA) - Instituto Nacional de Tecnología Agropecuariafalse
dc.title.none.fl_str_mv Desmosterol Incorporation Into Ram Sperm Membrane Before Cryopreservation Improves in vitro and in vivo Fertility
title Desmosterol Incorporation Into Ram Sperm Membrane Before Cryopreservation Improves in vitro and in vivo Fertility
spellingShingle Desmosterol Incorporation Into Ram Sperm Membrane Before Cryopreservation Improves in vitro and in vivo Fertility
Carro, María de las Mercedes
Criopreservación
Ovinos
Colesterol
Inseminación Artificial
Cryopreservation
Semen
Sheep
Cholesterol
Artificial Insemination
title_short Desmosterol Incorporation Into Ram Sperm Membrane Before Cryopreservation Improves in vitro and in vivo Fertility
title_full Desmosterol Incorporation Into Ram Sperm Membrane Before Cryopreservation Improves in vitro and in vivo Fertility
title_fullStr Desmosterol Incorporation Into Ram Sperm Membrane Before Cryopreservation Improves in vitro and in vivo Fertility
title_full_unstemmed Desmosterol Incorporation Into Ram Sperm Membrane Before Cryopreservation Improves in vitro and in vivo Fertility
title_sort Desmosterol Incorporation Into Ram Sperm Membrane Before Cryopreservation Improves in vitro and in vivo Fertility
dc.creator.none.fl_str_mv Carro, María de las Mercedes
Ramírez Vasquez, Rafael R.
Peñalva, Daniel A.
Buschiazzo, Jorgelina
Hozbor, Federico Andres
author Carro, María de las Mercedes
author_facet Carro, María de las Mercedes
Ramírez Vasquez, Rafael R.
Peñalva, Daniel A.
Buschiazzo, Jorgelina
Hozbor, Federico Andres
author_role author
author2 Ramírez Vasquez, Rafael R.
Peñalva, Daniel A.
Buschiazzo, Jorgelina
Hozbor, Federico Andres
author2_role author
author
author
author
dc.subject.none.fl_str_mv Criopreservación
Ovinos
Colesterol
Inseminación Artificial
Cryopreservation
Semen
Sheep
Cholesterol
Artificial Insemination
topic Criopreservación
Ovinos
Colesterol
Inseminación Artificial
Cryopreservation
Semen
Sheep
Cholesterol
Artificial Insemination
dc.description.none.fl_txt_mv Pregnancy rates in ewes are markedly low after cervical insemination with frozen-thawed sperm. Sensitivity of ram sperm to freeze-thawing is related to the lipid composition of the membrane, particularly to its low sterol content. Recently, we proved that sterol content of ram sperm can be increased by treatment with methyl-β-cyclodextrin-sterol complexes and we provided mechanistic based evidence on the differential behavior of cholesterol and desmosterol in the ram sperm membrane. In the present study, we evaluated the role of increasing cholesterol and desmosterol content of ram sperm before cryopreservation, on the extent and distribution of sterols, cryocapacitation status, acrosome integrity, DNA damage associated with apoptosis and fertility competence in vitro and in vivo of post-thawed sperm. After freeze-thawing, similar levels of sterol content were evidenced in control sperm cells and in those pre-incubated with either cholesterol or desmosterol. Still, moderately higher levels of sterols were registered in treated sperm compared to the control, indicating no physiological excess of sterols after thawing or sterol losses that exceed the control. Live cell imaging of fluorescent cholesterol evidenced the presence of sperm sub-populations differentially affected by freeze-thawing. Similar unimodal frequency profiles were observed between sterol-enriched groups, while the control exhibited a sub-population of sperm compatible with low sterol content. Tyrosine phosphorylation was significantly lower when ram sperm incorporated cholesterol compared to the control. No difference in this capacitation parameter was found between the latter and desmosterol-enriched sperm. The percentage of sperm with damaged acrosomes post-thawing, assessed by a fluorescent lectin, was reduced in sperm that incorporated sterols before freezing, irrespective of the sterol class. These results suggest that sterols exert a stabilizing effect on the acrosome. No differences were found in levels of apoptotic DNA fragmentation among experimental groups. As to fertility trials, desmosterol-enriched sperm gave rise to higher rates of in vitro activated oocytes by heterologous fertilization and to significantly lower pregnancy loss in vivo. Our research provides new insights on sterol incorporation into ram sperm prior to cryopreservation, in particular on the additional benefit of incorporating desmosterol as a strategy to improve fertility outcome.
EEA Balcarce
Fil: Carro, María de las Mercedes. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.
Fil: Ramírez Vasquez, Rafael R. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.
Fil: Peñalva, Daniel A. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Sur; Argentina. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina.
Fil: Buschiazzo, Jorgelina. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.
Fil: Hozbor, Federico Andrés. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.
description Pregnancy rates in ewes are markedly low after cervical insemination with frozen-thawed sperm. Sensitivity of ram sperm to freeze-thawing is related to the lipid composition of the membrane, particularly to its low sterol content. Recently, we proved that sterol content of ram sperm can be increased by treatment with methyl-β-cyclodextrin-sterol complexes and we provided mechanistic based evidence on the differential behavior of cholesterol and desmosterol in the ram sperm membrane. In the present study, we evaluated the role of increasing cholesterol and desmosterol content of ram sperm before cryopreservation, on the extent and distribution of sterols, cryocapacitation status, acrosome integrity, DNA damage associated with apoptosis and fertility competence in vitro and in vivo of post-thawed sperm. After freeze-thawing, similar levels of sterol content were evidenced in control sperm cells and in those pre-incubated with either cholesterol or desmosterol. Still, moderately higher levels of sterols were registered in treated sperm compared to the control, indicating no physiological excess of sterols after thawing or sterol losses that exceed the control. Live cell imaging of fluorescent cholesterol evidenced the presence of sperm sub-populations differentially affected by freeze-thawing. Similar unimodal frequency profiles were observed between sterol-enriched groups, while the control exhibited a sub-population of sperm compatible with low sterol content. Tyrosine phosphorylation was significantly lower when ram sperm incorporated cholesterol compared to the control. No difference in this capacitation parameter was found between the latter and desmosterol-enriched sperm. The percentage of sperm with damaged acrosomes post-thawing, assessed by a fluorescent lectin, was reduced in sperm that incorporated sterols before freezing, irrespective of the sterol class. These results suggest that sterols exert a stabilizing effect on the acrosome. No differences were found in levels of apoptotic DNA fragmentation among experimental groups. As to fertility trials, desmosterol-enriched sperm gave rise to higher rates of in vitro activated oocytes by heterologous fertilization and to significantly lower pregnancy loss in vivo. Our research provides new insights on sterol incorporation into ram sperm prior to cryopreservation, in particular on the additional benefit of incorporating desmosterol as a strategy to improve fertility outcome.
publishDate 2021
dc.date.none.fl_str_mv 2021-06
2022-11-28T10:26:14Z
2022-11-28T10:26:14Z
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dc.identifier.none.fl_str_mv http://hdl.handle.net/20.500.12123/13454
https://www.frontiersin.org/articles/10.3389/fcell.2021.660165/full
2296-634X
https://doi.org/10.3389/fcell.2021.660165
url http://hdl.handle.net/20.500.12123/13454
https://www.frontiersin.org/articles/10.3389/fcell.2021.660165/full
https://doi.org/10.3389/fcell.2021.660165
identifier_str_mv 2296-634X
dc.language.none.fl_str_mv eng
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dc.relation.none.fl_str_mv info:eu-repograntAgreement/INTA/PNSA-1115053/AR./Biotecnologías reproductivas y desarrollo de metodologías de diagnóstico, control y prevención de las enfermedades infecciosas y parasitarias que afectan la concepción, gestación y período neonatal en especies de interés zootécnico.
info:eu-repograntAgreement/INTA/PNSA-1115054/AR./Enfermedades parasitarias, infecciosas y tóxico metabólicas que afectan la productividad de los bóvidos para producción de carne y leche.
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
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Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)
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Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)
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