Effects of supplemental antioxidants on in vitro fertility measures for cryopreserved boar spermatozoa

André Furugen Cesar de Andrade; Kayode Balogun; Zoltan Machaty; Robert Victor Knox

doi:10.1016/j.theriogenology.2023.01.025

Effects of supplemental antioxidants on in vitro fertility measures for cryopreserved boar spermatozoa

Theriogenology. 2023 Apr 1:200:33-42. doi: 10.1016/j.theriogenology.2023.01.025. Epub 2023 Feb 2.

Authors

André Furugen Cesar de Andrade¹, Kayode Balogun², Zoltan Machaty², Robert Victor Knox³

Affiliations

¹ Department of Animal Reproduction, School of Veterinary Medicine and Animal Science, University of São Paulo, Pirassununga, São Paulo, Brazil; Department of Animal Sciences, College of Agricultural, Consumer & Environmental Sciences, University of Illinois at Urbana-Champaign, USA. Electronic address: andrefc@usp.br.
² Department of Animal Sciences, College of Agriculture, Purdue University, West Lafayette, IN, USA.
³ Department of Animal Sciences, College of Agricultural, Consumer & Environmental Sciences, University of Illinois at Urbana-Champaign, USA.

PMID: 36739670
DOI: 10.1016/j.theriogenology.2023.01.025

Abstract

This work aims to evaluate how supplementing a commercial freezing media with butylated hydroxytoluene (BHT), or reduced glutathione (GSH), or their combination affected in-vitro measures of boar sperm after cryopreservation. One ejaculate was collected from 30 high-fertility boars in a weekly collection rotation. Samples were diluted 1:1 in an extender and cooled before overnight shipping at 17 °C to the freezing lab. On arrival, samples were split into the treatments with the following additions before cryopreservation; 1) semen without additional antioxidants (Control), 2) semen with 1 mM BHT, 3) semen with 2 mM GSH, and 4) semen with 1 mM BHT+2 mM GSH. Semen was evaluated for motility kinetics at 30, 120, and 240 min after thawing. Flow cytometry assessments were performed at 60 min after thawing. At all-time points evaluated, total and progressive motility were greater (P ≤ 0.05) in semen cryopreserved with GSH than in Control. No (P > 0.05) differences between Control and other treatment groups were observed in viability, or acrosomal and mitochondrial membrane integrity; however, the proportion of capacitated spermatozoa were reduced (by -21.17%) in semen treated with BHT + GSH compared to Control (P ≤ 0.05). In contrast, there was a higher (P ≤ 0.05, +21.18%) superoxide anion production in the Control than in the BHT + GSH. For IVF, semen cryopreserved with both antioxidants (BHT + GSH) had a negative (P < 0.05) impact on fertilization rate (-54.11%) compared to Control. However, for the blastocysts rate, there were more (+22.75%) blastocysts (P ≤ 0.05) for BHT compared to Control. These results indicate that commercial media supplemented with GSH increased motility but impaired in vitro fertilization rate. On the other hand, media supplemented with BHT improved the in vitro fertilizing ability of the frozen-thawed sperm cells. Therefore, we suggest the supplementation with 1 mM of BHT in the formula of commercial freezing media used in the present experiment.

Keywords: Boar sperm; Cryopreservation; IVF; Motility; Viability.

MeSH terms

Animals
Antioxidants* / pharmacology
Butylated Hydroxytoluene / pharmacology
Cryopreservation / methods
Cryopreservation / veterinary
Fertility
Male
Semen
Semen Preservation* / methods
Semen Preservation* / veterinary
Sperm Motility
Spermatozoa
Swine

Substances

Antioxidants
Butylated Hydroxytoluene