Oxidative stress at different stages of two-step semen cryopreservation procedures in dogs

Lúcio, Cristina de Fátima; Regazzi, Fernanda Machado; Silva, L. C. G; Angrimani, Daniel de Souza Ramos; Nichi, Marcílio; Vannucchi, Camila Infantosi

doi:10.1016/j.theriogenology.2016.01.016

Cited by 58 publications

(43 citation statements)

References 49 publications

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“…However, this phenomenon was not observed in the present study: a massive effect on the sperm subpopulation displaying HMP, but not on motility parameters, was found. A positive correlation between static cells and high MMP has been reported for canine (Volpe et al, 2009;Lucio et al, 2016) and human (Donnelly et al, 2000) semen, which suggests that mitochondrial activity is not the only source of energy for sperm motility. Indeed, glycolysis in the midpiece is more likely than oxidative phosphorylation to provide the energy for sperm motility (Turner, 2003;Petrunkina and Harrison, 2011).…”

Section: Discussionmentioning

confidence: 92%

“…A significant difference in LPO was observed between cooled and frozen/thawed samples. This result is probably due to the stress of freezing procedure, including the manipulation, leading to an increase in ROS, which is an inducer of LPO and therefore an increase in ROS is another factor that indicates that a cell has suffered a stress (Donnelly et al, 2000;Câmara and Guerra, 2008;Kim et al, 2010a;Lucio et al, 2016;Ortega-Ferrusola et al, 2017).…”

Section: Discussionmentioning

confidence: 99%

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Effects of the cryopreservation process on dog sperm integrity

et al. 2020

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Sperm cryopreservation has become an indispensable tool in reproductive biology. However, frozen/thawed semen has a short lifespan due to loss of sperm cell integrity. To better understand which sperm cell structures are compromised by the cryopreservation process and apoptosis markers, the sperm of five healthy mature dogs was analyzed in this study. Analysis was performed after collection, cooling, and thawing via computer assisted sperm analyzer (CASA) and evaluation of membrane fluidity and permeability, phosphatidylserine translocation (Annexin V), membrane integrity, mitochondrial membrane potential, membrane lipid peroxidation (LPO) and activity of the apoptotic markers caspases 3 and 7 by flow cytometry. Cryopreservation decreased total and progressive motility and the percentage of rapid sperm (P < 0.01). Damage to sperm cells was confirmed by Annexin V (P < 0.01), indicating that capacitation-like changes were induced by the cryopreservation procedures. An increase in sperm membrane fluidity was also noted in frozen/thawed samples (P < 0.01). Plasma and acrosomal cell membranes were affected (P < 0.01), with decreases in the subpopulation displaying high membrane potential (P < 0.01). Membrane LPO was increased in thawed sperm compared to cooled sperm (P < 0.05) but was not different from that in fresh sperm. No differences were observed in caspase 3 and 7 activity after cooling, freezing, or thawing. In conclusion, total and progressive motility, plasma membrane integrity and mitochondrial membrane potential suffered from the deleterious effects caused by cryopreservation, unlike the activity of caspases that remained stable during the freezing process.

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Section: Discussionmentioning

confidence: 92%

Section: Discussionmentioning

confidence: 99%

Effects of the cryopreservation process on dog sperm integrity

et al. 2020

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“…Lucio et al (2016a) described that the changes in spermatozoa by the effects of cryopreservation begins during chilled and continues until thawed mainly by production of free radicals, which may occur due to mitochondrial dysfunctions, possibly in our study, by reductive stress (Korge and Weiss, 2014). Despite our results showed no changes in TBARS, it was possible to observe this injurious relationship between mitochondria and oxidative stress in the negative correlation of TBARS and high mitochondrial potential.…”

Section: Discussionmentioning

confidence: 51%

The use of reduced glutathione (GSH) as antioxidant for cryopreserved sperm in dogs

Angrimani

Nichi

Brito

et al. 2018

Arq. Bras. Med. Vet. Zootec.

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The aim of this study was to evaluate the effect of supplementation with different concentrations of reduced glutathione GSH (0; 5; 7.5; 10mM) in the extender for cryopreservation in dogs with evaluations performed after glycerolization (chilled) and thawing (thawed). For this purpose, we used 8 dogs and two semen collections were performed in a weekly interval, totaling 16 semen samples. The sperm were analyzed by automatic sperm motility (CASA) and flow cytometry analysis of mitochondrial potential (JC1 dye) and membrane/acrosome integrity (FITC-PI dyes). We evaluated subjectively the membrane and acrosome integrity, mitochondrial activity and DNA integrity. Seminal plasma was evaluated for lipid peroxidation (TBARS concentration). Chilled and thawed samples supplemented with 7.5 and 10mM of GSH had lower percentage of sperm with high (DAB -Class I) and medium (DAB -Class II) mitochondrial activity. And 10mM of GSH had higher percentage of low mitochondrial activity (DABClass III). Moreover, thawed samples of 10mM of GSH had high DNA fragmentation rates. Probably by a reductive stress effect on mitochondria which lead to an increase in reactive oxygen species, and a mitochondrial malfunction.Keywords: reactive oxygen species, cryopreservation, antioxidant, canine RESUMOO objetivo deste estudo foi avaliar o efeito da suplementação com diferentes concentrações de glutationa reduzida 5; 7,5; 10mM)

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“…In fact, it is important that the acrosome membrane remains intact for the successful binding to the oocyte, in order to avoid the loss of acrosomal enzymes (LUCIO et al, 2016). Therefore, the binding test to the perivitelline membrane not only indicates sperm fertilizing capacity, but also indirectly assures acrosome membrane integrity.…”

Section: Discussionmentioning

confidence: 99%

Sperm-binding to the perivitelline membrane of chicken egg yolk as a functional test for sperm evaluation in dogs

Brito¹,

Losano²,

Angrimani³

et al. 2017

Braz. J. Vet. Res. Anim. Sci.

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During fertilization, spermatozoa interact with the zona pellucida (ZP) through the binding between the acrosome and proteins 2 and 3 (ZP2 and ZP3). The perivitelline membrane of chicken egg yolk is homologous to the mammalian ZP3, which allows the binding of sperm of several species. The aim of this study was to standardize and evaluate the efficiency of sperm-binding to the perivitelline membrane of chicken eggs as a functional method for canine semen evaluation. For this purpose, nine post-thaw sperm samples were used, which were divided into two aliquots: the first was kept in water bath at 37ºC (live sample) and the second was submitted to cold shock to induce cellular damage (dead sample). The two aliquots were mixed on five proportions, corresponding to 0%, 25%, 50%, 75%, and 100% of viable cells, and the binding test was performed by analyzing the number of spermatozoa bonded to the perivitelline membrane by means of computerized assessment of sperm motility (CASA) or conventional microscopy. Additionally, samples were submitted to sperm motility analysis, evaluation of plasmatic and acrosomal membrane integrity, and sperm mitochondrial activity. The sperm-binding test to the perivitelline membrane of chicken egg yolk was considered a feasible sperm analysis test for both fertilizing capacity and overall sperm attributes evaluation, mainly when the analysis is performed by a conventional microscope, which expands its practicality to the majority of canine reproduction laboratories. Keywords: Sperm cryopreservation. Egg yolk. Sperm analysis. Perivitelline membrane. Canine. ResumoDurante a fecundação, os espermatozoides interagem com a zona pelúcida (ZP) por meio da ligação entre o acrossomo e as proteínas 2 e 3 (ZP2 e ZP3). A membrana perivitelínica da gema de ovo de galinhas é homóloga à ZP3 de mamíferos, possibilitando a ligação espermática de diversas espécies. Este trabalho padronizou e avaliou a eficiência do teste de ligação espermática à membrana perivitelínica da gema de ovo de galinhas como avaliação funcional do sêmen de cães. Para tal, foram utilizadas nove amostras seminais previamente criopreservadas. Cada amostra foi dividida em duas alíquotas: a primeira foi mantida em banho-maria à 37ºC (vivos) e a segunda submetida a choque térmico com o intuito de induzir dano celular (mortos). As duas alíquotas foram misturadas, correspondendo a 0, 25, 50, 75 e 100% de células viáveis. As amostras foram avaliadas quanto ao número de espermatozoides ligados à membrana perivitelínica por meio da análise computadorizada da motilidade (CASA) ou microscopia convencional. Ademais, as amostras foram avaliadas quanto à motilidade espermática, integridade das membranas acrossomal e plasmática e atividade mitocondrial espermática. O teste de ligação espermática à membrana perivitelínica de ovos de galinha foi considerado um teste de análise seminal exequível tanto para avaliar a capacidade fecundante dos espermatozoides como atributos seminais gerais, especialmente quando realizado em microscopia convenci...

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Oxidative stress at different stages of two-step semen cryopreservation procedures in dogs

Cited by 58 publications

References 49 publications

Effects of the cryopreservation process on dog sperm integrity

Effects of the cryopreservation process on dog sperm integrity

The use of reduced glutathione (GSH) as antioxidant for cryopreserved sperm in dogs

Sperm-binding to the perivitelline membrane of chicken egg yolk as a functional test for sperm evaluation in dogs

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