Cell cycle duration at the time of maternal zygotic transition for in vitro produced bovine embryos: effect of oxygen tension and transcription inhibition.
Early embryonic cleavages are mostly regulated by maternal components then control of development progressively depends on newly synthesized zygotic products. The timing of the first cleavages is a way to assess embryo quality. The goal of this study was to evaluate the duration of the fourth cell cycle, at the time of maternal-to-zygotic transition (MZT) in in vitro-produced bovine embryos by means of cinematographic analysis. We found that 75% of the embryos displayed a long fourth cycle (43.5 +/- 5.4 h) whereas the remaining embryos had a very short fourth cell cycle (8.9 +/- 2.9 h). Both groups did not differ in cleavage rhythm up to the eight-cell stage and timing of cavitation and blastocyst expansion was identical. However, embryos with a short fourth cell cycle had a better blastocyst rate than embryos with a long cycle (59% versus 38%, P < 0.01). Total cell number, inner cell mass (ICM):total cell ratio, and hatching rate were identical for blastocysts produced from embryos with either a long or a short fourth cell cycle. In a second experiment, we showed that increasing the oxygen tension, from 5% to 20%, decreased the percentage of embryos with a short fourth cell cycle, from 25% to 11% (P < 0.01), indicating that suboptimal culture conditions can influence the length of this cycle. Finally, we investigated whether fourth cell cycle duration could be influenced by transcription inhibition. With alpha-amanitin added at 18 h postinsemination (HPI), cleavage was reduced (66% versus 79%) and, at 70 HPI, the 9- to 16-cell rate increased (50% versus 25%) concomitantly with a 5- to 8-cell rate decrease (16% versus 47%). A similar pattern was observed when the drug was added at 6 HPI or 42 HPI but not at 0 HPI. Cinematographic analysis revealed that alpha-amanitin increased the first cell cycle duration whereas the second and third cell cycles were not affected. With the drug, one third of the embryos could develop up to the 9- to 16-cell stage and they all had a short fourth cell cycle (11.2 +/- 3.7 h) with a good synchrony of cleavage between blastomeres. These results suggest that duration of the fourth cell cycle of bovine embryo, during the MZT, is under a zygotic transcriptional control that can be affected by oxidative conditions.
Lequarre
A S
AS
Unité Vétérinaire, Institut des Sciences de la Vie, Université Catholique de Louvain, Louvain-la-Neuve B-1348, Belgium. Lequarre@vete.ucl.ac.be
Marchandise
J
J
Moreau
B
B
Massip
A
A
Donnay
I
I
D000546:Amanitins / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D002452:Cell Count; D002453:Cell Cycle / Q000187:drug effects / Q000502:physiology*; D002467:Cell Nucleus / Q000502:physiology; D004622:Embryo, Mammalian / Q000502:physiology*; D005314:Embryonic and Fetal Development / Q000502:physiology; D005260:Female; D005307:Fertilization in Vitro; D018496:Hyperoxia / Q000503:physiopathology; D066298:In Vitro Techniques; D018715:Microscopy, Video; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010100:Oxygen / Q000502:physiology*; D011247:Pregnancy; D012319:RNA Polymerase II / Q000037:antagonists & inhibitors; D017382:Reactive Oxygen Species / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D015053:Zygote / Q000187:drug effects / Q000502:physiology*
D000546:Amanitins; D019384:Nucleic Acid Synthesis Inhibitors; D017382:Reactive Oxygen Species; D012319:RNA Polymerase II; D010100:Oxygen