T Mamas

Detection of aneuploidy by array comparative genomic hybridization using cell lines to mimic a mosaic trophectoderm biopsy

OBJECTIVE To examine the effect of mosaicism in the array comparative genomic hybridization result during preimplantation genetic screening after blastocyst biopsy. DESIGN Experimental study. SETTING University laboratory. MATERIAL(S): Epithelial cell lines. INTERVENTION(S) Mixing of euploid and aneuploid cells to create mosaic trophectoderm and blastocyst models. MAIN OUTCOME MEASURE(S) The level of aneuploidy in samples with different ratios of aneuploid cells was measured after array comparative genomic hybridization. RESULT(S) A shift from normality was present when the level of aneuploid cells in the sample was >25%. Aneuploidy could be confidently called when the level of aneuploid cells was >50%. CONCLUSION(S) This study determined that aneuploidy in mosaic samples can be detected by array comparative genomic hybridization and that the result may also indicate the proportion of the aneuploid cells present in the sample.

Preimplantation genetic diagnosis for myotonic dystrophy type 1 in the UK.

Myotonic dystrophy type 1 (DM1) is a dominant multisystemic disorder caused by expansion of a trinucleotide repeat in a non-coding region of DMPK. Prenatal diagnosis (PND) is available; however, the decision to terminate affected pregnancies is difficult as the extent of disability is hard to predict from the size of the expansion. In preimplantation genetic diagnosis (PGD) genetic analysis is carried out before the establishment of pregnancy. This paper reviews the largest number of cycles of PGD for DM1 in the UK indicating that PGD is a practical option for affected couples.

Modification of the triplet repeat primed polymerase chain reaction method for detection of the CTG repeat expansion in myotonic dystrophy type 1: application in preimplantation genetic diagnosis

OBJECTIVE To overcome problems associated with the use of triplet repeat primed polymerase chain reaction (TP-PCR) in preimplantation genetic diagnosis (PGD) of myotonic dystrophy type 1 (DM1). DESIGN Clinical research study. SETTING UCL Centre for PGD and Centre for Reproductive and Genetic Health. PATIENT(S) Seven couples undergoing PGD for DM1. INTERVENTION(S) A modified TP-PCR protocol (mTP-PCR) for the reliable detection of both expanded and nonexpanded alleles in DMPK was optimized using single lymphocytes. Four cycles of PGD were performed with TP-PCR for diagnosis and a further 10 cycles with mTP-PCR. MAIN OUTCOME MEASURE(S) Amplification efficiency, allele dropout, diagnosis rate, and delivery rate. RESULT(S) Preliminary testing showed that the TP-PCR amplification efficiency was higher using lymphocytes versus buccal cells. Single lymphocytes gave very high amplification efficiencies for both protocols (99% to 100%). There were no false-positive or false-negative results for 148 single lymphocytes tested with mTP-PCR compared with 9% (5 out of 54) false-positive results with TP-PCR, indicating the improved accuracy of the modified protocol. In embryos, the diagnosis rate was 95.6% with mTP-PCR and 75% with TP-PCR. CONCLUSION(S) For PGD of DM1, mTP-PCR is recommended. It may also be applied as a rapid screen for DMPK expansions in individuals with symptoms of DM1, relatives of known mutation carriers, or in prenatal diagnosis.

The nature and origin of binucleate cells in human preimplantation embryos: relevance to placental mesenchymal dysplasia

Cleavage-stage embryos often have nuclear abnormalities, one of the most common being binucleate blastomeres, which may contain two diploid or two haploid nuclei. Biopsied cells from preimplantation genetic diagnosis (PGD) and preimplantation genetic screening (PGS) cycles were studied to determine the relative frequency of binucleate cells with two haploid versus two diploid nuclei. The frequency of mononucleate haploid biopsied blastomeres was also recorded. In the chromosomal PGD cycles 45.2% of the biopsied binucleate cells were overall diploid and 38.7% were overall tetraploid, compared with 50.0% and 29.2% for the PGS group, respectively. Placental mesenchymal dysplasia is a rare condition associated with intrauterine growth restriction, prematurity and intrauterine death. Recent work suggests that androgenetic diploid/haploid mosaicism may be a causal mechanism. There are two possible origins of haploid nuclei, either the cell contained only one parental genome initially or they may be derived from the cytokinesis of binucleate cells with two haploid nuclei. Binucleate formation therefore may be a way of doubling up the haploid genome, to produce diploid cells of androgenetic origin as seen in placental mesenchymal dysplasia.