PGD in female carriers of balanced Robertsonian and reciprocal translocations by first polar body analysis

Abstract

Preimplantation genetic diagnosis (PGD) using the first polar body (1PB) is a modality of PGD that can be used when the woman is the carrier of a genetic disease or of a balanced chromosomal reorganization. PGD using 1PB biopsy in carriers of balanced chromosome reorganizations has not become generalized. Here, we describe our experience based on the analysis of unfertilized or fresh, non-inseminated control oocytes, by fixing separately the 1PB and the corresponding oocyte, and on the study of six clinical cases of PGD using 1PB biopsy (four Robertsonian translocations and two reciprocal translocations). In fresh oocytes, the chromosome morphology of the 1PB was well preserved, and the results were always concordant for each oocyte-1PB pair. This indicates that the 1PB can be reliably used for the diagnosis of chromosome reorganizations. In these studies the technical problems encountered when performing PGD using 1PB biopsies for chromosome studies are also addressed. Three different strategies of 1PB biopsy (laser beam, partial zona dissection and acid Tyrode's) and two different protocols (intracytoplasmic sperm injection before or after 1PB biopsy) and their effect on the percentage of oocytes diagnosed and the fertilization rate, are discussed. In reciprocal translocation cases, published in the literature or studied by us, in which at least nine oocytes had been diagnosed, a correlation has been found between the frequency of nondisjunction observed and the theoretical recombination rate. To date, PGD by 1PB analysis alone or combined with blastomere biopsies in female carriers of chromosomal rearrangements has been used in 18 cases, with a further six cases reported here. A total of 325 cumulus-oocyte complexes have been obtained, of which 294 were biopsied and 224 were diagnosed. A total of 52 embryos was transferred, 19 of which implanted and 17 produced full-term pregnancies.

The importance of aneuploidy screening in reciprocal translocation carriers

“…A review on maternally derived reciprocal translocations (Durban et al 2001) showed that 56% of the oocytes diagnosed through first polar body (1PB) were unbalanced. On the other hand, Findikli (2003) also found a high percentage (69.2%) of unbalanced oocytes in female reciprocal translocation carriers.…”

“…Consequently, this method can be considered as not applicable for the detection of the unbalanced status of the translocations described. (d) PGD analysing of polar bodies (PBs), which can be used when the female is the carrier of a genetic disease or of a balanced chromosomal rearrangement (Durban et al 2001, Verlinsky et al 2004 or for aneuploidy screening (AS) in cases of advanced maternal age or in vitro fertilization (IVF) failures , Verlinsky et al 2004. This modality of PGD can be used alone or in combination with blastomere biopsy (Magli et al 2004).…”

Analysis of nine chromosome probes in first polar bodies and metaphase II oocytes for the detection of aneuploidies

“…Fixation of 1PBs was made using a modification of Durban et al, 24,32 which helps to reduce chromosome losses to a minimum and practically eliminates artefactual errors. Oocytes were fixed using a modification of a described method.…”

“…23 By analysing 1PBs, errors related to embryo mosaicism can be avoided, there is time to perform up to three rounds of FISH and, if problems have occurred during the process, a blastomere can be biopsied and analysed on time before transfer. Recently, a preimplantation genetic diagnosis using IPB (PB (PGDÀ1PB)) protocol in which ICSI was performed before 1PB biopsy was applied, 24 and the fertilisation rate achieved was comparable to the one obtained in IVF-ICSI cycles without PGD. Moreover, different types of FISH probes, (CEP), centromeric locus specific (LSI) and whole chromosome painting (WCP) can be used because the 1PB is in metaphase, and, when fixed, chromosome spreads are obtained.…”

“…Moreover, different types of FISH probes, (CEP), centromeric locus specific (LSI) and whole chromosome painting (WCP) can be used because the 1PB is in metaphase, and, when fixed, chromosome spreads are obtained. This technique has been applied when the female is a carrier of structural chromosome abnormalities (reviewed in Durban et al 24 ) or in patients with a high risk of aneuploidy. In the latter case, up to five different chromosomes have been analysed.…”