Chromosomal segregation in sperm of Robertsonian translocation carriers

Purpose

To study meiotic segregation patterns of Robertsonian translocations in sperm of male carriers and to assess the frequencies of unbalanced sperm formation.

Methods

FISH with combination of probes to detect all the variants of meiotic segregation was performed on decondensed sperm nuclei of 5 carriers of der(13;14), 3 carriers of der(14;21) and one carrier of a rare der(13;21) translocation.

Results

The frequency of sperm with alternate segregation and normal/balanced chromosomal complement ranged from 68 % to 94.4 % (mean 79.2 ± 8.4). Adjacent segregation was detected in 17.9 ± 7.3 % of sperm (from 5.6 % to 29 %). No significant differences in frequencies of gametes with nullisomies and disomies of chromosomes involved in translocations were observed. The mean frequency of 3:0 segregation products was 2.5 ± 1.4 %.

Conclusions

All analyzed patients showed homogenous segregation pattern with clear predominance of alternate segregation resulting in normal/balanced sperm production. Still, from 5.8–32 % (mean 20.4 ± 8.3 %) of sperm was unbalanced, which is the evidence of the increased risk of unbalanced offspring in carriers of Robertsonian translocations. Our results highlight the importance of genetic counseling of Robertsonian translocation carriers prior to ICSI or IVF.     

Sperm meiotic segregation, aneuploidy and high risk of delivering an affected offspring in carriers of non-Robertsonian translocation t(13;15)

Purpose

To determine the percentage of unbalanced spermatozoa and an interchromosomal effect in two carriers of balanced translocations t(13;15)(q32;q26) and t(13;15)(q32;p11.2).

Methods

Sperm nuclei analysis by fluorescent in situ hybridization for detection of percentage of unbalanced spermatozoa and sperm with disomy of chromosomes X, Y, 8, 18, 21 and diploidy.

Results

The incidence of unbalanced spermatozoa was 50.5 % and 44.6 % in patient 1 (P1) and patient 2 (P2), respectively. Partial disomy of chromosome 13 was detected in 13.4 % and 21.3 % of sperm in P1 and P2, respectively. The unbalanced karyotype der(15)t(13;15) was found previously in a son of P1 and in two adult relatives, and prenatally in the family of P2. This demonstrates a high risk of delivering an affected offspring. Significantly increased frequencies of chromosomes 8, 18, X and XY disomy and diploidy were observed in P2, which might either indicate an interchromosomal effect or be related to his asthenoteratozoospermia.

Conclusions

Since the proportions of unbalanced spermatozoa and the risk of delivering an affected offspring are high, prenatal or preimplantation genetic diagnosis is recommended for such patients.     

Meiotic segregation of a t(7;8)(q11.21;cen) translocation in two carrier brothers

OBJECTIVE: To determine the meiotic segregation of a t(7;8)(q11.21;cen) translocation in two carrier brothers. DESIGN: Analysis of sperm nuclei by fluorescence in situ hybridization (FISH). SETTING: Franche-Comté University Fertility Center in Besan?on, France. PATIENT(S): Two oligospermic brothers with a (7;8) translocation. INTERVENTION(S): Dual FISH for chromosomes 7 and 8, and multicolor FISH for chromosomes X, Y, 15, and 18 were performed. MAIN OUTCOME MEASURE(S): Meiotic segregation differences between both brothers and controls. RESULT(S): In dual FISH 7-8, a total of 34527 (older brother) and 10023 (younger brother) spermatozoa were analyzed. The frequencies of alternate, adjacent 1, adjacent 2, and 3:1 segregations and diploidies were 56.7%, 25.1%, 11.1%, 7.06%, and 0.04%, respectively, in the older brother and 62.84%, 17.61%, 12.8%, 6.47%, and 0.28% in the younger. Also, the disomy rates of some chromosomes not implicated in the translocation would suggest a possible interchromosomal effect. CONCLUSION(S): The similar segregation profiles for the same translocation, compared with those very divergent profiles of the other cases of studied translocations that were published in the literature, confirm that the risks of meiotic imbalances vary primarily according to the characteristics of the chromosomes involved in the rearrangement and the breakpoint position.     

Analysis of meiotic segregation patterns and interchromosomal effects in sperm from six males with Robertsonian translocations

Purpose To provide more genetic information about meiotic segregation behavior and the possibility of interchromosomal effects (ICE) in spermatozoa from carriers of Robertsonian (Rob) translocations. Materials and methods Meiotic segregation behavior in spermatozoa from six carriers of Rob translocations, four t(13;14), one t(14;22) and one t(13;21), was investigated by dual fluorescence in-situ hybridization (FISH). Aneuploidy for chromosomes 18, X and Y was studied by triple FISH. Results The rate of normal/balanced spermatozoa resulting from alternate segregation ranged from 78.14 to 86.88%. The frequency of unbalanced spermatozoa resulting from adjacent segregation varied between 11.70 and 19.53%. The higher frequencies of aneuploidy for sex chromosome were observed in three Rob translocation carriers. In addition, the increased rates of diploid were found in two t(13;14) carriers. Conclusions Alternate segregation is dominant in the different types of Rob translocations. Some carriers may be at an increased risk for ICE. Electronic Supplementary Material The online version of this article (doi:) contains supplementary material, which is available to authorized users. An increased aneuploidy for sex chromosome observed in three Robertsonian translocation carriers suggests that an interchromosomal effect is likely in some carriers.     

Infertile spermatozoa in a human carrier of robertsonian translocation 14;22

OBJECTIVE: To present the ultrastructural, functional, and chromosomal analyses of spermatozoa from an infertile man with normal phenotype and chromosomal translocation 14;22. DESIGN: Case report. SETTING: Regional Reference Center for Male Infertility in Siena, Italy. PATIENT(S): A 36-year-old man with primary infertility for 3 years and his parents. INTERVENTION(S): Family history and lymphocytic karyotypes, physical and hormonal assays, and semen analysis. MAIN OUTCOME MEASURE(S): Morphological sperm evaluation was performed by light, fluorescent, and electron microscopy; chromosomal constitution was examined by the fluorescence in situ hybridization (FISH) technique. The penetration ability of spermatozoa was checked by the hamster test. RESULT(S): The spermatozoa of the patient showed unusual ultrastructural defects. The nuclei were large, spheroidal, and generally uncondensed; the acrosomes were frequently absent or reduced; and the axonemes were often devoid of dynein arms or central singlet tubules. These characteristics are related to immaturity. The lymphocytic karyotype revealed a robertsonian translocation 14;22 in the sterile patient and his mother. FISH sperm analysis demonstrated a high frequency of diploidy for the chromosome 18,XY. The hamster penetration test gave negative results. CONCLUSION(S): The unusual structural sperm immaturity is associated with the translocation 14;22. This chromosomal anomaly may therefore negatively influence the spermatogenesis; an interchromosomal effect on meiosis segregation is also suggested.     

Higher degree of chromosome mosaicism in preimplantation embryos from carriers of robertsonian translocation t(13;14) in comparison with embryos from karyotypically normal IVF patients

Purpose : To compare the frequency and the degree of mosaicism in human embryos from Robertsonian translocation (RT) t(13;14) carriers, with embryos from karyotypically normal IVF patients. Methods : FISH analysis of embryos from PGD cycles for RT t(13;14), with probes for chromosomes 13, 14, and 18 (Group I) and of embryos from karyotypically normal IVF patients with probes for chromosomes 13, 18, 21, X, and Y (Group II). Results : The incidence of abnormal mosaic embryos was significantly higher in group I (38/51) as compared with group II (6/45) (²: P < 0.01). Furthermore, in group I the percentage of diploid cells per embryo was lower for chromosome 13 and 14 in comparison with 18, while in group II no differences were observed between the five chromosomes analyzed. Conclusions : RT induces a high frequency of mosaicism specifically for the chromosomes implicated in the translocation; the analysis by FISH of two blastomeres is strongly recommended for these patients.     

Clinical significance of 45XX, t(13q14Q) Robertsonian translocation: unexpected triploidy in third abortion of female carrier

A case report is described of a female t(13q14q) Robertsonian translocation carrier. Her firstborn daughter appeared to be a carrier of the same translocation. Chromosomal investigation of the third of three subsequent spontaneous abortions revealed a triploidy (69XXX). Literature shows discrepancies in the reported abortion rate in the reproductive performance of t(13q14q) carriers. It is concluded that these could in part be explained by heterogenicity of the study groups due to variable presence of other factors known to influence the abortion risk. In our case report treatment for epilepsy, a luteal phase defect and high numbers of spermatozoa in the sperm count of the husband must be considered as contributory factors. It is recommended to perform systematic investigations in all cases of recurrent abortion even when a chromosomal anomaly is found in one of the partners.     

Sequential FISH allows the determination of the segregation outcome and the presence of numerical anomalies in spermatozoa from a t(1;8;2)(q42;p21;p15) carrier

Purpose

To find out the meiotic segregation behaviour of the t(1;8;2)(q42;p21;p15), to evaluate the occurrence of interchromosomal effects, and to determine whether there is an accumulation of unbalanced products in aneuploid/diploid gametes.

Methods

A sequential FISH protocol based on two successive hybridization rounds over the same spermatozoa was performed to determine the segregation outcome of the rearranged chromosomes. The presence of numerical abnormalities for 13, 18, 21, X and Y was also evaluated by sperm FISH. Those aneuploid/diploid gametes were subsequently relocalized and analyzed for their segregation content through additional hybridization rounds.

Results

The segregation pattern observed reported a very low production of normal/balanced gametes (11.7 %). Significant increased frequencies of diploidies and disomies for chromosomes X/Y and 18 were detected (p < 0.001). Aneuploid and diploid spermatozoa displayed significant increases of 5:1, 6:0 and other unexpected disjunction modes (p < 0.001).

Conclusions

The strategy developed in this study is a reliable new approach to establish the full segregation pattern of complex chromosome rearrangements (CCR). Results corroborate the low number of normal/balanced spermatozoa produced by CCR carriers and support previous findings regarding an altered segregation pattern in gametes with numerical abnormalities. Altogether this confirms the importance of PGD as a tool to prevent the transmission of chromosomal abnormalities to the offspring in CCR patients.     

Balanced chromosomal translocations in men: relationships among semen parameters, chromatin integrity, sperm meiotic segregation and aneuploidy

Purpose

To analyse relationships between semen parameters, sperm chromatin integrity and frequencies of chromosomally unbalanced, disomic and diploid sperm in 13 Robertsonian and 37 reciprocal translocation carriers and to compare the results with data from 10 control donors.

Methods

Conventional semen analysis, Sperm Chromatin Structure Assay and FISH with probes for chromosomes involved in the individual translocations and for chromosomes X, Y, 7, 8, 13, 18 and 21.

Results

Normal semen parameters were found in 30.8 % of Robertsonian and 59.5 % of reciprocal translocation carriers. The rates of unbalanced sperm were 12.0 % in Robertsonian and 55.1 % in reciprocal translocation carriers with no difference between normospermic patients and those showing altered semen parameters. Significantly increased frequencies of spermatozoa showing defects in chromatin integrity and condensation, aneuploidy for chromosomes not involved in a translocation and diploidy were detected in translocation carriers with abnormal semen parameters. Normospermic reciprocal translocation carriers showed an increase in chromosome 13 disomy compared to the control group. There was no relationship between gametic and somatic aneuploidy in 12 translocation carriers studied by FISH on sperm and lymphocytes. The frequency of motile sperm was negatively correlated with the frequency of sperm showing disomy, diploidy and defective chromatin condensation.

Conclusions

Abnormal semen parameters can serve as indicators of an additional risk of forming spermatozoa with defective chromatin and aneuploidy in translocation carriers.     

Molecular cytogenetic characterization of a de novo small supernumerary marker chromosome derived from chromosome 15 in a pregnancy with incidental detection of a maternal Robertsonian translocation of 45,XX,der(13;14) (q10;q10)

ObjectiveWe present molecular cytogenetic characterization of a small supernumerary marker chromosome (sSMC) derived from chromosome 15 in a pregnancy with incidental detection of a maternal Robertsonian translocation of 45,XX,der(13; 14) (q10; q10).Case reportA 37-year-old, primigravid woman underwent amniocentesis at 16 weeks of gestation because of advanced maternal age. Amniocentesis revealed a karyotype of 47,XY,+mar. Simultaneous array comparative genomic hybridization (aCGH) analysis on the DNA extracted from uncultured amniocytes showed the result of no genomic imbalance or arr (1–22) × 2, (X,Y) × 1. Cytogenetic analysis of the parents showed a karyotype of 45,XX,der(13; 14) (q10; q10) in the mother and a karyotype of 46,XY in the father. Prenatal ultrasound was unremarkable. At 38 weeks of gestation, a 2790-g phenotypically normal male baby was delivered. The cord blood had a karyotype of 47,XY,+mar. Metaphase fluorescence in situ hybridization (FISH) analysis showed the result of +mar.ish dic(15) (D15Z1++, SNRPN-, PML-) (18/20). The extra chromosome was derived from chromosome 15.ConclusionMetaphase FISH analysis is useful for the identification of the origin of an sSMC in the presence of no genomic imbalance at aCGH analysis. Prenatal diagnosis of a de novo sSMC may be associated with a Robertsonian translocation in the parents, and parental cytogenetic analysis is necessary under such a circumstance.     

Meiotic segregation and interchromosomal effects in a rare (1:2:10) complex chromosomal rearrangement

Purpose  

Complex chromosomal rearrangements (CCR) are rare rearrangements involving more than two chromosomes and more than two breakpoints. CCR are associated with male infertility as a result of the disruption of spermatogenesis due to complex meiotic configurations and the production of chromosomally abnormal sperm. We examined a carrier of a t(1:2:10) CCR in order to determine the patterns of segregation and any presence of an interchromosomal effect (ICE).     

Preimplantation genetic diagnosis by fluorescence in situ hybridization of reciprocal and Robertsonian translocations

ObjectiveThe presence of reciprocal and Robertsonian chromosomal rearrangement is often related to recurrent miscarriage. Using preimplantation genetic diagnosis, the abortion rate can be decreased. Cases treated at our center were reviewed.Materials and methodsA retrospective analysis for either Robertsonian or reciprocal translocations was performed on all completed cycles of preimplantation genetic diagnosis at our center since the first reported case in 2004 until the end of 2010. Day 3 embryo biopsies were carried out, and the biopsied cell was checked by fluorescent in situ hybridization using relevant informative probes. Embryos with a normal or balanced translocation karyotype were transferred on Day 4.ResultsThirty-eight preimplantation genetic diagnosis cycles involving 17 couples were completed. A total of 450 (82.6%) of the total oocytes were MII oocytes, and 158 (60.0%) of the two-pronuclei embryos were biopsied. In 41.4% of the fluorescent in situ hybridization analyses, the results were either normal or balanced. Embryos were transferred back after 21 cycles. Three babies were born from Robertsonian translocation carriers and another two from reciprocal translocation carriers. The miscarriage rate was 0%. Among the reciprocal translocation group, the live delivery rate was 8.3% per ovum pick-up cycle and 18.2% per embryo transfer cycle. Among the Robertsonian translocation group, the live delivery rate was 14.3% per ovum pick-up cycle and 20.0% per embryo transfer cycle.ConclusionThere is a trend whereby the outcome for Robertsonian translocation group carriers is better than that for reciprocal translocation group carriers. Aneuploidy screening may possibly be added in order to improve the outcome, especially for individuals with an advanced maternal age. The emergence of an array-based technology should help improve this type of analysis.