Relative transcript level of BMP15, BAX and CASP3 with qRT-PCR in vitrified equine immature cumulus-oocytes complexes

• Autor(a) principal: Polenz, Mauro Flores
• Data de Publicação: 2022
• Outros Autores: Pereira, Gabriel Ribas, Rissi, Vitor Braga, Bertolin, Kalyne, Gonçalves, Paulo Bayard, Augusto, Geovani Celso, Rechsteiner, Sandra Mara Fiala
• Tipo de documento: Artigo
• Idioma: eng
• Título da fonte: Semina. Ciências Agrárias (Online)
• Texto Completo: https://ojs.uel.br/revistas/uel/index.php/semagrarias/article/view/44029
• topic: Bcl-2-associated X protein; Bone morphogenetic protein 15; Gene expression; Caspase 3; Horse.; Proteína X associada a Bcl-2; Proteína morfogenética óssea 15; Expressão genética; Caspase 3; Equinos.

Research focused on female gamete vitrification has increased attention to develop a reliable cryopreservation method to preserve immature equine oocytes. Despite the intensive implementation of biotechnological procedures for horse breeding, vitrification of immature equine cumulus-oocyte complexes (COCs) remain to be clearly elucidated. We aimed to determine the relative transcript level of target genes Bone morphogenetic protein 15 (BMP15); Bcl-2-associated X protein (BAX); and Caspase 3 (CASP3) in equine COCs prior to and after vitrification. Ovarian follicles were aspirated from ovaries collected from an abattoir. A total of 240 COCs were collected and distributed into vitrified COCs (VIT, n=120) and non-vitrified (Non-VIT, n=120) groups. Then, COCs were preserved and relative transcript expressions of BMP15, BAX, CASP3 were measured and normalized against GAPDH performed by qRT-PCR. In addition, 38 COCs were evaluated to assess chromatin configuration of germinal vesicle stage prior and after vitrification by exposure to 10 ug/ml of bisbenzimide. A difference was observed in the COCs mRNA level of abundance for the BAX gene between the VIT (2.05 ± 0.47) and (0.85 ± 0.08) Non-VIT groups. There was no difference in mRNA relative transcript level of CASP3 and BMP15 in Non-VIT (0.63 ± 0.20 and 1.55 ± 0.73, respectively) compared to VIT (0.64 ± 0.01 and 2.84 ± 2.20, respectively) equine COCs. All COCs where considered at immature stage of development even though COCs in Non-VIT group showed higher condensed chromatin configuration compared to VIT (100% vs 60.7%, respectively). We demonstrate that BMP15 and CASP3 are detected in VIT and Non-VIT immature COCs. In conclusion, BAX is expressed highly in vitrified immature equine COCs and indicates that activation of apoptosis signaling cascades in cells exposed to vitrification.