The CBA mouse as a model for age-related aneuploidy in man: studies of oocyte maturation, spindle formation and chromosome alignment during meiosis

Abstract

To elucidate the possible mechanism of disturbances in chromosome segregation leading to the increase in aneuploidy in oocytes of aged females we examined the meiotic spindles of CBA/Ca mice. Employing immunofluorescence with an anti-tubulin antibody, and human scleroderma serum, as well as 4′-6-diamidino-2-phenylindole (DAPI) staining of chromosomes the microtubular cytoskeleton could be visualized, and the behaviour of chromosomes and centromeres of oocytes spontaneously maturing in vitro could be studied. The morphology of spindles during the first meiotic division was not conspiciously different in oocytes from young and aged mice as far as the cytoskeletal elements were concerned. Neither multipolar spindles nor pronounced cytoplasmic asters appeared in oocytes of mice approaching the end of their reproductive life (9 months and older). Oocytes of aged females also did not exhibit any sign of premature separation of parental chromosomes at prophase, obvious malorientations of bivalents, or significant lagging of chromosomes during ana and telophase. Metaphase I with all bivalents aligned at the spindle equator appeared to be a relatively brief stage in oocyte development compared with pro-and prometaphase. Therefore, already slight disturbances occuring in the timing of the developmental programme which leads to a premature anaphase transition may be responsible for the high incidence of chromosomally unbalanced gametes in aged females, rather than non-separation and lagging of chromosomes during late ana-and telophase. In a second set of experiments we compared the metaphase II spindles of spontaneously ovulated oocytes obtained from animals at different ages. Previous studies have shown that spindle length and chromosome alignment may be altered in cells predisposed to aneuploidy. To distinguish between the significance of the chronological age of the female and the physiological age of the ovaries (as indicated by the total number of oocytes remaining) we examined the spindle apparatus in young (3–4 months old) and aged (9 months and older) mice as well as CBA females which had been unilaterally ovariectomized (uni-ovx) early in adult life and were approaching the end of their reproductive life at 6–7 months of age. Measurements of the pole-to-pole distance implied that spindle length may be related to maternal age. In oocytes of aged (9 month), uni-ovx (6 month) as well as 6-month-old sham-operated controls the metaphase II spindle was significantly shorter than in oocytes of young mice. By contrast, chromosome disorder and displacement was most pronounced in the aged and uni-ovx mice whilst most oocytes from young mice and moderately aged shamtreated controls exhibited a more regular alignment of chromosomes. These results, which are consistent with recent findings in CBA mice of an increased rate of aneuploidy in females approaching the end of their reproductive life, are discussed with respect to the hypothesis that the aetiology of aneuploidy rests on the critical timing of different events in oocyte development.