Karina Cañón-Beltrán, Encina M González, Alfonso Gutiérrez-Adán, Y. N. Cajas, Ekaitz Agirregoitia, Cláudia Lima Verde Leal, Dimitrios Rizos, Serafín Pérez-Cerezales, Ministerio de Ciencia e Innovación (España), Canon-Beltran, K, Cajas, YN, Cerezales, S, Leal, CLV, Agirregoitia, E, Gutierrez-Adan, A, Gonzalez, EM, Rizos, D, Canon-Beltran, K [0000-0002-0279-0857], Cajas, YN [0000-0001-9791-6733], Cerezales, S [0000-0003-2119-6228], Leal, CLV [0000-0002-8180-5825], Agirregoitia, E [0000-0001-7987-4963], Gutierrez-Adan, A [0000-0001-9893-9179], Gonzalez, EM [0000-0002-6217-866X], and Rizos, D [0000-0001-6813-3940]
In vitro culture can alter the development and quality of bovine embryos. Therefore, we aimed to evaluate whether nobiletin supplementation during EGA improves embryonic development and blastocyst quality and if it affects PI3K/AKT signaling pathway. In vitro zygotes were cultured in SOF + 5% FCS (Control) or supplemented with 5, 10 or 25 µM nobiletin (Nob5, Nob10, Nob25) or with 0.03% dimethyl-sulfoxide (CDMSO) during minor (2 to 8-cell stage; MNEGA) or major (8 to 16-cell stage; MJEGA) EGA phase. Blastocyst yield on Day 8 was higher in Nob5 (42.7 ± 1.0%) and Nob10 (44.4 ± 1.3%) for MNEGA phase and in Nob10 (61.0 ± 0.8%) for MJEGA phase compared to other groups. Mitochondrial activity was higher and lipid content was reduced in blastocysts produced with nobiletin, irrespective of EGA phase. The mRNA abundance of CDK2, H3-3B, H3-3A, GPX1, NFE2L2 and PPARα transcripts was increased in 8-cells, 16-cells and blastocysts from nobiletin groups. Immunofluorescence analysis revealed immunoreactive proteins for p-AKT forms (Thr308 and Ser473) in bovine blastocysts produced with nobiletin. In conclusion, nobiletin supplementation during EGA has a positive effect on preimplantation bovine embryonic development in vitro and corroborates on the quality improvement of the produced blastocysts which could be modulated by the activation of AKT signaling pathway., This work was funded by the Spanish Ministry of Science and Innovation (PID2019-111641RB-I00 to D.R. and RTI2018-093548-B-I00 to A.G.-A). Y.N.C. was supported by a predoctoral fellowship from the Secretaría Nacional de Educación Superior, Ciencia, Tecnología e Innovación (Convocatoria abierta 2017, SENESCYT-Ecuador). C.L.V.L. was supported by a BPE grant from Fundação de Amparo à Pesquisa do Estado de São Paulo, Brazil (FAPESP #2017/20339-3).