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Effects of antifreeze protein from Lolium perenne L. (LpAFP) in the vitrification of in vitro-produced bovine embryos

Published online by Cambridge University Press:  27 June 2023

R.A. Silva Júnior Open the ORCID record for R.A. Silva Júnior [Opens in a new window] ,
R. Desenzi Open the ORCID record for R. Desenzi [Opens in a new window] ,
M.M.S. Ramires ,
A.F. Souza ,
M.A.M. Donato ,
C.A. Peixoto ,
T. Nascimento ,
C.C. Bartolomeu  and
A.M. Batista Open the ORCID record for A.M. Batista [Opens in a new window]
R.A. Silva Júnior*
Affiliation:
Laboratório de Biotécnicas Aplicadas à Reprodução, Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Recife, Pernambuco, Brazil
R. Desenzi
Affiliation:
Laboratório de Biotécnicas Aplicadas à Reprodução, Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Recife, Pernambuco, Brazil
M.M.S. Ramires
Affiliation:
Departamento de Zootecnia, Universidade Federal Rural de Pernambuco, Recife, Pernambuco, Brazil
A.F. Souza
Affiliation:
Departamento de Zootecnia, Universidade Federal Rural de Pernambuco, Recife, Pernambuco, Brazil
M.A.M. Donato
Affiliation:
Departamento de Histologia e Embriologia, Universidade Federal de Pernambuco, Recife, Pernambuco, Brazil
C.A. Peixoto
Affiliation:
Laboratory of Ultrastructure, Aggeu Magalhães Institute (IAM), Recife, PE, Brazil; National Institute of Science and Technology on Neuroimmunomodulation (INCT-NIM, CNPq), Oswaldo Cruz Foundation, Rio de Janeiro, Brazil
T. Nascimento
Affiliation:
Departamento de Botânica, Universidade Federal de Pernambuco, Recife, PE, Brazil
C.C. Bartolomeu
Affiliation:
Laboratório de Biotécnicas Aplicadas à Reprodução, Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Recife, Pernambuco, Brazil
A.M. Batista*
Affiliation:
Laboratório de Biotécnicas Aplicadas à Reprodução, Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Recife, Pernambuco, Brazil
*
Corresponding authors: R.A. Silva Júnior; Email: artur_rjs@hotmail.com; A.M. Batista; Email: andre.batista@ufrpe.br
Corresponding authors: R.A. Silva Júnior; Email: artur_rjs@hotmail.com; A.M. Batista; Email: andre.batista@ufrpe.br
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Summary

In the present study, the cryoprotective effects of Lolium perenne antifreeze protein (LpAFP) on the vitrification of bovine embryos were evaluated. In vitro-produced blastocysts were divided into two groups: the control group (CG) without the addition of LpAFP and the treatment group (TG) with the addition of 500 ng/ml of LpAFP in the equilibrium and vitrification solution. Vitrification was carried out by transferring the blastocysts to the equilibrium solution [7.5% ethylene glycol (EG) and 7.5% dimethyl sulfoxide (DMSO)] for 2 min and then to the vitrification solution (15% EG, 15% DMSO and 0.5M sucrose). The blastocysts were deposited on a cryotop device and submerged in liquid nitrogen. Warming was carried out in three steps in solutions with different sucrose concentrations (1.0, 0.5, and 0.0 M, respectively). Embryos were evaluated for re-expansion/hatching, the total cell count, and ultrastructural analysis. There was no significant difference in the re-expansion rate 24 h after warming; however, there was variation (P < 0.05) in the hatching rate in the TG and the total number of cells 24 h after warming was higher in the TG (114.87 ± 7.24) when compared with the CG (91.81 ± 4.94). The ultrastructural analysis showed changes in organelles related to the vitrification process but, in the TG, there was less damage to mitochondria and rough endoplasmic reticulum compared with the CG. In conclusion, the addition of 500 ng/ml of LpAFP during the vitrification of in vitro-produced bovine embryos improved the hatching rate and total cell number of blastocysts after warming and mitigated intracellular damage.

Keywords

Assisted reproductionBlastocystsCryopreservationCryotopIVF
Type
Research Article
Information
Zygote , Volume 31 , Issue 5 , October 2023 , pp. 468 - 474
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Copyright
© The Author(s), 2023. Published by Cambridge University Press

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