Y Saglam

Short tandem repeats haplotyping of the HLA region in preimplantation HLA matching.

Recently, preimplantation genetic diagnosis (PGD) has been considered for several indications beyond its original purpose, not only to test embryos for genetic disease but also to select embryos for a nondisease trait, such as specific human leukocyte antigen (HLA) genotypes, related to immune compatibility with an existing affected child in need of a haematopoetic stem cell (HSC) transplant. We have optimized an indirect single-cell HLA typing protocol based on a multiplex fluorescent polymerase chain reaction (PCR) of short tandem repeat (STR) markers scattered throughout the HLA complex. The assay was clinically applied in 60 cycles from 45 couples. A conclusive HLA-matching diagnosis was achieved in 483/530 (91.1%) of the embryos tested. In total, 74 (15.3%) embryos revealed an HLA match with the affected siblings, 55 (11.4%) of which resulted unaffected and 46 (9.5%) have been transferred to the patients. Nine pregnancies were achieved, five healthy HLA-matched children have already been delivered and cord blood HSCs, were transplanted to three affected siblings, resulting in a successful haematopoietic reconstruction.

Embryo aneuploidy screening for repeated implantation failure and unexplained recurrent miscarriage

Among other factors, chromosomal abnormalities that originate from gametogenesis and preimplantation embryonic development are thought to be one of the major contributing factors for early embryonic death and failure of pregnancy. However, so far, no non-invasive technique exists that allows the detection of the chromosomal complement of an oocyte or a developing embryo as a whole. Rather, by removing polar bodies/blastomeres, recent developments on preimplantation genetic diagnosis for aneuploidy screening (PGD-AS) have paved the way to detect and possibly eliminate the majority of chromosomally abnormal embryos, thereby increasing the chance of a healthy pregnancy. This article summarizes the origin and impact of chromosomal abnormalities on human reproduction in cases with repeated implantation failure (RIF) and unexplained recurrent miscarriage. It also discusses recent advances regarding the possible benefits of PGD-AS in such cases.

Preliminary FISH studies on spermatozoa and embryos in patients with variable degrees of teratozoospermia and a history of poor prognosis.

The aim of this study was to analyse to what extent sperm aneuploidy is associated with sperm morphology and subsequently with embryo aneuploidy. Fifty-nine men with variable degrees of teratozoospermia and previously poor assisted reproduction prognosis were included in the study. Samples from 10 normozoospermic men with proven fertility were used as controls. Individual spermatozoa were scored for chromosomes 13, 21 and for 18, X, Y separately. Compared with controls, 23 out of 59 cases (39.0%) were found to have increased sperm aneuploidy for at least one of the chromosomes analysed in a treatment cycle. Fifty-two patients underwent a treatment cycle and were documented according to the pregnancy and spermatozoa fluorescence in-situ hybridization results. A total of 121 previous unsuccessful assisted reproduction cycles of the cases were then retrospectively reviewed. In 23 of the latest cycles, preimplantation genetic diagnosis was applied to 106 cleavage stage embryos and 47 of 94 embryos analysed (50.0%) were found to be chromosomally abnormal. Furthermore, 16 of 47 (34.0%) embryos with chromosomal abnormality were carrying complex chromosomal defects. The results imply that increased aneuploidy is present in both spermatozoa and embryos in couples with severe male infertility with a history of repeated unsuccessful attempts. Therefore, proper genetic counselling should be considered in these cases.

Genetic diagnosis in infertile men with numerical and constitutional sperm abnormalities.

Infertile men having numerical or structural sperm defects may carry several genetic abnormalities (karyotype abnormalities, Y chromosome microdeletions, cystic fibrosis transmembrane conductance regulator (CFTR) gene mutations, androgen receptor gene mutations, and abnormalities seen in sperm cells) leading to this situation. First we aimed to investigate the relationship between the numerical and constitutional (morphological) sperm anomalies and the genetic disorders that can be seen in infertile males. Our other aim was to compare two different kinds of kits that we use for the detection of Y chromosome microdeletions. Sixty-three infertile males [44 nonobstructive azoospermic, 8 severe oligozoospermic, and 11 oligoasthenoteratozoospermic] were investigated in terms of somatic chromosomal constitutions and microdeletions of the Y chromosome. Sperm aneuploidy levels were analyzed by fluorescence in situ hybridization (FISH) in sperm cells obtained from the semen of six OAT patients. Microdeletion and sex chromosome aneuploidy (47,XXY) rates in somatic cells were found to be approximately 3.2% and 4.7%, respectively. Sperm aneuploidy rates were determined as 9%, 22%, and 47% in three patients out of six. Two of these three patients also had high rates of head anomalies in semen samples. High correlation was found between sperm aneuploidy rates and sperm head anomalies. Since the introduction of the assisted reproductive techniques for the treatment of severe male infertility, genetic tests and genetic counseling became very important due to the transmission of genetic abnormalities to the next generation. Thus in a very near future, for a comprehensive male infertility panel, it will be essential to include additional genetic tests, such as CFTR gene mutations, sperm mitochondrial DNA mutations, and androgen receptor gene mutations, besides the conventional chromosomal analyses, Y chromosome microdeletion detection, and sperm-FISH analyses.

Medical and social perspectives of PGD for single gene disorders and human leukocyte antigen typing

Preimplantation genetic diagnosis (PGD) for single gene disorders combined with human leukocyte antigen (HLA) typing has recently emerged as a therapeutic tool. For couples who are at risk of passing on a genetic disease to their offspring, preimplantation embryos can be selected according to their genetic status as well as a possible HLA matching with the affected sibling. Stem cells from the resulting babys umbilical cord blood have, therefore, a great therapeutic value for haematopoietic and other life threatening diseases, as stem cells in the cord blood from a HLA-compatible newborn can be used for transplantation without graft rejection, thus saving an affected childs life. However, apart from being a valuable treatment option, there exist several medical and social aspects that should be evaluated and discussed. From the ethical and the social aspects, although PGD for single gene disorders is well defined and accepted, application of PGD combined with HLA typing is less obvious and under extensive debate. This article is aimed at summarizing the current results and limitations of PGD with HLA typing that are related to the successful medical outcome. It further discusses the ethical and social issues that have recently been raised on the application of this technique.

The Mayer-Rokitansky-Kuster-Hauser and gonadal dysgenesis anomaly in a girl with 45,X/46,X,del(X)(p11.21).

The Mayer–Rokitansky–Kuster–Hauser (MRKH) anomaly is characterized by uterine aplasia or hypoplasia associated with normal ovaries [Morcel et al., 2007]. The co-occurrence of gonadal dysgenesis and mullerian agenesis has been previously described as a rare event [Alper et al., 1985; Mohapatra et al., 1992; Guitron-Cantu et al., 1999]. Structural or numerical abnormalities of X chromosome can cause gonadal dysgenesis as absent or streak gonads [Goldman et al., 1982]. Although gonadal dysgenesis has been reported with structural abnormalities in X chromosome such as 46Xi(Xq),45,X and 45X/46XX, neither structural nor numerical abnormalities of X chromosome have been associated with the MRKH anomaly [Gardo et al., 1971; De Leon et al., 1984; Gray et al., 2001]. Associated skeletal abnormalities including spine, face and limb such as asymmetric or fused vertebra, ectrodactly, radial agenesis and facial asymmetry have been reported in patients with MRKH [Behera et al., 2005; Morcel et al., 2007]. The etiology of this condition is still unknown. Here, we report on a girl whohas 45,X/46,X,del(X)(p11.21!pter)withMRKH and gonadal dysgenesis. A 17-year-old girl with complaints short stature, primary amenorrhea, hypertrophy of both second toes was admitted to the hospital. She was attending high school, and her school performance was good. She had two sisters and two brothers who were chromosomal and physically normal. There was no consanguinity in her parents who were also normal. Physical examination showed that her height was 129.9 cm (<3rd centile); weight 28.8 kg (<3rd centile). Height SDS was – 4.5, bone age was 12. She had downslanting palpebral features, a webbed neck, low posterior hairline, cubitus valgus, and short 4th metacarpals. She also had long and hyperplastic second toes in both feet (Fig. 1) and a grade 1/6 systolic murmur. Development of breast was Tanner phase I, pubic hair Tanner phase II. Axillary hair was absent. Serum gonadotropin levels were high (LH: 14.43 ml U/ml, FSH: 86.59 ml U/ml) and estradiol level were low (10.44 pg/ml). Thyroid function test was normal. Serum fasting glucose, lipids and insulin levels were within normal range. Bone mineral density (BMD) obtained from lumbar vertebral and femoral neck was osteopenic (Z score: 1.93 and 1.21, respectively). Echocardiographic and abdominal ultrasonographic examinations were normal. Ultrasonographic examination and magnetic resonance imaging of the pelvis revealed absence of uterus and ovaries and hypoplastic vagina. Cytogenetic analysis was performed on peripheral blood lymphocytes. A 100 GTG banded metaphases were analyzed utilizing the following DNA probe sequences: CEP X (DXZ1) SpectrumAquaTM; TelVysion Xp/Yp SpectrumGreenTM TelVysion Xq/Yq SpectrumOrangeTM (Vysis). Of 100 metaphase cells examined from PHA-stimulated blood cultures by conventional G-banding techniques, 30 were characterized as missing a sex chromosome (45,X) while the remaining 70 were characterized as containing number of chromosomes with deletion

An epileptic case with mosaic ring chromosome 6 and 6q terminal deletion

Ring chromosomes are rare chromosome disorders that arise usually de novo. Children with ring chromosome 6 have a wide range of intellectual functioning and congenital anomalies. We report an epileptic case of a 10-year-old boy to be mild psychomotor retardation and dysmorphic traits including microcephaly, brachycephaly, flat occiput, large and apparently low set ears, and bilateral syndactyly between his second and third fingers with mosaic ring chromosome 6 and 6q terminal deletion. Peripheral chromosome and fluorescent in situ hybridisation (FISH) analysis of the patient showed mos46,XY,r(6)(p24;q26),del(6)(q27) [30]/46,XY,del(6)(q27) [20] de novo. We presented the patient in the light of literature because the mosaic ring 6 and 6q terminal deletion was different caryotypically from other mosaic ring 6 patients.

Preimplantation genetic diagnosis (PGD) for extremes—successful birth after PGD for a consanguineous couple carrying an identical balanced reciprocal translocation

OBJECTIVE To report a healthy birth after preimplantation genetic diagnosis (PGD) performed for a consanguineous couple carrying an identical familial reciprocal translocation in both partners. DESIGN Case report. SETTING In vitro fertilization (IVF) clinic and genetic laboratory in a private hospital. PATIENT(S) Consanguineous couple carrying the same balanced reciprocal translocation: 46,XX,t(1;16)(q12;q11.2) and 46,XY,t(1;16)(q12;q11.2). INTERVENTION(S) 25 oocyte-cumulus complexes were retrieved 36 hours after human chorionic gonadotropin injection; metaphase II oocytes were fertilized by intracytoplasmic sperm injection; single blastomere biopsy was performed on 15 embryos on day 3; one embryo was found to be normal or balanced according to fluorescent in situ hybridization studies, embryo transfer was performed on day 4. MAIN OUTCOME MEASURE(S) Healthy birth of homozygous double translocation carrier twins with 46,XY,t(1;16)(q12;q11.2)mat,t(1;16)(q12;q11.2)pat karyotype. RESULT(S) Healthy monozygotic male twins were born at 36 weeks of gestation. Karyotype studies of the babies revealed that they are double translocation homozygotes: 46,XY,t(1;16)(q12;q11.2)mat,t(1;16)(q12;q11.2)pat. They are healthy and more than 4 years old later show no physical or mental abnormalities. CONCLUSION(S) To our knowledge, this is the first PGD study performed for a couple who carry the same reciprocal translocation. The twins born after this study are rare examples in the literature of healthy balanced reciprocal translocation homozygotes.