34 results on '"Fernández Alegre, Estela"'
Search Results
2. Single layer centrifugation (SLC) for bacterial removal with Porcicoll positively modifies chromatin structure in boar spermatozoa
- Author
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Lacalle, Estíbaliz, Fernández-Alegre, Estela, Soriano-Úbeda, Cristina, Martínez-Martínez, Sonia, Domínguez, Juan Carlos, González-Montaña, J. Ramiro, Morrell, Jane M., and Martínez-Pastor, Felipe
- Published
- 2023
- Full Text
- View/download PDF
3. Extension of the equilibration period up to 24h maintains the post-thawing quality of Holstein bull semen frozen with OPTIXcell®
- Author
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Salman, Amer, Fernández-Alegre, Estela, Francisco-Vázquez, Rubén, Gómez-Martín, Rubén, Fernández-Fernández, Alejandro, Areán-Dablanca, Héctor, Domínguez, Juan Carlos, González-Montaña, J. Ramiro, Caamaño, J. Néstor, and Martínez-Pastor, Felipe
- Published
- 2023
- Full Text
- View/download PDF
4. Application of Flow Cytometry Using Advanced Chromatin Analyses for Assessing Changes in Sperm Structure and DNA Integrity in a Porcine Model
- Author
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Lacalle, Estíbaliz, primary, Fernández-Alegre, Estela, additional, Gómez-Giménez, Belén, additional, Álvarez-Rodríguez, Manuel, additional, Martín-Fernández, Beatriz, additional, Soriano-Úbeda, Cristina, additional, and Martínez-Pastor, Felipe, additional
- Published
- 2024
- Full Text
- View/download PDF
5. Supplementation of the BIOXcell extender with the antioxidants crocin, curcumin and GSH for freezing bull semen
- Author
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Salman, Amer, Caamaño, J. Néstor, Fernández-Alegre, Estela, Hidalgo, Carlos O., Nadri, Touba, Tamargo, Carolina, Fueyo, Carmen, Fernández, Ángel, Merino, María J., and Martínez-Pastor, Felipe
- Published
- 2021
- Full Text
- View/download PDF
6. Application of Flow Cytometry Using Advanced Chromatin Analyses for Assessing Changes in Sperm Structure and DNA Integrity in a Porcine Model
- Author
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Ministerio de Ciencia, Innovación y Universidades (España), Agencia Estatal de Investigación (España), European Commission, Junta de Castilla y León, Lacalle, E., Fernández-Alegre, Estela, Gómez-Giménez, Belén, Álvarez-Rodríguez, Manuel, Martín-Fernández, B., Soriano-Úbeda, C., Martínez-Pastor, F., Ministerio de Ciencia, Innovación y Universidades (España), Agencia Estatal de Investigación (España), European Commission, Junta de Castilla y León, Lacalle, E., Fernández-Alegre, Estela, Gómez-Giménez, Belén, Álvarez-Rodríguez, Manuel, Martín-Fernández, B., Soriano-Úbeda, C., and Martínez-Pastor, F.
- Abstract
Chromatin status is critical for sperm fertility and reflects spermatogenic success. We tested a multivariate approach for studying pig sperm chromatin structure to capture its complexity with a set of quick and simple techniques, going beyond the usual assessment of DNA damage. Sperm doses from 36 boars (3 ejaculates/boar) were stored at 17 °C and analyzed on days 0 and 11. Analyses were: CASA (motility) and flow cytometry to assess sperm functionality and chromatin structure by SCSA (%DFI, DNA fragmentation; %HDS, chromatin maturity), monobromobimane (mBBr, tiol status/disulfide bridges between protamines), chromomycin A3 (CMA3, protamination), and 8-hydroxy-2′-deoxyguanosine (8-oxo-dG, DNA oxidative damage). Data were analyzed using linear models for the effects of boar and storage, correlations, and multivariate analysis as hierarchical clustering and principal component analysis (PCA). Storage reduced sperm quality parameters, mainly motility, with no critical oxidative stress increases, while chromatin status worsened slightly (%DFI and 8-oxo-dG increased while mBBr MFI—median fluorescence intensity—and disulfide bridge levels decreased). Boar significantly affected most chromatin variables except CMA3; storage also affected most variables except %HDS. At day 0, sperm chromatin variables clustered closely, except for CMA3, and %HDS and 8-oxo-dG correlated with many variables (notably, mBBr). After storage, the relation between %HDS and 8-oxo-dG remained, but correlations among other variables disappeared, and mBBr variables clustered separately. The PCA suggested a considerable influence of mBBr on sample variance, especially regarding storage, with SCSA and 8-oxo-dG affecting between-sample variability. Overall, CMA3 was the least informative, in contrast with results in other species. The combination of DNA fragmentation, DNA oxidation, chromatin compaction, and tiol status seems a good candidate for obtaining a complete picture of pig sperm nucleus status.
- Published
- 2024
7. First evaluations of extracts from grape marc as antibiotic substitutes in semen extenders
- Author
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European Commission, Fernández-Alegre, Estela [0000-0002-2223-1788], Faldyna, Martin [0000-0002-5448-7235], Boryshpolets, S. [0000-0002-7346-8734], Santiago Moreno, Julián [0000-0001-5551-8120], Martínez-Pastor, F. [0000-0003-2987-4302], Lacalle, E., Fernández-Alegre, Estela, Martínez, S., Dzyuba, B., Martín-Fernández, B., Mendoza, N., Ausejo, R., Faldyna, Martin, Boryshpolets, S., Santiago Moreno, Julián, Martínez-Pastor, F., European Commission, Fernández-Alegre, Estela [0000-0002-2223-1788], Faldyna, Martin [0000-0002-5448-7235], Boryshpolets, S. [0000-0002-7346-8734], Santiago Moreno, Julián [0000-0001-5551-8120], Martínez-Pastor, F. [0000-0003-2987-4302], Lacalle, E., Fernández-Alegre, Estela, Martínez, S., Dzyuba, B., Martín-Fernández, B., Mendoza, N., Ausejo, R., Faldyna, Martin, Boryshpolets, S., Santiago Moreno, Julián, and Martínez-Pastor, F.
- Published
- 2023
8. Pre-freezing selection of Holstein bull semen with the BoviPure colloid as double- or single-layer centrifugation improves the post-thawing quality
- Author
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Ministerio de Economía y Competitividad (España), Ministerio de Ciencia e Innovación (España), Agencia Estatal de Investigación (España), European Commission, Salman, A. [0000-0003-4960-9020], Fernández-Alegre, Estela [0000-0002-2223-1788], Domínguez, Juan C. [0000-0003-3654-1573], Álvarez-Rodríguez, Manuel [0000-0003-0120-354X], Caamaño, J. N. [0000-0001-6449-5784], Martínez-Pastor, F. [0000-0003-2987-4302], Fernández-Fernández, Alejandro [0000-0003-2524-3597], Areán-Dablanca, Héctor [0000-0002-6811-8983], Salman, A., Fernández-Alegre, Estela, Francisco-Vázquez, Rubén, Domínguez, Juan C., Álvarez-Rodríguez, Manuel, Caamaño, J. N., Martínez-Pastor, F., Gómez-Martín, Rubén, Fernández-Fernández, Alejandro, Areán-Dablanca, Héctor, Ministerio de Economía y Competitividad (España), Ministerio de Ciencia e Innovación (España), Agencia Estatal de Investigación (España), European Commission, Salman, A. [0000-0003-4960-9020], Fernández-Alegre, Estela [0000-0002-2223-1788], Domínguez, Juan C. [0000-0003-3654-1573], Álvarez-Rodríguez, Manuel [0000-0003-0120-354X], Caamaño, J. N. [0000-0001-6449-5784], Martínez-Pastor, F. [0000-0003-2987-4302], Fernández-Fernández, Alejandro [0000-0003-2524-3597], Areán-Dablanca, Héctor [0000-0002-6811-8983], Salman, A., Fernández-Alegre, Estela, Francisco-Vázquez, Rubén, Domínguez, Juan C., Álvarez-Rodríguez, Manuel, Caamaño, J. N., Martínez-Pastor, F., Gómez-Martín, Rubén, Fernández-Fernández, Alejandro, and Areán-Dablanca, Héctor
- Abstract
Artificial insemination (AI) is critical for breeding in the dairy industry. High-merit bulls can present low freezability, hampering genetic dissemination. Thawed semen can be improved using density gradient centrifugation (DGC) with colloids, but little information deals with the pre-freezing application. Thus, the BoviPure colloid (optimized for bull spermatozoa) was tested for pre-freezing application as the usual double-layer (DLC) versus single-layer (SLC, quick and economical). Semen from twelve Holstein-Friesian bulls was extended with OPTIXcell extender, frozen (Control), or processed by SLC or DLC and frozen. Sperm were assessed pre-freezing for motility and viability and post-thawing (directly and after 4 h 38 °C) for apoptosis, capacitation status, acrosomal damage, mitochondrial activity, cytoplasmic and mitochondrial reactive oxygen species (ROS), and chromatin status (SCSA for DNA fragmentation and chromatin compaction and monobromobimane, mBBr, for disulfide bridges evaluation). The DGC improved parameters post-thawing (e.g., 57.5%±10.1 motility vs. control 53.3% ± 11.2) at the cost of sperm loss (sperm recovery of DGC 14.4% ± 2.5 and SLC 17.4% ± 2.5). DNA fragmentation (%DFI) decreased (0.21% ± 0.53 vs. control 1.30% ± 0.10), and SLC reduced chromatin compaction. A clustering procedure separated lesser (LF) and greater freezability (GF) bulls. LF samples were especially benefited by DGC, with SLC providing better post-thawing results for this group. In conclusion, pre-freezing DGC improved sperm parameters post-thawing, potentially improving the cryopreservation of low-freezability semen from high-merit bulls. SLC, quicker and economical, would be preferable since it showed similar or higher performance than DLC.
- Published
- 2023
9. Seminal plasma proteins modify the distribution of sperm subpopulations in cryopreserved semen of rams with lesser fertility
- Author
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Ledesma, Alba, Zalazar, Lucía, Fernández-Alegre, Estela, Hozbor, Federico, Cesari, Andreina, and Martínez-Pastor, Felipe
- Published
- 2017
- Full Text
- View/download PDF
10. Extension of the equilibration period up to 24 h maintains the post-thawing quality of Holstein bull semen frozen with OPTIXcell®
- Author
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Salman, Amer, primary, Fernández-Alegre, Estela, additional, Francisco-Vázquez, Rubén, additional, Gómez-Martín, Rubén, additional, Fernández-Fernández, Alejandro, additional, Areán-Dablanca, Héctor, additional, Domínguez, Juan Carlos, additional, González-Montaña, J. Ramiro, additional, Caamaño, J.Néstor, additional, and Martínez-Pastor, Felipe, additional
- Published
- 2023
- Full Text
- View/download PDF
11. Seminal plasma proteins interacting with sperm surface revert capacitation indicators in frozen-thawed ram sperm
- Author
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Ledesma, Alba, Fernández-Alegre, Estela, Cano, Adriana, Hozbor, Federico, Martínez-Pastor, Felipe, and Cesari, Andreína
- Published
- 2016
- Full Text
- View/download PDF
12. Bos taurus and Cervus elaphus as Non-Seasonal/Seasonal Models for the Role of Melatonin Receptors in the Spermatozoon
- Author
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Fernández-Alegre, Estela, primary, Lacalle, Estíbaliz, additional, Soriano-Úbeda, Cristina, additional, González-Montaña, José Ramiro, additional, Domínguez, Juan Carlos, additional, Casao, Adriana, additional, and Martínez-Pastor, Felipe, additional
- Published
- 2022
- Full Text
- View/download PDF
13. Melatonin affects red deer spermatozoa motility and physiology in capacitating and non‐capacitating conditions
- Author
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Fernández‐Alegre, Estela, primary, Lacalle, Estíbaliz, additional, Soriano‐Úbeda, Cristina, additional, Carlos Domínguez, Juan, additional, Casao, Adriana, additional, and Martínez‐Pastor, Felipe, additional
- Published
- 2022
- Full Text
- View/download PDF
14. Cold-Shock Test Is a Practical Method for Selecting Boar Ejaculates Yielding Appropriate Seminal Plasma for Post-Thawing Supplementation
- Author
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Lacalle, Estíbaliz, primary, Núñez, Andrea, additional, Fernández-Alegre, Estela, additional, Crespo-Félez, Itxaso, additional, Domínguez, Juan Carlos, additional, Alonso, Marta Elena, additional, González-Urdiales, Raúl, additional, and Martínez-Pastor, Felipe, additional
- Published
- 2021
- Full Text
- View/download PDF
15. Melatonin Non-Linearly Modulates Bull Spermatozoa Motility and Physiology in Capacitating and Non-Capacitating Conditions
- Author
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Fernández-Alegre, Estela, primary, Álvarez-Fernández, Indira, additional, Domínguez, Juan Carlos, additional, Casao, Adriana, additional, and Martínez-Pastor, Felipe, additional
- Published
- 2020
- Full Text
- View/download PDF
16. Removal of bacteria from boar semen using a low-density colloid
- Author
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Morrell, Jane Margaret, Núñez González, A., Crespo Félez, I., Martínez Martínez, Sonia, Martínez Alborcia, María J., Fernández Alegre, Estela, Domínguez Fernández De Tejerina, Juan Carlos, Gutiérrez Martín, César Bernardo, Martínez Pastor, Felipe, Biologia Celular, and Facultad de Ciencias Biologicas y Ambientales
- Subjects
endocrine system ,urogenital system ,Semen ,Jabalíes ,Centrifugación ,Veterinaria ,Coloides - Abstract
P. 272-278 Antibiotics are added to semen extenders when preparing commercial semen doses for artificial insemination according to national and international guidelines. However, this addition of antibiotics represents non-therapeutic usage and could be contributing to the development of antibiotic resistance. Colloid centrifugation was shown to reduce the load of bacteria present in boar semen and was capable of removing all bacteria if performed directly after semen collection, albeit with some loss of spermatozoa. The present experiment was conducted with a low density colloid to investigate whether it was possible to separate all of the spermatozoa from seminal plasma i.e. without selection for robust spermatozoa, or whether this would have a detrimental effect on sperm quality. Ejaculates from nine boars were extended in Beltsville Thawing Solution without antibiotics and were transported to the laboratory for Single Layer Centrifugation (SLC) on modified Porcicoll i.e. at a low density (S). A further modification was that a sterile inner tube was included inside some of the 50 mL centrifuge tubes to facilitate harvesting of the sperm pellet (M). Aliquots of all samples (control, S and M) were cultured for bacterial quantification and identification using standard microbiological methods. Sperm quality was evaluated daily. Three of the C and M samples and five of the S samples did not contain any bacteria. Mean bacterial counts for the remaining samples (colony forming units/mL) were as follows: C 259 ± 216; S 30 ± 22; M 33 ± 15 (P
- Published
- 2019
17. Seminal plasma proteins modify the distribution of sperm subpopulations in cryopreserved semen of rams with lesser fertility
- Author
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Ledesma, Alba, Biologia Celular, Ledesma, Alba, Zalazar, Lucía, Fernández Alegre, Estela, Hozbor, Federico A., Cesari, Andreína, Martínez Pastor, Felipe, Ledesma, Alba, Biologia Celular, Ledesma, Alba, Zalazar, Lucía, Fernández Alegre, Estela, Hozbor, Federico A., Cesari, Andreína, and Martínez Pastor, Felipe
- Abstract
Any physiological mechanism involved in sperm selection and semen improvement has effects on heterogeneous sperm populations. This is mainly due to the fact that sperm populations within a single ejaculate have considerable heterogeneity for many variables, such as motility which is meaningful in terms of understanding how some sperm cells possess fertility advantages as compared with other cells. In the present research, initially there was a multivariate and clustering analysis used to assess sperm motility data from cryopreserved ram semen to identify subpopulations and compare the distribution of these clusters between rams with lesser and greater fertility. There were four classifications made of sperm subpopulations (clusters): CL1 fast/linear/progressive sperm; CL2 fast/non-linear sperm; CL3 very fast/linear sperm with vigorous beating and CL4 slow/non-linear sperm. Rams with greater fertility had a lesser proportion of sperm considered as “hyperactivated” (CL2) and a greater proportion of slow and non-linear sperm (CL4) than sperm of rams with lesser fertility. In addition, the effects were assessed for the capacity of seminal plasma (SP) and interacting SP proteins (iSPP) that were present during different seasons of the year to improve the distribution of sperm within subpopulations of semen from rams with lesser fertility. The iSPP and SP were obtained by artificial vagina (AV) and electroejaculation (EE) during breeding and non-breeding seasons and added to thawed semen. All the aggregates had a significant effect on the distribution of sperm subpopulations and effects differed among seasons of the year and depending on collection method used. Even though, future studies are needed to assess the contribution of each subpopulation on ram sperm fertility, it is important that a multivariate analysis be used to evaluate the effect of a treatment on sperm quality variables.
- Published
- 2019
18. Seminal plasma proteins interacting with sperm surface revert capacitation indicators in frozen-thawed ram sperm
- Author
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Ledesma, Alba, Biologia Celular, Ledesma, Alba, Fernández Alegre, Estela, Cano, Adriana, Hozbor, Federico A., Martínez Pastor, Felipe, Cesari, Andreína, Ledesma, Alba, Biologia Celular, Ledesma, Alba, Fernández Alegre, Estela, Cano, Adriana, Hozbor, Federico A., Martínez Pastor, Felipe, and Cesari, Andreína
- Abstract
This study was conducted to evaluate the effects of interacting seminal plasma proteins (iSPP) obtained by AV or EE on frozen-thawed ram sperm in order to test the hypothesis whether this fraction could be sufficient to emulate the effect of complete seminal plasma (SP). Additionally, we evaluated whether these proteins have a differential effect between spermatozoa from high and low fertility rams and between breeding and non-breeding seasons. We assessed sperm motility, quality parameters (intracellular reactive oxygen species, membrane fluidity, plasma membrane permeability and mitochondrial activity) and capacitation status. The main findings from this work were: i) iSPP had no effect on sperm motility, whereas SP (AV or EE) addition produced the highest values of total motility (74.13 ± 2.99 and 72.27 ± 2.99 for AV and EE, respectively) and progressive motility (64.97 ± 2.64 and 63.73 ± 2.64 for AV and EE, respectively); ii) iSPP had no effect on sperm quality parameters (p > 0.05), but whole SP improved all parameters evaluated. Moreover, SP collected by AV yielded significantly higher viability (44.60 ± 2.87) and sperm with stable plasma membrane (44.56 ± 2.49) comparing with the addition of SP collected by EE (35.80 ± 2.47 and 36.67 ± 1.71, respectively); iii) iSPP and SP collected by EE, but not by AV, reverted molecular signals of capacitation as protein tyrosine phosphorylation caused by freezing temperatures; iv) there were no effects of fertility or season in sperm quality parameters evaluated. This study demonstrated that, although the iSPP have a clear decapacitating effect, including the ability to revert cryo-capacitation indicators, they are not sufficient to emulate the effects of complete SP regarding sperm functional parameters.
- Published
- 2019
19. Melatonin receptors MT1 and MT2 are expressed in spermatozoa from several seasonal and nonseasonal breeder species
- Author
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González Arto, Marta, Biologia Celular, González Arto, Marta, Vicente Carrillo, Alejandro, Martínez Pastor, Felipe, Fernández Alegre, Estela, Roca Aleu, Jorge, Miró Roig, Jordi, Rigau, Teresa, Rodríguez Gil, Joan Enric, Pérez Pé, Rosaura, Muiño Blanco, María Teresa, Cebrián Pérez, José A., Casao Gascón, Adriana, González Arto, Marta, Biologia Celular, González Arto, Marta, Vicente Carrillo, Alejandro, Martínez Pastor, Felipe, Fernández Alegre, Estela, Roca Aleu, Jorge, Miró Roig, Jordi, Rigau, Teresa, Rodríguez Gil, Joan Enric, Pérez Pé, Rosaura, Muiño Blanco, María Teresa, Cebrián Pérez, José A., and Casao Gascón, Adriana
- Abstract
Melatonin is a ubiquitous and multipurpose molecule, and one of its roles is to regulate reproduction in some seasonal mammals. Our group has previously reported the variation in the melatonin levels in ram seminal plasma along the year and identified MT1 and MT2 receptors in ram spermatozoa. The objective of this study was to elucidate whether the presence of melatonin receptors (MT1 and MT2) in the sperm plasma membrane, and melatonin in the seminal plasma is related to seasonal breeding. For this purpose, the presence of melatonin receptors and the levels of melatonin in seminal plasma have been examined in several species: donkey and stallion as long-day breeders; red deer as a wild, short-day, highly seasonal breeder (epididymal spermatozoa); bull as a conventional nonseasonal breeder; boar as a seasonal breeder under management techniques; and dog as possible a seasonal breeder not regulated by melatonin. We have detected measurable levels of melatonin in the seminal plasma of all ejaculated semen samples (from donkey, stallion, boar, bull, and dog). Also, and for the first time, we have demonstrated the presence of MT1 and MT2 melatonin receptors in the spermatozoa of all these species, regardless their type of reproduction or sperm source (ejaculated or epididymal), using indirect immunofluorescence techniques and Western blotting. Our findings suggest that melatonin and melatonin receptors may be universally distributed in the reproductive system of mammals and that the sperm melatonin receptors cells may not be necessarily related with seasonal reproduction. Furthermore, the presence of MT1 at the cytoplasmic droplet in immature ejaculated stallion spermatozoa found in one sample and epididymal red deer spermatozoa suggests that melatonin may be involved in specific functions during spermatogenesis and sperm maturation, like protecting spermatozoa from oxidative damage, this activity being mediated through these receptors.
- Published
- 2019
20. Seminal plasma proteins modify the distribution of sperm subpopulations in cryopreserved semen of rams with lesser fertility
- Author
-
Biologia Celular, Ledesma, Alba, Zalazar, Lucía, Fernández Alegre, Estela, Hozbor, Federico A., Cesari, Andreína, Martínez Pastor, Felipe, Biologia Celular, Ledesma, Alba, Zalazar, Lucía, Fernández Alegre, Estela, Hozbor, Federico A., Cesari, Andreína, and Martínez Pastor, Felipe
- Abstract
Any physiological mechanism involved in sperm selection and semen improvement has effects on heterogeneous sperm populations. This is mainly due to the fact that sperm populations within a single ejaculate have considerable heterogeneity for many variables, such as motility which is meaningful in terms of understanding how some sperm cells possess fertility advantages as compared with other cells. In the present research, initially there was a multivariate and clustering analysis used to assess sperm motility data from cryopreserved ram semen to identify subpopulations and compare the distribution of these clusters between rams with lesser and greater fertility. There were four classifications made of sperm subpopulations (clusters): CL1 fast/linear/progressive sperm; CL2 fast/non-linear sperm; CL3 very fast/linear sperm with vigorous beating and CL4 slow/non-linear sperm. Rams with greater fertility had a lesser proportion of sperm considered as “hyperactivated” (CL2) and a greater proportion of slow and non-linear sperm (CL4) than sperm of rams with lesser fertility. In addition, the effects were assessed for the capacity of seminal plasma (SP) and interacting SP proteins (iSPP) that were present during different seasons of the year to improve the distribution of sperm within subpopulations of semen from rams with lesser fertility. The iSPP and SP were obtained by artificial vagina (AV) and electroejaculation (EE) during breeding and non-breeding seasons and added to thawed semen. All the aggregates had a significant effect on the distribution of sperm subpopulations and effects differed among seasons of the year and depending on collection method used. Even though, future studies are needed to assess the contribution of each subpopulation on ram sperm fertility, it is important that a multivariate analysis be used to evaluate the effect of a treatment on sperm quality variables.
- Published
- 2019
21. Melatonin receptors MT1 and MT2 are expressed in spermatozoa from several seasonal and nonseasonal breeder species
- Author
-
Biologia Celular, González Arto, Marta, Vicente Carrillo, Alejandro, Martínez Pastor, Felipe, Fernández Alegre, Estela, Roca Aleu, Jorge, Miró Roig, Jordi, Rigau, Teresa, Rodríguez Gil, Joan Enric, Pérez Pé, Rosaura, Muiño Blanco, María Teresa, Cebrián Pérez, José A., Casao Gascón, Adriana, Biologia Celular, González Arto, Marta, Vicente Carrillo, Alejandro, Martínez Pastor, Felipe, Fernández Alegre, Estela, Roca Aleu, Jorge, Miró Roig, Jordi, Rigau, Teresa, Rodríguez Gil, Joan Enric, Pérez Pé, Rosaura, Muiño Blanco, María Teresa, Cebrián Pérez, José A., and Casao Gascón, Adriana
- Abstract
Melatonin is a ubiquitous and multipurpose molecule, and one of its roles is to regulate reproduction in some seasonal mammals. Our group has previously reported the variation in the melatonin levels in ram seminal plasma along the year and identified MT1 and MT2 receptors in ram spermatozoa. The objective of this study was to elucidate whether the presence of melatonin receptors (MT1 and MT2) in the sperm plasma membrane, and melatonin in the seminal plasma is related to seasonal breeding. For this purpose, the presence of melatonin receptors and the levels of melatonin in seminal plasma have been examined in several species: donkey and stallion as long-day breeders; red deer as a wild, short-day, highly seasonal breeder (epididymal spermatozoa); bull as a conventional nonseasonal breeder; boar as a seasonal breeder under management techniques; and dog as possible a seasonal breeder not regulated by melatonin. We have detected measurable levels of melatonin in the seminal plasma of all ejaculated semen samples (from donkey, stallion, boar, bull, and dog). Also, and for the first time, we have demonstrated the presence of MT1 and MT2 melatonin receptors in the spermatozoa of all these species, regardless their type of reproduction or sperm source (ejaculated or epididymal), using indirect immunofluorescence techniques and Western blotting. Our findings suggest that melatonin and melatonin receptors may be universally distributed in the reproductive system of mammals and that the sperm melatonin receptors cells may not be necessarily related with seasonal reproduction. Furthermore, the presence of MT1 at the cytoplasmic droplet in immature ejaculated stallion spermatozoa found in one sample and epididymal red deer spermatozoa suggests that melatonin may be involved in specific functions during spermatogenesis and sperm maturation, like protecting spermatozoa from oxidative damage, this activity being mediated through these receptors.
- Published
- 2019
22. Seminal plasma proteins interacting with sperm surface revert capacitation indicators in frozen-thawed ram sperm
- Author
-
Biologia Celular, Ledesma, Alba, Fernández Alegre, Estela, Cano, Adriana, Hozbor, Federico A., Martínez Pastor, Felipe, Cesari, Andreína, Biologia Celular, Ledesma, Alba, Fernández Alegre, Estela, Cano, Adriana, Hozbor, Federico A., Martínez Pastor, Felipe, and Cesari, Andreína
- Abstract
This study was conducted to evaluate the effects of interacting seminal plasma proteins (iSPP) obtained by AV or EE on frozen-thawed ram sperm in order to test the hypothesis whether this fraction could be sufficient to emulate the effect of complete seminal plasma (SP). Additionally, we evaluated whether these proteins have a differential effect between spermatozoa from high and low fertility rams and between breeding and non-breeding seasons. We assessed sperm motility, quality parameters (intracellular reactive oxygen species, membrane fluidity, plasma membrane permeability and mitochondrial activity) and capacitation status. The main findings from this work were: i) iSPP had no effect on sperm motility, whereas SP (AV or EE) addition produced the highest values of total motility (74.13 ± 2.99 and 72.27 ± 2.99 for AV and EE, respectively) and progressive motility (64.97 ± 2.64 and 63.73 ± 2.64 for AV and EE, respectively); ii) iSPP had no effect on sperm quality parameters (p > 0.05), but whole SP improved all parameters evaluated. Moreover, SP collected by AV yielded significantly higher viability (44.60 ± 2.87) and sperm with stable plasma membrane (44.56 ± 2.49) comparing with the addition of SP collected by EE (35.80 ± 2.47 and 36.67 ± 1.71, respectively); iii) iSPP and SP collected by EE, but not by AV, reverted molecular signals of capacitation as protein tyrosine phosphorylation caused by freezing temperatures; iv) there were no effects of fertility or season in sperm quality parameters evaluated. This study demonstrated that, although the iSPP have a clear decapacitating effect, including the ability to revert cryo-capacitation indicators, they are not sufficient to emulate the effects of complete SP regarding sperm functional parameters.
- Published
- 2019
23. Removal of bacteria from boar semen using a low-density colloid
- Author
-
Biologia Celular, Morrell, Jane Margaret, Núñez González, A., Crespo Félez, I., Martínez Martínez, Sonia, Martínez Alborcia, María J., Fernández Alegre, Estela, Domínguez Fernández De Tejerina, Juan Carlos, Gutiérrez Martín, César Bernardo, Martínez Pastor, Felipe, Biologia Celular, Morrell, Jane Margaret, Núñez González, A., Crespo Félez, I., Martínez Martínez, Sonia, Martínez Alborcia, María J., Fernández Alegre, Estela, Domínguez Fernández De Tejerina, Juan Carlos, Gutiérrez Martín, César Bernardo, and Martínez Pastor, Felipe
- Abstract
Antibiotics are added to semen extenders when preparing commercial semen doses for artificial insemination according to national and international guidelines. However, this addition of antibiotics represents non-therapeutic usage and could be contributing to the development of antibiotic resistance. Colloid centrifugation was shown to reduce the load of bacteria present in boar semen and was capable of removing all bacteria if performed directly after semen collection, albeit with some loss of spermatozoa. The present experiment was conducted with a low density colloid to investigate whether it was possible to separate all of the spermatozoa from seminal plasma i.e. without selection for robust spermatozoa, or whether this would have a detrimental effect on sperm quality. Ejaculates from nine boars were extended in Beltsville Thawing Solution without antibiotics and were transported to the laboratory for Single Layer Centrifugation (SLC) on modified Porcicoll i.e. at a low density (S). A further modification was that a sterile inner tube was included inside some of the 50 mL centrifuge tubes to facilitate harvesting of the sperm pellet (M). Aliquots of all samples (control, S and M) were cultured for bacterial quantification and identification using standard microbiological methods. Sperm quality was evaluated daily. Three of the C and M samples and five of the S samples did not contain any bacteria. Mean bacterial counts for the remaining samples (colony forming units/mL) were as follows: C 259 ± 216; S 30 ± 22; M 33 ± 15 (P < 0.01). Citrobacter spp., Staphylococcus simulans, Klebsiella variicola, Escherichia coli, Myroides odoratimimus, Proteus spp. and Enterococcus faecalis were identified in the control samples. There were marginal differences in sperm quality among treatments, with sperm velocity and linearity being higher in S and M samples than in C at all time points. However, sperm viability, capacitation and acrosome status were marginally better in contro
- Published
- 2019
24. Caracterización morfológica del caballo de la montaña asturiana
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Lomillos Pérez, Juan Manuel, Armenteros Santos, José Ángel, Álvarez Espada, Aitor, Gómez Seco, Cristina, Fernández Alegre, Estela, Alonso De La Varga, Marta Elena, Producción Animal, and Facultad de Veterinaria
- Subjects
Asturias (Comunidad Autónoma) ,Caballos ,Veterinaria ,Carne de caballo ,2401.01 Anatomía Animal - Abstract
Las razas de caballos autóctonos españoles existentes hasta el siglo XIX, no eran adecuadas para el tiro pesado y desde que a finales del siglo XVI, Juan Jerónimo de Tiutti introdujera en nuestro país sementales flamencos, napolitanos y daneses, buscando aumentar la corpulencia de los caballos autóctonos, se ha recorrido un largo camino de intentos, fracasos y éxitos hasta conseguir actualmente la consolidación de diversas poblaciones dedicadas a la producción de caballos cuya aptitud principal es la producción de carne. Tomando como base la cabaña de yeguas autóctonas, los servicios de cría caballar del Ministerio de Defensa realizaron, a partir de los años 30 del siglo pasado, una destacada labor de mejora utilizando diversas razas, de las cuales el postier bretón resultó ser la más adecuada para formar las principales razas de carácter cárnico de España cuyos estándares se fijaron en las siguientes fechas: hispano-bretón (H-B) en 1998, en 1999 el burguete (B), en 2001 la jaca navarra (JN), en 2008 el caballo pirenaico catalán (CPC) y en 2015 el caballo del monte del País Vasco (CMPV) En el Principado de Asturias existen efectivos de caballos pesados criados en la montaña sin estar encuadrados en ninguna agrupación racial reconocida. El objetivo del presente trabajo es analizar y caracterizar racialmente dichos efectivos comprobando si existen diferencias con los efectivos H-B que puedan sustentar su consideración como nueva raza.
- Published
- 2017
25. Melatonin receptors MT1 and MT2 are expressed in spermatozoa from several seasonal and nonseasonal breeder species
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González Arto, Marta, Vicente Carrillo, Alejandro, Martínez Pastor, Felipe, Fernández Alegre, Estela, Roca Aleu, Jorge, Miró Roig, Jordi, Rigau, Teresa, Rodríguez Gil, Joan Enric, Pérez Pé, Rosaura, Muiño Blanco, María Teresa, Cebrián Pérez, José A., Casao Gascón, Adriana, Biologia Celular, and Facultad de Ciencias Biologicas y Ambientales
- Subjects
endocrine system ,Stallion ,urogenital system ,animal diseases ,Donkey ,Deer ,Boar ,Dog ,Veterinaria ,Bull ,hormones, hormone substitutes, and hormone antagonists ,reproductive and urinary physiology - Abstract
P. 1958-1968 Melatonin is a ubiquitous and multipurpose molecule, and one of its roles is to regulate reproduction in some seasonal mammals. Our group has previously reported the variation in the melatonin levels in ram seminal plasma along the year and identified MT1 and MT2 receptors in ram spermatozoa. The objective of this study was to elucidate whether the presence of melatonin receptors (MT1 and MT2) in the sperm plasma membrane, and melatonin in the seminal plasma is related to seasonal breeding. For this purpose, the presence of melatonin receptors and the levels of melatonin in seminal plasma have been examined in several species: donkey and stallion as long-day breeders; red deer as a wild, short-day, highly seasonal breeder (epididymal spermatozoa); bull as a conventional nonseasonal breeder; boar as a seasonal breeder under management techniques; and dog as possible a seasonal breeder not regulated by melatonin. We have detected measurable levels of melatonin in the seminal plasma of all ejaculated semen samples (from donkey, stallion, boar, bull, and dog). Also, and for the first time, we have demonstrated the presence of MT1 and MT2 melatonin receptors in the spermatozoa of all these species, regardless their type of reproduction or sperm source (ejaculated or epididymal), using indirect immunofluorescence techniques and Western blotting. Our findings suggest that melatonin and melatonin receptors may be universally distributed in the reproductive system of mammals and that the sperm melatonin receptors cells may not be necessarily related with seasonal reproduction. Furthermore, the presence of MT1 at the cytoplasmic droplet in immature ejaculated stallion spermatozoa found in one sample and epididymal red deer spermatozoa suggests that melatonin may be involved in specific functions during spermatogenesis and sperm maturation, like protecting spermatozoa from oxidative damage, this activity being mediated through these receptors. SI
- Published
- 2016
26. Melatonin receptors MT1 and MT2 are expressed in spermatozoa from several seasonal and nonseasonal breeder species
- Author
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González-Arto, Marta, primary, Vicente-Carrillo, Alejandro, additional, Martínez-Pastor, Felipe, additional, Fernández-Alegre, Estela, additional, Roca, Jordi, additional, Miró, Jordi, additional, Rigau, Teresa, additional, Rodríguez-Gil, Joan E., additional, Pérez-Pé, Rosaura, additional, Muiño-Blanco, Teresa, additional, Cebrián-Pérez, José A., additional, and Casao, Adriana, additional
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- 2016
- Full Text
- View/download PDF
27. Complete genome sequence of ‘Mycobacterium neoaurum’ NRRL B-3805, an androstenedione (AD) producer for industrial biotransformation of sterols
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Rodríguez‐García, Antonio, primary, Fernández‐Alegre, Estela, additional, Morales, Alejandro, additional, Sola‐Landa, Alberto, additional, Lorraine, Jess, additional, Macdonald, Sandy, additional, Dovbnya, Dmitry, additional, Smith, Margaret C.M., additional, Donova, Marina, additional, and Barreiro, Carlos, additional
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- 2016
- Full Text
- View/download PDF
28. Metabolismo de la melatonina en el tracto genital masculino de rumiante. Implicación de esta hormona en la calidad seminal y fisiología de los espermatozoides = Metabolism of melatonin in the male genital tract of ruminal. Involvement of this hormone in semen quality and sperm physiology
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Fernández Alegre, Estela, primary
- Full Text
- View/download PDF
29. Caracterización morfológica del caballo de la montaña asturiana
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Lomillos Pérez, Juan Manuel, Producción Animal, Lomillos Pérez, Juan Manuel, Armenteros Santos, José Ángel, Álvarez Espada, Aitor, Gómez Seco, Cristina, Fernández Alegre, Estela, Alonso De La Varga, Marta Elena, Lomillos Pérez, Juan Manuel, Producción Animal, Lomillos Pérez, Juan Manuel, Armenteros Santos, José Ángel, Álvarez Espada, Aitor, Gómez Seco, Cristina, Fernández Alegre, Estela, and Alonso De La Varga, Marta Elena
- Abstract
Las razas de caballos autóctonos españoles existentes hasta el siglo XIX, no eran adecuadas para el tiro pesado y desde que a finales del siglo XVI, Juan Jerónimo de Tiutti introdujera en nuestro país sementales flamencos, napolitanos y daneses, buscando aumentar la corpulencia de los caballos autóctonos, se ha recorrido un largo camino de intentos, fracasos y éxitos hasta conseguir actualmente la consolidación de diversas poblaciones dedicadas a la producción de caballos cuya aptitud principal es la producción de carne. Tomando como base la cabaña de yeguas autóctonas, los servicios de cría caballar del Ministerio de Defensa realizaron, a partir de los años 30 del siglo pasado, una destacada labor de mejora utilizando diversas razas, de las cuales el postier bretón resultó ser la más adecuada para formar las principales razas de carácter cárnico de España cuyos estándares se fijaron en las siguientes fechas: hispano-bretón (H-B) en 1998, en 1999 el burguete (B), en 2001 la jaca navarra (JN), en 2008 el caballo pirenaico catalán (CPC) y en 2015 el caballo del monte del País Vasco (CMPV) En el Principado de Asturias existen efectivos de caballos pesados criados en la montaña sin estar encuadrados en ninguna agrupación racial reconocida. El objetivo del presente trabajo es analizar y caracterizar racialmente dichos efectivos comprobando si existen diferencias con los efectivos H-B que puedan sustentar su consideración como nueva raza.
30. Extension of the equilibration period up to 24 h maintains the post-thawing quality of Holstein bull semen frozen with OPTIXcell®
- Author
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Biologia Celular, Salman, Amer, Fernández-Alegre, Estela, Francisco-Vázquez, Rubén, Gómez-Martín, Rubén, Fernández-Fernández, Alejandro, Areán-Dablanca, Héctor, Domínguez Fernández de Tejerina, Juan Carlos, González Montaña, José Ramiro, Caamaño Gualdoni, José Néstor, Martínez Pastor, Felipe, Biologia Celular, Salman, Amer, Fernández-Alegre, Estela, Francisco-Vázquez, Rubén, Gómez-Martín, Rubén, Fernández-Fernández, Alejandro, Areán-Dablanca, Héctor, Domínguez Fernández de Tejerina, Juan Carlos, González Montaña, José Ramiro, Caamaño Gualdoni, José Néstor, and Martínez Pastor, Felipe
- Abstract
[EN] Semen cryopreservation in bovine livestock is well established, but logistics often require deviations from standard protocols. Extending the equilibration time to the following day is convenient in many situations. To improve our knowledge of the effects of this modification, we studied the post-thawing and post-incubation (4 h, 38 °C) sperm quality after freezing with 4 or 24-h extension in the OPTIXcell extender by using an ample panel of analyses: CASA for motility; flow cytometry for viability, physiology, oxidative stress, and chromatin parameters (DNA fragmentation, chromatin compaction, and thiol groups status); and spectrometry for malondialdehyde production. Semen was obtained from 12 Holstein bulls. The 24-h equilibration time showed few significant effects, with only a tiny decrease in progressive motility and a positive impact on chromatin structure. The incubation removed some of these effects, with the pattern for chromatin compaction remaining the same. No detrimental oxidative stress or increase in apoptotic or capacitation markers was detected. Additionally, the individual bull interacted with the effects of the incubation and the equilibration, especially regarding the chromatin status. Whereas this interaction did not critically affect sperm quality, it could be relevant in practice. Bull fertility as non-return rates (NRR56) was associated with some sperm parameters (especially with an improved chromatin structure) but not in the 4-h post-thawing analysis. Our study supports that extending the equilibration time by at least 24-h is feasible for bull semen freezing with the OPTIXcell extender.
31. Melatonin affects red deer spermatozoa motility and physiology in capacitating and non‐capacitating conditions
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Biologia Celular, Fernández‐Alegre, Estela, Lacalle Fernández, Estivaliz, Soriano‐Úbeda, Cristina, Domínguez Fernández de Tejerina, Juan Carlos, Casao, Adriana, Martínez Pastor, Felipe, Biologia Celular, Fernández‐Alegre, Estela, Lacalle Fernández, Estivaliz, Soriano‐Úbeda, Cristina, Domínguez Fernández de Tejerina, Juan Carlos, Casao, Adriana, and Martínez Pastor, Felipe
- Abstract
[EN] Melatonin affects sperm physiology, possibly through membrane receptors. Effects were tested at low concentrations (1 pM, 100 pM, 10 nM and 1 µM) in red deer epididymal spermatozoa as a model for high-seasonality species. Samples were incubated with melatonin as uncapacitated or capacitating conditions (heparin) and evaluated for motility and physiology (flow cytometry). Most effects occurred at low concentrations (nM-pM), mainly protecting from apoptosis and maintaining acrosomal integrity, suggesting a role for membrane receptors rather than a direct antioxidant effect. Intracellular calcium was not affected, differing from other studies and perhaps because of the epididymal origin. This study supports the relevance of melatonin on sperm physiology and could contribute to the application of reproductive technologies in wild ruminants.
32. Cold-Shock Test Is a Practical Method for Selecting Boar Ejaculates Yielding Appropriate Seminal Plasma for Post-Thawing Supplementation
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Bioquimica y Biologia Molecular, Lacalle, Estíbaliz, Núñez, Andrea, Fernández-Alegre, Estela, Crespo-Félez, Itxaso, Domínguez Fernández de Tejerina, Juan Carlos, Alonso de la Varga, Marta Elena, González Urdiales, Raúl, Martínez Pastor, Felipe, Bioquimica y Biologia Molecular, Lacalle, Estíbaliz, Núñez, Andrea, Fernández-Alegre, Estela, Crespo-Félez, Itxaso, Domínguez Fernández de Tejerina, Juan Carlos, Alonso de la Varga, Marta Elena, González Urdiales, Raúl, and Martínez Pastor, Felipe
- Abstract
Artificial insemination (AI) with cryopreserved semen is still unreliable for extensive pig industry application. Adding seminal plasma (SP) could improve post-thawing quality, but its suitability could vary. We applied a simple cold-shock test (CST, 5 min at 0 °C) on neat semen for classifying ejaculates (n = 63) as resistant or sensitive, obtaining two SP pools (CST-resistant: SPr, sensitive: SPs). Subsequently, frozen/thawed spermatozoa from six boars were incubated (37 °C) in MR-A® extender (control), 20% SPr, or 20% SPs, and analyzed at 0, 2, and 4 h. SP improved total and progressive motility, with a higher effect for SPr and STR (p < 0.05), decreasing kinematic parameters VCL and VAP, ALH, and BCF. Sperm viability was unaffected. SP increased apoptotic and membrane disorder ratios, and acrosomal damage, not affecting the chromatin structure (DNA fragmentation and immaturity by SCSA), protamination (CMA3), or disulfide levels (mBBr). However, the proportion of spermatozoa with elevated free thiols (disulfide bridges reduction) significantly increased. Results support a stimulatory role of SP on thawed semen, with additional benefits from SPr. The effect of SP and especially SPr after AI should be tested since CST could be a practical test for selecting suitable ejaculates in AI centers.
33. Caracterización morfológica del caballo de la montaña asturiana
- Author
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Producción Animal, Lomillos Pérez, Juan Manuel, Armenteros Santos, José Ángel, Álvarez Espada, Aitor, Gómez Seco, Cristina, Fernández Alegre, Estela, Alonso De La Varga, Marta Elena, Producción Animal, Lomillos Pérez, Juan Manuel, Armenteros Santos, José Ángel, Álvarez Espada, Aitor, Gómez Seco, Cristina, Fernández Alegre, Estela, and Alonso De La Varga, Marta Elena
- Abstract
Las razas de caballos autóctonos españoles existentes hasta el siglo XIX, no eran adecuadas para el tiro pesado y desde que a finales del siglo XVI, Juan Jerónimo de Tiutti introdujera en nuestro país sementales flamencos, napolitanos y daneses, buscando aumentar la corpulencia de los caballos autóctonos, se ha recorrido un largo camino de intentos, fracasos y éxitos hasta conseguir actualmente la consolidación de diversas poblaciones dedicadas a la producción de caballos cuya aptitud principal es la producción de carne. Tomando como base la cabaña de yeguas autóctonas, los servicios de cría caballar del Ministerio de Defensa realizaron, a partir de los años 30 del siglo pasado, una destacada labor de mejora utilizando diversas razas, de las cuales el postier bretón resultó ser la más adecuada para formar las principales razas de carácter cárnico de España cuyos estándares se fijaron en las siguientes fechas: hispano-bretón (H-B) en 1998, en 1999 el burguete (B), en 2001 la jaca navarra (JN), en 2008 el caballo pirenaico catalán (CPC) y en 2015 el caballo del monte del País Vasco (CMPV) En el Principado de Asturias existen efectivos de caballos pesados criados en la montaña sin estar encuadrados en ninguna agrupación racial reconocida. El objetivo del presente trabajo es analizar y caracterizar racialmente dichos efectivos comprobando si existen diferencias con los efectivos H-B que puedan sustentar su consideración como nueva raza.
34. Metabolismo de la melatonina en el tracto genital masculino de rumiante. Implicación de esta hormona en la calidad seminal y fisiología de los espermatozoides = Metabolism of melatonin in the male genital tract of ruminal. Involvement of this hormone in semen quality and sperm physiology
- Author
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Martínez Pastor, Felipe, Domínguez Fernández de Tejerina, Juan Carlos, Bioquimica y Biologia Molecular, Fernández Alegre, Estela, Martínez Pastor, Felipe, Domínguez Fernández de Tejerina, Juan Carlos, Bioquimica y Biologia Molecular, and Fernández Alegre, Estela
- Abstract
El control de los procesos reproductivos es indispensable para el sector ganadero, en el caso de especies de producción, pero también es crucial para el éxito de programas de recuperación de especies silvestres y para la optimización de cría en cautividad. En la reproducción de rumiantes, uno de los factores limitantes más relevantes es la estacionalidad reproductiva ligada al fotoperiodo. La melatonina pineal es mediadora de los efectos producidos por el fotoperiodo en estas especies, influyendo sobre la actividad sexual y gonadal. El primer objetivo de esta tesis fue caracterizar la distribución de los receptores sobre la membrana plasmática de ciervo y toro mediante inmunofluorescencia. En segundo lugar, se estudió la posible afectación de la capacitación espermática en la distribución de estos receptores en la membrana celular. Además, el estudio pretende determinar el papel de la melatonina en los principales parámetros fisiológicos de espermatozoides de ciervo y toro, incluyendo el efecto sobre la capacitación.
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