Peter M.M. Kastrop

Analysis of spermatozoa from seven ICSI males with constitutional sex Chromosomal abnormalities by fluorescent in situ hybridization

AbstractPurpose: The objective was to estimate the risk for subfertilemales with a constitutional sex chromosomal abnormalityof transmitting such a chromosome abnormality to theirchildren, conceived by intracytoplasmic sperm injection(ICSI). Methods: Semen samples were obtained from seven severelyoligospermic ICSI candidates. Six of them had a numericalsex chromosomal abnormality, including mosaic 45,X/46,XY,mosaic 46,XY/47, XXY, 47,XXY (Klinefelters syndrome), and47,XYY. One male had a structural abnormality, namely, aninversion of the Y chromosome. The semen was studied bythree-color fluorescent in situ hybridization (FISH) withprobes specific for chromosomes 18,X, and Y. Results: Chromosomal aneuploidy rates of any of the threechromosomes were significantly higher than the aneuploidyrates observed in three control samples but comparable tothe rates observed in 10 ICSI candidates witholigoasthenoteratozoospermia (OAT) and a normal constitutionalkaryotype. Conclusions: Our data indicate that males with (mosaic) sexchromosomal abnormalities have no higher risk of producingoffspring with a sex chromosomal abnormality by ICSI thanOAT males with a normal karyotype.

PGD for hereditary breast and ovarian cancer: the route to universal tests for BRCA1 and BRCA2 mutation carriers.

Preimplantation Genetic Diagnosis (PGD) is a method of testing in vitro embryos as an alternative to prenatal diagnosis with possible termination of pregnancy in case of an affected child. Recently, PGD for hereditary breast and ovarian cancer caused by BRCA1 and BRCA2 mutations has found its way in specialized labs. We describe the route to universal single-cell PGD tests for carriers of BRCA1/2 mutations. Originally, mutation-specific protocols with one or two markers were set up and changed when new couples were not informative. This route of changing protocols was finalized after 2 years with universal tests for both BRCA1 and BRCA2 mutation carriers based on haplotyping of, respectively, 6 (BRCA1) and 8 (BRCA2) microsatellite markers in a multiplex PCR. Using all protocols, 30 couples had a total of 47 PGD cycles performed. Eight cycles were cancelled upon IVF treatment due to hypostimulation. Of the remaining 39 cycles, a total of 261 embryos were biopsied and a genetic diagnosis was obtained in 244 (93%). In 34 of the 39 cycles (84.6%), an embryo transfer was possible and resulted in 8 pregnancies leading to a fetal heart beat per oocyte retrieval of 20.5% and a fetal heart beat per embryonic transfer of 23.5%. The preparation time and costs for set-up and validation of tests are minimized. The informativity of microsatellite markers used in the universal PGD-PCR tests is based on CEPH and deCODE pedigrees, making the tests applicable in 90% of couples coming from these populations.

The effect of colchicine treatment on spermatozoa: a cytogenetic approach.

Colchicine is an alkaloid which exerts its main effect at the cellular level by its interference with microtubule formation, thereby affecting mitosis and other microtubule-dependent functions (1). In routine cytogenetic diagnostics, colchicine is commonly used in in vitro culture systems to block spindle formation and arrest cells undergoing mitosis at the metaphase stage (2,3). On the other hand, due to its antiinflammatory effect, colchicine is used for the treatment of several diseases, including gouty arthritis (4), familial Mediterranean fever (FMF) (5), and Behcets disease (6). The effects of colchicine on sperm production and function in men are controversial. Recently, HaimovKochman and Ben-Chetrit (7) reviewed all papers published from 1966 to April 1997 regarding this effect in healthy individuals as well as in patients. The authors concluded that colchicine seems to have no significant direct adverse effect on sperm production and function as determined by routine sperm analysis. However, it is unknown whether the incidence of aneuploidy in spermatozoa of men treated with colchicine is affected, since colchicine can act directly on chromosomes by detaching chromosomes from the meiotic spindle. Recently, two infertile couples of which the male partners were treated with colchicine attended our in vitro fertilization (IVF) center. In both couples routine sperm analyses were performed and an additional ejaculate was prepared for cytogenetic analysis by three-color fluorescence in situ hybridization (FISH) with probes specific for chromosomes 18, X, and Y(8).

Can Characteristics of Reciprocal Translocations Predict the Chance of Transferable Embryos in PGD Cycles

Translocation carriers have an increased risk of miscarriage or the birth of a child with congenital anomalies. Preimplantation genetic diagnosis (PGD) is performed in translocation carriers to select for balanced embryos and, thus, increase the chance of an ongoing pregnancy. However, a common experience is that reciprocal translocation carriers produce a high percentage of unbalanced embryos, which cannot be transferred. Therefore, the pregnancy rates in PGD in this patient group are low. In a cohort of 85 reciprocal translocation carriers undergoing PGD we have searched for cytogenetic characteristics of the translocations that can predict the percentage of balanced embryos. Using shape algorithms, the most likely segregation mode per translocation was determined. Shape algorithm, breakpoint location, and relative chromosome segment sizes proved not to be independent predictors of the percentage of balanced embryos. The ratio of the relative sizes of the translocated segments of both translocation chromosomes can give some insight into the chance of transferable embryos: Very asymmetrical translocations have a higher risk of unbalanced products (p = 0.048). Counseling of the couples on the pros and cons of all their reproductive options remains very important.

Does the genetic and familial background of males undertaking ICSI affect the outcome

PurposeTo investigate whether the success rate of ICSI is (1) related to the etiology of infertility or (2) adversely affected by a family history of potential genetic disorders.MethodsAll men with an ICSI indication in our hospital between 1994 and 2005 were included in our cohort study. Data on the ICSI process, etiology of infertility, and family history were collected. ICSI success rates of infertility subgroups and a subgroup with a positive family history were compared to a group with unknown etiology and a negative family history.ResultsThere was no significant difference in clinical pregnancy or delivery rates between the subgroups. Couples achieving a pregnancy underwent significantly more ICSI cycles compared to couples not achieving a pregnancy.ConclusionOur results suggest that the success rate of ICSI treatment is not related to the cause of infertility or a family history positive for potential genetic disorders.