Impact of cryopreservation protocols (one- and two-step) on boar semen quality at 5 degrees C and post-thawing

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Monteiro, Matheus Saliba ; Torres, Mariana Andrade ; Passarelli, Marina da Silva ; Martins, Matheus Passini ; Ravagnani, Gisele Mouro ; Papa, Frederico Ozanam ; Alvarenga, Marco Antonio ; Dell'Aqua Junior, Jose Antomo ; Yasui, George Shigueki ; Martins, Simone Maria Massami Kitamura ; de Andrade, Andre Furugen Cesar

• Tipo de documento: Artigo Científico
• Fonte: Animal Reproduction Science; v. 247, p. 9-pg., 2022-10-20.

Resumo

The two-step protocol (2 S) is currently used for boar semen cryopreservation. In this method, the cryoprotectant penetrant agents (CPAs) are added at 5 degrees C to reduce the toxicity of CPAs. An alternative is the one-step protocol (1 S), which is easier, cheaper, and reduces the necessity of equipment, but could increase the toxicity of CPAs. Currently, there are no studies that compared both protocols for boar semen cryopreservation. This experiment aimed to study the effect of cryopreservation protocol (1 S vs 2 S) on boar spermatozoa. In the one-step protocol, after centrifugation, the spermatozoa pellet was resuspended at 17 degrees C in the extender containing CPAs to achieve a concentration of 1 x 109 spermatozoa/mL and then submitted to cryopreservation. For the two-step protocol, the sperm pellet was resuspended in fraction A at 17 degrees C to achieve a concentration of 1.5 x 109 spermatozoa/ mL, and then allowed to cool to 5o C before fraction B with CPA was added to the sample to achieve a final concentration of 1 x 109 spermatozoa/mL and followed by freezing. The cryopreservation protocol did not impact total motility at 5 degrees C (1 S: 78.5 % vs 2 S: 79 %, p > 0.05). After thawing, the two-step protocol improved (p < 0.05) total (1 S: 18.2 % vs 2 S: 29.5 %) and progressive motility (1 S: 9 % vs 2 S: 15%). Further, the 2 S protocol increased (p < 0.05) the percentage of rapid spermatozoa (1 S: 8.7 % vs 2 S: 14.6 %) and spermatozoa with intact plasma and acrosomal membrane (IAIP) (1 S: 40.5 % vs 2 S: 61.5 %), and increased (p < 0.05) live sperm cells with high mitochondrial potential (MHIP) (1 S: 42.9 % vs 2 S: 60 %). The boar semen cryopreservation method (TRT) did not (p > 0.05) alter membrane lipid disorder, lipid peroxidation, and superoxide anion. Thus, the best method for boar semen cryo-preservation is the two-step protocol. (AU)

• Processo FAPESP: 17/16987-0 - Validação de melhor crioprotetor intracelular para a metodologia One Step de criopreservação de sêmen suíno seguido de comparação com a metodologia Two Steps
• Beneficiário: Matheus Saliba Monteiro
• Modalidade de apoio: Bolsas no Brasil - Iniciação Científica
• Processo FAPESP: 16/24690-4 - Assinaturas metabolômicas da influência do holding time sobre o aumento da criotolerância dos espermatozóides suínos
• Beneficiário: André Furugen Cesar de Andrade
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 17/10821-2 - Avaliação dos efeitos do tempo de equilíbrio nos protocolos de criopreservação de sêmen suíno: um estudo fisiológico e morfofuncional.
• Beneficiário: Marina da Silva Passarelli Zanatta
• Modalidade de apoio: Bolsas no Brasil - Mestrado