RESUMEN
Preimplantation embryos often consist of a combination of euploid and aneuploid cells, suggesting that safeguards preventing the generation and propagation of aneuploid cells in somatic cells might be deficient in embryos. In somatic cells, a mitotic timer mechanism has been described, in which even a small increase in the duration of M phase can cause a cell cycle arrest in the subsequent interphase, preventing further propagation of cells that have undergone a potentially hazardously long M phase. Here, we report that cell divisions in the mouse embryo and embryonic development continue even after a mitotic prolongation of several hours. However, similar M-phase extensions caused cohesion fatigue, resulting in prematurely separated sister chromatids and the production of micronuclei. Only extreme prolongation of M phase caused a subsequent interphase arrest, through a mechanism involving DNA damage. Our data suggest that the simultaneous absence of a robust mitotic timer and susceptibility of the embryo to cohesion fatigue could contribute to chromosome instability in mammalian embryos. This article has an associated 'The people behind the papers' interview.
Asunto(s)
Aneuploidia , Blastocisto , Animales , Blastocisto/metabolismo , Inestabilidad Cromosómica , Desarrollo Embrionario/genética , Fatiga/metabolismo , Femenino , Humanos , Mamíferos , Ratones , EmbarazoRESUMEN
OBJECTIVE: To determine whether human oocytes possess a checkpoint to prevent completion of meiosis I when DNA is damaged. DESIGN: DNA damage is considered a major threat to the establishment of healthy eggs and embryos. Recent studies found that mouse oocytes with damaged DNA can resume meiosis and undergo germinal vesicle breakdown (GVBD), but then arrest in metaphase of meiosis I in a process involving spindle assembly checkpoint (SAC) signaling. Such a mechanism could help prevent the generation of metaphase II (MII) eggs with damaged DNA. Here, we compared the impact of DNA-damaging agents with nondamaged control samples in mouse and human oocytes. SETTING: University-affiliated clinic and research center. PATIENT(S): Patients undergoing ICSI cycles donated GV-stage oocytes after informed consent; 149 human oocytes were collected over 2 years (from 50 patients aged 27-44 years). INTERVENTIONS(S): Mice and human oocytes were treated with DNA-damaging drugs. MAIN OUTCOME MEASURE(S): Oocytes were monitored to evaluate GVBD and polar body extrusion (PBE), in addition to DNA damage assessment with the use of γH2AX antibodies and confocal microscopy. RESULT(S): Whereas DNA damage in mouse oocytes delays or prevents oocyte maturation, most human oocytes harboring experimentally induced DNA damage progress through meiosis I and subsequently form an MII egg, revealing the absence of a DNA damage-induced SAC response. Analysis of the resulting MII eggs revealed damaged DNA and chaotic spindle apparatus, despite the oocyte appearing morphologically normal. CONCLUSION(S): Our data indicate that experimentally induced DNA damage does not prevent PBE in human oocytes and can persist in morphologically normal looking MII eggs.
Asunto(s)
Daño del ADN , Meiosis , Oocitos/patología , Adulto , Animales , Carbazoles/toxicidad , Células Cultivadas , Etopósido/toxicidad , Femenino , Histonas/metabolismo , Humanos , Ratones , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Cuerpos Polares/patología , Pirimidinas/toxicidad , Especificidad de la Especie , Huso Acromático/patología , Tionas/toxicidad , Factores de TiempoRESUMEN
Chromosome segregation errors in mammalian embryos are common and jeopardize embryo health. Here, we perform for the first time 4-Dimensional imaging and tracking of chromosomes and centromeres through each preimplantation mitotic cell division in mouse embryos to define the normal dynamics of chromosome segregation. We show that a microtubule (MT)-dependent inward movement of chromosomes occurs at the time of nuclear envelope breakdown (NEBD), particularly in the earliest cell divisions, to position chromosomes prior to spindle assembly. Establishment of a rudimentary metaphase plate occurs immediately after NEBD, and is followed by a progressive alignment and biorientation of mitotic chromosomes. Stable end-on kinetochore-MT attachments form rapidly and attachment errors are uncommon. Altogether our data describe a rapid and efficient spindle assembly pathway that apparently minimizes the need for canonical MT attachment error correction in normally dividing embryos.
Asunto(s)
Embrión de Mamíferos/fisiología , Cinetocoros/fisiología , Microtúbulos/fisiología , Huso Acromático/fisiología , Animales , Segregación Cromosómica/fisiología , Femenino , Masculino , RatonesRESUMEN
Brominated flame retardants (BFRs), including polybrominated diphenyl ethers and hexabromocyclododecane, leach out from consumer products into the environment. Exposure to BFRs has been associated with effects on endocrine homeostasis. To test the hypothesis that in utero and lactational exposure to BFRs may affect the reproductive system of female offspring, adult female Sprague Dawley rats were fed diets formulated to deliver nominal doses (0, 0.06, 20, or 60 mg/kg/day) of a BFR dietary mixture mimicking the relative congener levels in house dust from prior to mating until weaning. Vaginal opening and the day of first estrus occurred at a significantly earlier age among offspring from the 20 mg/kg/day BFR group, indicating that the onset of puberty was advanced. Histological analysis of ovaries from postnatal day 46 offspring revealed an increase in the incidence of abnormal follicles. A toxicogenomic analysis of ovarian gene expression identified upstream regulators, including HIF1A, CREB1, EGF, the ß-estradiol, and PPARA pathways, predicted to be downregulated in the 20 or 60 mg/kg/day group and to contribute to the gene expression patterns observed. Thus, perinatal exposure to BFRs dysregulated ovarian folliculogenesis and signaling pathways that are fundamental for ovarian function in the adult.