Chromosomal analysis of unfertilized human oocytes prepared by a gradual fixation-air drying method (original) (raw)
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Chromosomal FISH analysis of unfertilized human oocytes and polar bodies
Journal of Human Genetics, 2002
The incidence of chromosomal aneuploidy was studied in 208 unfertilized metaphase II human oocytes from an in vitro fertilization program by fluorescence in situ hybridization using probes for chromosomes 18, 21, and X. Chromosome spreads were prepared by a gradual fixationair-drying method. We obtained analyzable results from 183 oocytes and 93 polar bodies; 167 oocytes (91%) were normal, 11 (6%) were diploid, and 5 (3%) were aneuploid. Of the five aneuploid oocytes, four involved chromosome 21, and one involved the X chromosome. In this study, oocyte aneuploidy caused by both nondisjunction of bivalent chromosomes and predivision of univalent chromosomes was observed. The aneuploidy rate (9.8%) in the oocytes from women aged м35 years was significantly higher than that (0.7%) in those aged 23 to 34 years (P ϭ 0.0017).
A New Technique for the Cytogenetic Study of Human Oocytes
1991
=Abstract=A technique for making good quality chromosome preparations from human oocytes is described. Analyzable good quality chromosomes were present in human oocytes that were hypotonically treated and fixed. Ninety-eight oocytes were processed. Chromosome analysis could be performed on 80 (82%) oocytes. Seven oocytes showed an absence of chromosomes, mainly due to degeneration. Chromosome analysis could not be performed on 11 oocytes because the chromosome number could not be determined. Of 80 analyzable preparations, a 63% (50) incidence of aneuploidy withlwithout premature chromosome condensation was observed in the human oocytes that failed to fertilize in vitro with 3:4 ratios of hypo and hyperhaploidy. Our method presented in this study gives an excellent scattering of nuclei and metaphase plates in one layer as well as a good spreading of chromosomes. It may be applied to all stages of human oocytes in IVF-ET programs. This method can also be useful in pre-embryo biopsy and in secondhand quality control for an IVF-ET program. This may help resolve the present limitations of oocyte chromosome preparation and analysis, and therefore lead to further improvements in germ cell cytogenetics.
Journal of In Vitro Fertilization and Embryo Transfer, 1985
Chromosomal abnormalities and abnormal embryonic development have previously been observed after human in vitro fertilization (IVF). Chromosomal abnormalities may arise not only after fertilization but even earlier during meiotic maturation of human ooeytes in culture. Since chromosomal analysis is simple in oocytes during meiotic maturation, the chromosomal status was analyzed in oocytes which remained unfertilized in a human in vitro fertilization program. In 50fertilization attempts the chromosomes of 62 unfertilized oocytes could be analyzed; 45 of them were in the process of meiotic maturation. In three oocytes two small polar bodies were observed 16-18 hr after insemination in the absence offertilization. In one oocyte abnormal chromosome behavior was found during the first meiotic division, and in four oocytes during metaphase of the second meiotic division. These data suggest that chromosomal analysis of unfertilized oocytes in human IVF may improve the understanding of human oocyte maturation and fertilization.
Human Reproduction, 1998
The incidence of chromosomal abnormalities was studied in 719 unfertilized human oocytes obtained from our invitro fertilization (IVF) programme. To make chromosome preparations, a gradual fixation/air-drying method was utilized. Of 388 oocytes successfully karyotyped, 70 (18.0%) were abnormal. The abnormalities included 33 aneuploidies (8.5%) (14 hyperhaploidies and 19 hypohaploidies), 25 diploidies (6.4%) and 15 structural abnormalities (3.9%), three of them being accompanied by aneuploidy. Of the 33 aneuploidies, 16 (48.5%) showed the loss or gain of dyads (so-called non-disjunction), while 17 (51.5%) showed the loss or gain of monads (so-called predivision). There was no maternal age-dependent increase in the incidence of aneuploidy. Unfertilized oocytes from patients with a high fertilization rate (>25%) had a significantly higher (11.4%, P < 0.05) incidence of diploidy compared with the oocytes from the remaining patients (4.3 and 4.0%), suggesting that diploid oocytes might have a lower fertilizing ability.
Cytogenetics of unfertilized human oocytes
Reproduction, 1988
During an in-vitro fertilization programme 150 oocytes from 62 women with a mean age of 31 years (range 24-39) remained unfertilized. Successful chromosome analysis was carried out on 96 oocytes by Q-banding: 59 (61.5%) oocytes bore a normal haploid complement, 8 (8.3%) were diploid and 3 (3.1%) tetraploid. In 26 (27.1%) oocytes aneuploidy was observed; these included 9 (9.4%) nullisomic, 5 (5.2%) double nullisomic, 4 (4.2%) triple nullisomic and 2 (2.1%) disomic oocytes. The remaining 54 (36.0%) oocytes could not be evaluated. A nearly uniform rate of aneuploidy was found for unfertilized oocytes among different donor age groups.
Journal of In Vitro Fertilization and Embryo Transfer, 1990
In vitro fertilization cycles yield a low percentage of pregnancies, Eighty-five to ninety percent of the transferred embryos do not implant, and the abortion rate approaches 30%. Aneuploidy is assumed to be responsible for a major portion of this pregnancy wastage. The purpose of this study was to determine if there was any correlation between morphology and chromosomal content of unfertilized oocytes and rejected embryos. To assess the chromosomal content of oocytes and embryos, we used the method described by Tarkowsld in 1966. Sixty oocytes from 28 women, aged between 27 and 41 years, were analyzed. Sixty-seven percent were aneuploid; of these, 23.35% were hyperhaploid, 23.35% were hypohaploid, 8.35% were hyperdiploid, 3.35% were diploid, and 8.35% showed premature chromosome condensation. Of 20 preimplantation embryos analyzed, 80% were aneuploid, 10% were diploid, 5% were haploid, and 5% showed structural anomaly. Correlation was found between maternal age and aneuploidy in oocytes and between morphology and genetic balance in preimplantation embryos.
Non-meiotic chromosome instability in human immature oocytes
European Journal of Human Genetics, 2013
Aneuploidy has been a major issue in human gametes and is closely related to fertility problems, as it is known to be present in cleavage stage embryos and gestational losses. Pre-meiotic chromosome abnormalities in women have been previously described. The aim of this study is to assess the whole-chromosome complement in immature oocytes to find those abnormalities caused by mitotic instability. For this purpose, a total of 157 oocytes at the germinal vesicle or metaphase I stage, and discarded from IVF cycles, were analysed by CGH. Fifty-six women, between 18 and 45 years old (mean 32.5 years), including 32 IVF patients (25-45 years of age) and 24 IVF oocyte donors (18-33 years of age), were included in the study. A total of 25/157 (15.9%) of the oocytes analysed, obtained from three IVF clinics, contained chromosome abnormalities, including both aneuploidy (24/157) and structural aberrations (9/157). Independently of the maternal age, the incidence of abnormal oocytes which originated before meiosis is 15.9%, and these imbalances were found in 33.9% of the females studied. This work sheds light on the relevance of mitotic instability responsible for the generation of the abnormalities present in human oocytes.
Human Reproduction, 2004
BACKGROUND: The object of this study was to determine the mechanisms that produce aneuploidy in oocytes and establish which chromosomes are more prone to aneuploidy. METHODS: A total of 54 oocytes from 36 women were analysed. The whole chromosome complement of the first polar body (1PB) was analysed by comparative genomic hybridization (CGH), while the corresponding metaphase II (MII) oocyte was analysed by fluorescence in situ hybridization (FISH) to confirm the results. RESULTS: Matched CGH-FISH results were obtained in 42 1PB -MII doublets, of which 37 (88.1%) showed reciprocal results. The aneuploidy rate was 57.1%. Two-thirds of the aneuploidy events were chromatid abnormalities. Interestingly, the chromosomes more frequently involved in aneuploidy were chromosomes 1, 4 and 22 followed by chromosome 16. In general, small chromosomes (those equal to or smaller in size than chromosome 13) were more prone to aneuploidy (x 2 -test, P 5 0.07); 25% of the aneuploid doublets would have been misdiagnosed as normal using FISH with probes for nine-chromosomes. CONCLUSIONS: The combination of two different techniques, CGH and FISH, for the study of 1PB and MII allowed the identification and confirmation of any numerical chromosome abnormality, as well as helping to determine the mechanisms involved in the genesis of maternal aneuploidy.
2009
The aim of this work is to analyze, using the comparative genomic hybridization technique, the frequencies and the mechanisms involved in the production of aneuploidy events in donor oocytes. The results showed that 32.1% of them were aneuploid, with 51.7% of those originating from first meiotic division errors and 48.3% from the presence of aneuploid oogonium. (Fertil Steril Ò 2010;93:675-9. Aneuploidy is one of the main causes behind low pregnancy rates in humans (1). Chromosome abnormalities are found in both early cleavage stage embryos and first trimester, spontaneously aborted fetuses (2, 3). Most of the aneuploidy is due to errors in the female first meiotic division (4, 5); therefore, the cytogenetic study of oocytes may provide useful data. To describe the aneuploidy rate in oocytes, several groups have analyzed the first polar body (1PB) and its corresponding metaphase II (MII) stage oocyte. Most of the studies were performed using discarded oocytes, generally MI immature oocytes, from IVF patients that were in vitro matured (IVM) until they reached the MII stage. Different investigators describe an aneuploidy rate of around 50%, analyzing IVM-MII oocytes from IVF patients with a mean age of 40 (6-11). The considerably high aneuploidy rate found in this population of patients is concordant with their low pregnancy expectations; in addition, it must also be taken into consideration that they are patients who are participating in an IVF program and for the most part they have an advanced maternal age indication.