Huntington's disease (HD) is an autosomal dominant disease characterized by motor disturbance, cognitive loss and psychiatric manifestations, starting between the fourth and the fifth decade, followed by death within 10–20 years of onset of the disease. The disease‐causing mutation is an expansion of a CAG triplet repeat at the 5′ coding end of the Huntington gene. We have developed a single‐cell PCR assay for the HD gene in order to propose preimplantation genetic diagnosis (PGD) for the couples at risk. We present here our first results with our first nine PGD cycles and also discuss the behaviour of the disease‐causing expansion in pre‐implantation embryos. Copyright © 1998 John Wiley & Sons, Ltd.
Myotonic dystrophy (DM), or Steinert's disease, is an autosomal dominant disease characterized by myotonia, muscular weakness and atrophy, as well as lens opacities, cardiomyopathy and mild endocrine changes. The gene for DM located on 19q contains a triplet repeat at the 3' end of the gene. In DM patients, this repeat is found to be expanded. We have previously described a preimplantation genetic diagnosis (PGD) for DM using polymerase chain reaction (PCR) followed by conventional analysis on ethidium bromide-stained gels. The major drawback of this system was that allelic dropout occurred in >20% of the cells, leading to the loss of healthy embryos for transfer. To resolve this problem, we developed a PGD for DM using fluorescent PCR followed by fragment analysis on an automated DNA sequencer and made a comparison between the conventional PCR described earlier and fluorescent PCR, which turned out to be superior in accuracy and efficiency. Three PGD cycles were performed using fluorescent PCR and are described here.
Preimplantation genetic diagnosis (PGD) can be offered as an alternative to prenatal diagnosis (PND) to couples at risk of having a child with a genetic disease. The affected embryos are detected before implantation by fluorescent in situ hybridisation (FISH) for sexing (X‐linked diseases) and chromosomal disorders (numerical and structural) or by polymerase chain reaction (PCR) for monogenic disorders (including some X‐linked diseases). The accuracy and reliability of the diagnosis is increased by analysing two blastomeres of the embryo. However, the removal of two blastomeres might have an effect on the implantation capacity of the embryo. We have evaluated the implantation of embryos after the removal of one, two or three cells in 188 PGD cycles where a transfer was done. The patients were divided into five groups: a first group which received only embryos from which one cell had been removed, a second group which received only embryos from which two cells had been removed, a third group which received a mixture of embryos from which one and two cells had been taken, a fourth group where two and three cells had been removed, and a fifth group where three cells had been removed. The overall ongoing pregnancy rate per transfer was 26.1%, the overall implantation rate per transfer was 15.2% and the overall birth rate was 14.2%. Although pregnancy rates between the groups cannot be compared because the second group (two cells removed) contains more rapidly developing and therefore ‘better quality’ embryos, an ongoing pregnancy rate of 29.1% and an implantation rate of 18.6% per transferred embryo in this group is acceptable, and we therefore advise analysing two cells from a ≥7‐cell stage embryo in order to render the diagnosis more accurate and reliable. Copyright © 2000 John Wiley & Sons, Ltd.
A controlled comparison between conventional in-vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) has been carried out for patients with =5% rapid progressive (type A) spermatozoa in their fresh semen. Twenty couples were allocated to the study. All semen samples fulfilled the minimum criteria to be allocated to the conventional IVF programme. The two pronuclear (2PN) fertilization rate per retrieved cumulus-oocyte complex was significantly higher after ICSI (63.4%) than after conventional IVF (22.9%), with complete fertilization failure after IVF in 10 out of the 20 cycles. Embryo quality was similar for both treatments. Sixteen patients received only ICSI embryos, two patients only IVF embryos and two patients received IVF and ICSI embryos. The ongoing clinical pregnancy rate was 45%, the implantation rate was 37%. Comparison of the characteristics of patients/cycles with and without 2PN fertilization revealed a higher proportion of progressively motile spermatozoa in the prepared sperm fraction for the group of patients with fertilization after conventional IVF. It can be concluded from this study that absence of or an extremely low proportion of rapid progressive motility in fresh semen indicates a high risk of complete fertilization failure with conventional IVF.
This paper describes the 5 years' experience of preimplantation genetic diagnosis (PGD) at the Brussels Free University. Our first PGD was carried out in February 1993. Up to October 1998, we carried out 183 PGD cycles on fresh cleavage embryos of 92 couples for 25 different conditions. Patients were treated for autosomal recessive (n = 39), autosomal dominant (n = 65) and X-linked recessive (n = 47) monogenic disorders as well as for autosomal structural aberrations (n = 10), sex chromosome numerical and structural aberrations (n = 21) and a combination of the two latter (n = 1). Specific diagnosis was carried out by polymerase chain reaction (n = 108). Fluorescence in-situ hybridization was used for sexing (n = 64) and structural aberrations (n = 11). We transferred 1.6 +/- 1.1 embryos per cycle, resulting in an implantation rate of 12.0% per replaced embryo. Ongoing pregnancies were achieved in 29 cycles, i.e. 23 singletons, five twins and one dichorionic triplet with an acardius acranius. The ongoing pregnancy rates per cycle, per transfer and per couple were 16.4, 19.9 and 31.5% respectively. While 28 ongoing pregnancies resulted in the births of 34 infants, one pregnancy was terminated after misdiagnosis. The results of 24 PGD were confirmed by prenatal diagnosis or after birth while no information was available in four pregnancies. Our series demonstrates that PGD is a feasible technique by which to avoid the birth of genetically affected children to couples at risk.
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