Cytogenetic abnormalities (Chandley, 1998) and microdeletions of the Y chromosome (Vogt et al., 1996) are the major genetic causes of low sperm concentrations due to varying degrees of distur-bance of meiotic processes.
In the treatment strategies for couples with idiopathic male infertility, routine cytogenetic and molecular genetic screening tests are useful to identify the reason in significant part of infertile couples and provide help in choosing the best treat-ment options. In general male populations, the prevalence of chromosomal abnormality ranges between 0.7-1% (Lange et al., 1991), while this rate is approximately 10.6% among azoospermic and oli-gozoopermic men (Dohle et al., 2002). Furthermore, the frequency of karyotypic abnormalities increases with the severity of the semen parameters (Van As-sche et al., 1996).
The deletion types of AZF a,b and c loci on Yq11, are the potential prognostic factors in patients planned to undergo testicular surger-ies such as testicular sperm extraction (TESE) and microdissection-TESE procedures (micro-TESE).
More than half of men with AZFc deletions, ma-ture spermatozoa can be obtained in ejaculate or in testis via surgery, whereas patients with complete deletions in AZFa or AZFb regions, to obtain ma-ture spermatozoa is nearly impossible (Hopps et al., 2003).
To evaluate the incidence of cytogenetic abnor-malities and Y-microdeletions in infertile men, we
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Translocation carrier couples are the most chal-lenging ones among the other PGD couples, the reason for poor prognosis comes from very low chances of producing normal or balanced gametes.
Attempts have been made to determine any possi-ble correlation between chromosomal abnormalities in gametes and embryos. With fluorescent in situ hybridization (FISH) technique applied on sperm cells, it is possible to determine the rate of
unbal-anced sperms in the ejaculate. The rate of normal or balanced sperm is especially low for reciprocal translocation carriers (reviewed by Estop et al., 1996) and higher for robertsonian translocation car-riers (reviewed by Escudero et al., 2000). To detect whether there is a relationship between unbalanced rate in gametes and embryos, we conduct a study, which revealed a strong correlation between sperm FISH results and abnormal embryo rates in PGD
TABLE I. Distribution of normal and abnormal karyotypes in infertile men according to indications KARYOTYPE
Category of Male Infetility
NOA n, (%) OAT n, (%) TOTAL n, (%)
NORMAL (46,XY) 973 (80.15) 645 (89.46) 1618 (83.6)
ABNORMAL 199 (16.4) 42 (5.83) 241 (12.45)
Klinefelter’s Syndrome (47,XXY) 133 (10.95) 5 (0.69) 138 (7.13)
Mosaic Klinefelter 10 (0.82) 6 (0.83) 16 (0.83)
Other sex chromosomemosaicisms 13 (1.07) 2 (0.28) 15 (0.78)
45,X(1)/46,XX(10) Males 11 (0.90) 0 (0.00) 11 (0.57)
Other Sex Chromosomal Abnormalities 10 (0.82) 1 (0.14) 11 (0.57)
Reciprocal Translocation 12 (0.98) 13 (1.80) 25 (1.29)
Robertsonian Translocation 1 (0.08) 12 (1.66) 13 (0.67)
Inversions 4 (0.33) 1 (0.14) 5 (0.26)
Markers 1 (1.08) 1 (0.14) 2 (0.10)
Other Abnormalities 4 (0.33) 1 (0.14) 5 (0.26)
Normal Variable Features/Polymorphisms 42 (3.46) 34 (4.72) 76 (3.93)
TOTAL 1214 721 1935
NOA= Non-Obstructive Azoospermia, OAT= Oligoasthenoteratozoospermia
Figure I. a) Karyotype of a robertsonian translocation carrier man: 45, XY, der (13;14) (q10;q10), b) FISH image using LSI, WCP and telomeric probes.
a b
2. GÜNCEL ÜREME ENDOKRİNOLOJİSİ, YARDIMCI ÜREME TEKNİKLERİ KONGRESİ ve 1. ÜREME TIBBI DERNEĞİ KONGRESİ
cycles of male reciprocal translocation carriers, while the rate of chromosomal abnormality in em-bryos was found to be higher than sperm samples of robertsonian translocation carriers (presented in PGDIS meeting, 2007).
By removing one cell on day 3 of embryonic de-velopment, it is possible to eliminate unbalanced embryos that may not implant or may end with miscarriage. In approximately the same time pe-riod, 104 PGD cycles have been performed for 77 translocation carrier men in Memorial Hospital ART and Reproductive Genetics Centre. One cell was biopsied from 6-8 cell stage embryos via laser.
Blastomeres were fixed according to hypotonic so-lution/3:1 fixative method. Commercially supplied telomeric, locus specific and centromeric FISH probes were used; for cases with advanced maternal age (AMA), at least chromosomes 13 and 21 were included in the study. Almost all transfers were performed on day 4. High clinical pregnancy rates and abortion rates show benefit of PGD especially in young maternal age group (table 2). Although, prenatal genetic diagnosis is still considered as an option for some centers, PGD technique decreases abortion rates while increasing pregnancy and implantation rates among carriers of chromosomal rearrangements (Otani et al, 2006).
There are growing evidences that point out the paternal contribution to early embryonic develop-ment and aneuploidy. Morphological abnormalities of head and tail as well as poor motility have been as-sociated with increased chromosomal abnormalities in sperm (Calegro et al., 2001; Tempest et al., 2004) and embryos (Kahraman et al., 2004). Furthermore, the incidence of aneuploidy in sperm increases with the severity of infertility condition (Gianaroli et al., 2005). Several studies have been performed which examined the benefit of PGD procedure for couples with male infertility, but prospective randomize studies are needed. In our clinic 445 couples with
male infertility have been undergone genetic diag-noses on their embryos, however only minor portion (29.4%) has male infertility as the sole reason, such as Klinefelter’s syndrome. PGD technique was ap-plied as indicated above with changes in the probe panel used. In these cases; 9-probe panel containing 13,15,16,17,1,8,21,22 and sex chromosomes were screened. Results indicated high rate of abnormality (60%) in embryos from infertile men, and this rate was even higher when additional indications were present with male infertility such as + advanced maternal age (AMA) (64%) and +AMA+ repeated im-plantation failures (RIF) (72%). Also, we compared the aneuploidy rate according to sperm source; em-bryos fertilized with ejaculate sperm versus testicu-lar sperm contained less aneuploidy although did not reached significance (56% versus 62%). In addition, 45.5% of embryos were abnormal according to the results of 16 PGD cycles performed for patients with Klinefelter’s syndrome. It should be noted that; sig-nificant part of the abnormalities were coming from haploidy and aneuploidy of sex chromosomes.
These results show that patients with male infertility, regardless of the reason, are at risk of developing abnormal embryos, and transferring the genetic defect to their children. The high frequen-cies of cytogenetic abnormalities and Y microdele-tions definitely suggest the need for genetic screen-ing and counselscreen-ing in these particular cases. Karyo-typing should be regarded as a mandatory part of the pre-treatment screening process for all men referred for ICSI and Y microdeletion analysis test is necessary to avoid useless TESE procedures.
In the case of men with Y chromosome microde-letions, with the aid of assisted reproductive tech-niques, such as ICSI, TESE and mic-TESE, inevi-tably, the situation is transmitting the genetically aberrant Y chromosome to the male offspring for that reason replacing only female embryos (Athalye et al., 2004), or if possible; freezing of the sperm in
Table II. PGD results of male translocation carriers according to maternal age
Type of Translocation Robertsonian Reciprocal
Maternal age <37 ≥37 <37 ≥37
PGD cycles/patients, n 46/35 21/13 27/21 10/8
Maternal age, mean 29.1 40.3 29.7 40.9
Abnormal embryos, % 80* 77.8 53.2* 69
CPR,% 27.3** 6** 37.5 14.2
IR,% 16 5 23 10
PGD= preimplantation genetic diagnosis, CPR=clinical pregnancy rate, IR=implantation rate
* p<0.001 , **p<0.05
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male offspring in puberty have been considered as options for these particular couples. In the case of chromosomal abnormalities, it is clear that PGD increases the success rate in these particular pa-tients. Even in couples with no identifiable genetic abnormality, the high rates of aneuploidies and mosaicisms observed in embryos suggest incom-plete maturity of sperm machinery such as defec-tive centrioles which is responsible for divisions and chromosomal segregations eventually causing developmental arrest in preimplantation embryos.