168 Improvement of in vivo-produced bovine embryo competence by using a bovine recombinant FSH in a superovulation protocol
L. Rodriguez-Alvarez A , M. A. Gutiérrez-Reinoso B , C. J. Aguilera A , F. Navarrete A , J. Cabezas A , O. Sanchez C , I. Cabezas A , F. O. Castro A , M. García-Herreros D and Y. Wong AA Universidad de Concepción, Chillán, Chile
B Universidad Técnica de Cotopaxi, Latacunga, Ecuador
C Centro de Biotecnología y Biomedicina Spa, Concepción, Chile
D Instituto Nacional de Investigaçao Agrária e Veterinária, Santarém, Portugal
Reproduction, Fertility and Development 34(2) 322-323 https://doi.org/10.1071/RDv34n2Ab168
Published: 7 December 2021
© 2022 The Author(s) (or their employer(s)). Published by CSIRO Publishing on behalf of the IETS
Superovulation (SOV) protocols are crucial to improve in vivo embryo production in cattle. FSH induces the recruitment and growth of follicles, guaranteeing the competence of oocytes, which is reflected in embryo quality. The aim of this study was to assess the quality of in vivo-produced bovine embryos by using recombinant FSH (bscFSH-r) vs. conventional FSH (NIH-FSH-p) superovulation (SOV) protocols. Embryo quality was assessed by the analysis of the expression of developmentally important genes and by in vitro development up to Day 11. Embryos were in vivo-produced from Red Angus cows after NIH-FSH-p (Group (G)1: FSH from purified pig pituitary extract) and bscFSH-r (G2: Recombinant FSH) derived SOV protocols. Regarding G1 SOV, a conventional protocol was applied (Day 0: intravaginal P4 device (CIDR: 1.38 g) + 2.5 mg intramuscular (IM) oestradiol benzoate E2B + 100 mg P4 (IM); Day 4: 280 mg NIH-FSH-p/12-h intervals/4 day/8 decreasing doses; Day 6: fifth and sixth NIH-FSH-p dose + two PGF2α IM doses (500 µg of D-cloprostenol each); Day 7: CIDR removal at the seventh NIH-FSH-p dose application. Same protocol was applied in G2 with modifications (170 µg of bscFSH-r/24-h intervals/4 day/4 decreasing doses). Embryo quality and classification were performed by morphological criteria according to the IETS guidelines. Collected grade I embryos were kept in culture (G1: n = 26; G2: n = 33) or used for gene expression analysis (G1: n = 10; G2: n = 11). Embryos were individually cultured in synthetic oviductal fluid (SOF) medium or lysed and then subjected to reverse transcription using the Cells-to-cDNA™ II Kit (ThermoFisher Scientific). The diameter of embryos in culture was recorded on Days 7, 9, and 11. Expression analyses of OCT4, SOX2, NANOG, FGF4, CDX2, IFNtau, BAX, and BCL2 were performed by real-time PCR using the 2−ΔΔCT method. In all cases, samples with a cycle threshold (Ct) ≤35 were considered for analysis. The Ct values were normalised using the geometric mean of the expression of GAPDH and ACTB. The Wilcoxon non-parametric test was used for data analysis. At Days 7 and 9, embryos had a similar diameter (P > 0.05). However, embryos from G2 had a greater diameter at Day 11 (P = 0.003) and showed linear growth from Day 7 to 11. SOX2, NANOG, and FGF4 were amplified in fewer than 60% of samples regardless of the experimental group; therefore, they were not included in the expression analysis. IFNtau and BAX were overexpressed in embryos from G2 (P < 0.05) with a fold change of 16 and 1.3, respectively. However, no statistical differences were detected regarding OCT4, CDX2, BCL2, and BCL2/BAX expression ratio (P > 0.05). In conclusion, embryos produced using a SOV protocol that includes bscFSH-r seems to have better quality, as reflected by the higher diameter and expression level of IFNtau, which would influence maternal recognition of the pregnancy.
This research was partially supported by Fondef ID18I10082-ANID 21201280.
