BACKGROUND: A correlation between morphology, developmental competence and chromosome abnormalities is established. However, since absolute correlations are rare, embryo selection remains one of the most arduous tasks in assisted reproduction. This study was undertaken in order to determine which chromosomal abnormalities are compatible with development to the blastocyst stage. METHODS: Embryos diagnosed by preimplantation genetic diagnosis (PGD) as chromosomally abnormal or unsuitable for transfer were cultured to day 5 or 6. Morphology and development were observed daily. After extended culture, embryos were fixed and analysed by two rounds of FISH with the same probes used for PGD. RESULTS: Some types of numerical chromosome abnormalities do not preclude full differentiation in vitro. For instance, extensive mosaicism was detected in blastocysts and trisomic embryos reached the blastocyst stage with a frequency of 37%. Interestingly, only those monosomies compatible with first trimester development (monosomy X and 21) were detected at blastocyst stage. CONCLUSION: Even though there is a strong selection against chromosomally abnormal embryos, extended culture to day 5 or 6 cannot be used as a reliable tool to select against clinically relevant chromosome abnormalities such as trisomies.

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Developmental ability of chromosomally abnormal human embryos to develop to the blastocyst stage

In the human, IVF and ET was developed for the treatment of tubal infertility, but it is already clear that the technique has wider applications. In this chapter, the present position of IVF and ET in infertility based on tubal, male factor and undiscovered causes (idiopathic) is discussed. There will be very rapid progress in the future, and thus some trends in research and extensions of IVF are also covered. It is anticipated that this procedure will greatly expand knowledge of the nature of human fertility disorders, particularly those that are currently obscure, such as male factor and idiopathic infertility.

Current Status and Future Prospects

In the human, mosaicism may occur before implantation; but, to determine when it first occurs, it is necessary to study the chromosomal complement of all blastomeres. Full karyotypes of blastomeres from 2- to 8-cell human embryos by conventional karyotyping of metaphase spreads are difficult to obtain. The aim of this study was to assess the stage at which mosaicism occurred in preimplantation human embryos through use of fluorescence in situ hybridization (FISH) with multiple probes. All or most blastomeres from 2- to 12-cell human embryos were analyzed by FISH using probes for gonosomes and chromosome 18. FISH was performed on blastomeres from 117 morphologically normal monospermic embryos that were not transferred after preimplantation diagnosis because of their risk of carrying X-linked disease; 20 (17.1%) of these embryos were mosaic. Another group of 163 arrested or morphologically abnormal monospermic embryos were also analyzed by FISH; 47 (28.8%) of these embryos were mosaic. In addition, 37 dispermic embryos were analyzed, and 28 (75.7%) of these were found to be mosaic. Mosaicism first occurred at the second cleavage division when the monospermic embryo was mostly diploid and at the first cleavage division when the embryo was mostly haploid, polyploid, or dispermic.

Chromosome mosaicism in human embryos.

We describe a quick and efficient method of determining the sex of DNA samples from humans, cattle, sheep, and goats. Using universal primers we have amplified 447 or 445 bp fragments from male or female genomic DNA corresponding to the ZFY or ZFX genes. Restriction fragment length polymorphism (RFLP) analysis of the fragments yielded specific banding patterns between the two sexes in these species. Horse, pig and rainbow trout fragments did not show RFLPs with the group of enzymes chosen for the other species. Use of this method would allow animal breeders to determine the sex of preimplantation embryos before transfer to recipient females. Human embryos at risk for sex-linked disorders could be sexed soon after in vitro fertilization (IVF) and options considered before implantation.

Amplification of the ZFY and ZFX genes for sex identification in humans, cattle, sheep and goats.

BACKGROUND: A large percentage of in-vitro generated cleavage stage human embryos are chromosomally mosaic, consisting of both normal (diploid) and abnormal (non-diploid) cells. The present study characterized mosaicism at each stage of cleavage division and examined its effect on preimplantation development in vitro. METHODS: A total of 216 normally fertilized (two-pronucleate) embryos which were not selected for transfer to the patients were analysed for chromosomal abnormalities using multi-colour fluorescence in-situ hybridization DNA probes specific for three to five of nine different chromosomes (X, Y, 2, 7, 13, 16, 18, 21, 22). RESULTS: Overall, 48.1% of embryos were mosaic. The frequency of mosaic embryos increased from 15.2 to 49.4 to 58.1%, from the 2-4-cell to 5-8-cell to morula stages respectively, and the types of non-diploid cells detected were mostly aneuploid or chaotic. The incidence of mosaicism at the blastocyst stage was 90.9%; however, most of the mosaicism comprised diploid and polyploid cells. Arrested mosaic embryos had a higher incidence of chaotic abnormalities, and higher proportions of abnormal cells compared with the non-arrested group. CONCLUSIONS: Post-zygotic errors leading to mosaicism may occur, and persist throughout preimplantation development in vitro. Our results suggest that mosaicism involving multiple chromosomal imbalances and/or imbalances affecting a high proportion of cells in an embryo appear to impair development to the blastocyst stage.

Chromosomal mosaicism throughout human preimplantation development in vitro: incidence, type, and relevance to embryo outcome

One hundred and fifty-nine embryos were analysed in conjunction with embryo transfer studies. Chromosome preparations were made from small biopsy fragments of trophoblast, from large portions of trophoblast and the inner cell mass, or from the entire embryo. Results obtained from fragments were similar to those obtained from large portions of trophoblast and inner cell mass. No structural chromosomal abnormalities were observed. Single triploid, diploid-triploid and diploid-hexaploid and 66 diploid-tetraploid embryos were found. The 41.5% incidence of diploid-tetraploid embryos was relatively high, and appeared to be associated with a donor factor. The transfer of 49 biopsied and analysed embryos to recipients resulted in 15 pregnancies, seven with diploid-tetraploid embryos having up to 25% of polyploid cells. The diploid-triploid and diploid-hexaploid embryos were among the transfers that did not result in pregnancy. The low incidence of embryos with chromosomal abnormalities, excluding the diploid-tetr...

Chromosomal analysis of 159 bovine embryos collected 12 to 18 days after estrus.

Embryo transfer as a means of controlling the transmission of viral infections. I. The in vitro exposure of preimplantation bovine embryos to akabane, bluetongue and bovine viral diarrhea viruses

Sexing two-week old bovine embryos by chromosomal analysis prior to surgical transfer: Preliminary methods and results

Embryo transfer as a means of controlling the transmission of viral infections. II. The in vitro exposure of preimplantation bovine embryos to infectious bovine rhinotracheitis virus.

W.C.D. Hare

Papers

Chromosomal analysis of 159 bovine embryos collected 12 to 18 days after estrus.

Embryo transfer as a means of controlling the transmission of viral infections. I. The in vitro exposure of preimplantation bovine embryos to akabane, bluetongue and bovine viral diarrhea viruses

Sexing two-week old bovine embryos by chromosomal analysis prior to surgical transfer: Preliminary methods and results

Embryo transfer as a means of controlling the transmission of viral infections. II. The in vitro exposure of preimplantation bovine embryos to infectious bovine rhinotracheitis virus.

Embryo transfer as a means of controlling the transmission of viral infections