Effects of α-tocopherol and freezing rates on the quality and heterologous in vitro fertilization capacity of stallion sperm after cryopreservation.
Abstract: The effects of supplementation of α-tocopherol and different freezing rates (FRs) on the ability of stallion sperm to fertilize bovine oocytes with intact zona pellucida were investigated, in an attempt to develop a model to assess cryopreserved sperm function. Semen was obtained from four purebred Lusitano stallions (n = 4). Each ejaculate was subjected to cryopreservation with a commercial extender (Ghent, Minitub Iberia, Spain), without any supplementation (control) or supplemented with 2-mM α-tocopherol. The semen was exposed to two different FRs between 5 °C and -15 °C: slow (5 °C/min) and moderate (10 °C/min). After thawing, the viability (SYBR®-14 and propidium iodide [PI]), mitochondrial membrane potential (JC-1, 5,5',6,6'-tetrachloro-1,1',3,3'tetraethylbenzimidazolyl carbocyanine iodine) and membrane lipid peroxidation (C11-BODIPY(581/591)) of each sample were determined by flow cytometry. Moreover, the heterologous IVF rate was measured to evaluate the fertilization capacity of postthaw semen in the four different treatments. For both extenders, the viability was higher for spermatozoa cooled slowly (39.40 ± 2.17 vs. 17.59 ± 2.25-control; 31.96 ± 2.19 vs. 11.46 ± 1.34-Tocopherol; P < 0.05). The α-tocopherol extender improved (P < 0.05) postthaw lipid peroxidation (10.28 ± 0.70 vs. 15.40 ± 0.95-slow FR; 10.14 ± 0.40 vs. 13.48 ± 0.34-moderate FR); however, it did not improve viability and mitochondrial membrane potential. Regarding the IVF rate, in the moderate FR, α-tocopherol supplementation reported a higher percentage of IVF (20.50 ± 2.11; P < 0.05), comparing with the control (14.00 ± 1.84). Regarding the slow FR, no significance differences were observed for percentage of IVF between the two extenders and the FRs. However, it seems that the α-tocopherol supplementation improved the IVF rate. In conclusion, this research reported that bovine oocytes intact zona pellucida can be used to evaluate the quality of postthaw stallion semen and α-tocopherol supplementation in the stallion freezing extender might exert a protective effect against oxidative damage during heterologous IVF.
Copyright © 2016 Elsevier Inc. All rights reserved.
Publication Date: 2016-03-19 PubMed ID: 27125690DOI: 10.1016/j.theriogenology.2016.03.019Google Scholar: Lookup
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Summary
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The research article focuses on how the addition of α-tocopherol and different freezing rates (FRs) can influence the quality and potential to fertilize of cryopreserved stallion sperm.
Overview of the Research
In this study, the researchers intended to establish a model to assess the functionality of cryopreserved sperm. They used the semen collected from four purebred Lusitano stallions for this experiment. Their control group used regular freezing extender without any supplements while in the other group they added 2-mM α-tocopherol to the freezing extender.
The Experiment Process and Its Results
- The researchers subjected the semen to two different freezing rates between 5 -15 degrees Celsius, one being slow (5 degrees/minute) and the other moderate (10 degrees/minute).
- After thawing the samples, they were inspected for their viability, mitochondrial membrane potential, and membrane lipid peroxidation. These parameters were assessed by flow cytometry.
- The results showed an improvement on viability for semen which had been cooled in a slower rate. However, it should be noted that the introduction of α-tocopherol did not enhance the semen’s viability and mitochondrial membrane potential.
- In terms of membrane lipid peroxidation, the semen which was introduced to α-tocopherol showed improved post-thaw conditions irrespective of the freezing rate.
- They also did a detailed analysis on the rate of heterologous in vitro fertilization (IVF). Significantly higher IVF rates were recorded when α-tocopherol was supplemented, especially when the semen was subjected to moderate freezing rates.
- No significant differences were noticed in the IVF rates between the two extenders and freezing rates for semen which was frozen slowly. But still, a subtle improvement in IVF rate was noted with α-tocopherol supplementation
Conclusion Derived From the Research
- The researchers concluded that bovine oocytes with intact zona pellucida could be effectively used to gauge the quality of post-thaw stallion semen.
- Additionally, they inferred that inclusion of α-tocopherol in the stallion freezing extender might yield protective effects against oxidative damage during heterologous in vitro fertilization.
Cite This Article
APA
de Vasconcelos Franco JS, Faheem M, Chaveiro A, Moreira da Silva F.
(2016).
Effects of α-tocopherol and freezing rates on the quality and heterologous in vitro fertilization capacity of stallion sperm after cryopreservation.
Theriogenology, 86(4), 957-962.
https://doi.org/10.1016/j.theriogenology.2016.03.019 Publication
Researcher Affiliations
- Animal Reproduction, Department of Agrarian Sciences, University of the Azores, CITA-A, Angra do Heroísmo, Portugal. Electronic address: joanna@uac.pt.
- Animal Reproduction, Department of Agrarian Sciences, University of the Azores, CITA-A, Angra do Heroísmo, Portugal; Animal Production Department, Faculty of Agriculture, Cairo University, Giza, Egypt.
- Animal Reproduction, Department of Agrarian Sciences, University of the Azores, CITA-A, Angra do Heroísmo, Portugal.
- Animal Reproduction, Department of Agrarian Sciences, University of the Azores, CITA-A, Angra do Heroísmo, Portugal.
MeSH Terms
- Animals
- Antioxidants / pharmacology
- Cryopreservation / veterinary
- Fertilization in Vitro / veterinary
- Freezing
- Horses / physiology
- Male
- Semen Preservation / veterinary
- Sperm-Ovum Interactions / drug effects
- Spermatozoa / drug effects
- alpha-Tocopherol / pharmacology
Citations
This article has been cited 3 times.- Barcelona B, Ramos Z, Viñoles C, Rodríguez-Osorio N, Báez F. Season-specific effects of α-tocopherol supplementation during bovine oocyte in vitro maturation on embryo yield and quality. Anim Reprod 2025;22(2):e20240136.
- Loureiro KC, Lima-Verde IB, Johannisson A, Ntallaris T, Jager A, Štěpánek P, da Costa Mendonça M, Severino P, Morrell JM. Effects of cashew gum and nanoparticles on cooled stallion semen. Acta Vet Scand 2020 Jun 18;62(1):31.
- Rahman MS, Kang KH, Arifuzzaman S, Pang WK, Ryu DY, Song WH, Park YJ, Pang MG. Effect of antioxidants on BPA-induced stress on sperm function in a mouse model. Sci Rep 2019 Jul 22;9(1):10584.
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