Abstract
Purpose
To evaluate whether the morphologically normal spermatozoa selected for intracytoplasmic sperm injection (ICSI) under microscope had a higher rate of normal/balanced chromosome contents than that in the whole unselected sperm from reciprocal translocation carriers.
Methods
Five hundred unselected spermatozoa from each of 40 male translocation carriers were performed with fluorescence in situ hybridization (FISH), to determine the rates of gametes with different meiotic contents of translocated chromosomes. Meanwhile, 3030 biopsied blastocysts from 239 male and 293 female reciprocal translocation carriers were detected with the microarray technique to analyze the rates of embryos with different translocated chromosome contents.
Results
The D3 embryo rate, blastocyst formation rate, and euploid rate of blastocysts were remarkably higher in male carriers than those in female (p = 0.001, p = 0.004, and p = 0.035, respectively). In addition, the percentages of alternate products, which contained normal/balanced chromosome contents, in embryos from male carriers were markedly higher than those in sperm FISH (p = 2.48 × 10−5 and p = 2.88 × 10−10), while the percentages of adjacent-2 and 3:1 products were lower than those in sperm FISH (p = 0.003 and p = 5.28 × 10−44). Moreover, consistent results were obtained when comparing the rates of products in embryos between male and female carriers. Specifically, the incidence of alternate products in male carriers was higher than those in female carriers (p = 0.022). However, no similar differences were seen between sperm and embryos of female carriers.
Conclusion
ICSI facilitates the selection of spermatozoa with normal/balanced chromosome contents and improves the D3 embryo rate, blastocyst formation rate, and the euploid embryo rate in male carriers.
Supplementary Information
The online version contains supplementary material available at 10.1007/s10815-020-02013-z.
Keywords: Male reciprocal translocation, Sperm FISH, ICSI, Quadrivalent structure, Segregation patterns, Alternate products
Introduction
Palermo et al. had first reported the microscopic selection of morphologically normal spermatozoon (magnifications: × 200/400) to be used in intracytoplasmic sperm injection (ICSI) [1]. Typically, introducing the ICSI procedures into the assisted reproduction techniques has marked an important milestone in the development of that field. Currently, ICSI is deemed as an efficient method of micromanipulation assisted fertilization, by which one motile and normal-looking spermatozoon is selected, aspirated into a microinjection needle and injected into the oocyte cytoplasm. With this procedure, patients with oligospermia or asthenospermia can achieve pregnancy successfully [1]. Notably, the microscopic selection of sperm is processed based on the judgment of an embryologist who conducts this operation under a microscope. Sperm morphology evaluation plays a crucial role in diagnosing male fertility potential and has a predictive value for the fertilization and pregnancy outcomes in IVF treatments [2].
Reciprocal translocations are common structural chromosome rearrangements, with an estimated prevalence of 0.14% in newborns and 0.6% in infertile population [3]. Male translocation carriers usually present with azoospermia or oligospermia, and female translocation carriers may develop premature ovarian failure [4, 5]. These carriers may be encountered with the fertility problems, mainly as a consequence of producing unbalanced gametes [6, 7]. During the pachytene stage of prophase I, these two translocated chromosomes have formed a quadrivalent structure with the corresponding normal chromosomes, which allows for the pairing of homologous regions. At anaphase I, the meiosis segregation of this structure will lead to the formation of either balanced (alternate segregation mode) or unbalanced (adjacent-1, adjacent-2, 3:1 and 4:0 segregation modes) gametes. To be specific, a total of 32 aspects of gametes can be generated when taking the crossover and non-disjunction into consideration [8]. Notably, among all these possible gametes, only 2 from the alternate segregation mode have normal or balanced karyotype, whereas the others are genetically unbalanced. Typically, fluorescence in situ hybridization (FISH) using appropriate probes is a useful technique to predict the rate of each class of segregation products and to evaluate the prognosis for male translocation carriers [9, 10]. For translocation carriers, it is recommended that embryos with normal/balanced chromosome content be selected for transfer through preimplantation genetic testing (PGT) to achieve successful pregnancies with ICSI procedures [11, 12], which is the routine procedure in reproductive centers. ICSI prevents sperm around the embryo’s zona pellucida from interfering with genetic testing, just as IVF does.
Chelli et al. reported that high-magnification motile sperm organelle morphology examination (MSOME) or ICSI could not improve the selection of normal/balanced spermatozoa compared with that of the whole sperm in patients with heterozygous reciprocal translocations [13]. Nonetheless, only a limited number of patients were enrolled in that study. On the contrary, Wilding et al. suggested that MSOME during ICSI procedures could deselect the physiologically abnormal by testing the level of DNA fragmentation using TUNEL assay and improved clinical results [14]. Besides, other researches described that the microinjection into oocytes of motile spermatozoa with morphologically normal nuclei, which was strictly defined by high-power light microscopy (> × 6000), could improve the IVF/ICSI pregnancy rate for patients experiencing repeated failures or with severe oligozoospermia [15–17]. The research in this field is far from sufficient.
However, the problem of whether the morphologically normal spermatozoon selected by conventional ICSI has a higher rate of normal/balanced chromosomes than that of the whole unselected sperm and can thereby improve the clinical outcomes in male reciprocal translocation carriers remains unclear so far. In this study, sperm FISH with appropriate probes was performed to evaluate the rate of gametes with each class of segregation products in the unselected ejaculated spermatozoa from male reciprocal translocation carriers. For male carriers, the gametes of sperm undergo the selection process of ICSI, while for female carriers, the gametes of eggs will not undergo the selection process. Biopsied trophectoderm cells from male and female carriers were detected with single-nucleotide polymorphism microarray (aSNP); then, the rates of gametes with normal/balanced chromosome contents between sperm FISH and embryos were compared to evaluate the effect of ICSI.
Materials and methods
Clinical subjects and recruitment of participants
A total of 532 translocation carriers expected to undergo assisted reproduction were enrolled from Shanghai Ji Ai Genetics & IVF Institute from March 2014 to August 2018, including 239 male and 293 female carriers. A total of 10,673 oocyte-cumulus complexes were retrieved in 767 controlled ovarian hyperstimulation (COH) cycles, 8954 metaphase II oocytes were performed with routine ICSI, and 3030 blastocysts were biopsied. And a total of 20,000 spermatozoa from each of 40 male carriers were analyzed by FISH; some of these male carriers were included in the males undergoing PGT. All translocation carriers had a history of recurrent spontaneous abortions, infertility, or pregnancies with congenital anomalies. Semen was collected in the laboratory by masturbation and liquefied at 37 °C for 30 min. Subsequently, the sperm quality parameters were analyzed according to the World Health Organization recommendations. Written informed consent was obtained from each family before initiating the PGT cycle. The study protocol was approved by the Ethics Committee of Human Subject Research of the Obstetrics and Gynecology Hospital, Fudan University.
FISH probes and procedures
For each patient, two non-commercial subtelomeric probes labeled by spectrum green and spectrum orange, respectively, as well as one commercial centromeric probe labeled by spectrum aqua, were used. Meanwhile, the probes used in each patient were first tested on metaphase chromosomes according to classic cytogenetic and FISH techniques, and the probes with diffuse signals were excluded. Sperm FISH was then carried out as previously described [18]. Briefly, semen samples were acquired by masturbation and incubated until liquefaction. Later, the spermatozoa were isolated using the pure sperm technique (JCD, Lyon, France), diluted in phosphate-buffered saline, and washed three times by centrifugation for 10 min at 750×g. The pellet was resuspended in fresh fixative prepared by methanol and acetic acid (3:1). Afterwards, the spermatozoon suspension was spread onto clean glass slides, followed by air-drying. Then, the sperm nuclei were decondensed and denatured by incubation in 1 M NaOH for 5 min at room temperature, dehydrated in an ethanol gradient (70, 85, and 100%), and air-dried, while the probe mixture was denatured at 72 °C for 10 min. After applying the three-probe mixture onto the slide, the hybridization was performed overnight at 37 °C in a moist chamber. The slides were later washed with 0.4 saline sodium citrate (SSC)/0.1% Tween 20 for 2 min at 72 °C and with 2 SSC/0.1% Tween 20 at room temperature, and then stained with antifade medium containing 4′,6-diamidino-2-phenyl-indole (DAPI).
Later, signals were observed using Leica GSL-120 Cytogenetics Platform (Leica Biosystem, Inc., CA, USA), and 500 unselected spermatozoa were analyzed for each patient. Typically, spermatozoa were identified as carrying two signals of the same probe if the two fluorescent spots were of the same color and of similar size and intensity, separated by a distance of at least one spot, and not connected by a fluorescent bridge, compared with the neighboring germ cells [19–21]. The FISH pattern of a sperm cell carrying a normal or balanced chromosome content consisted of one blue, red, and green spot, respectively, whereas other patterns of spermatozoa were defined as recombinant unbalanced chromosomes.
In vitro fertilization, blastocyst biopsy, and whole genomic amplification
Standard techniques were employed in IVF in Shanghai Ji Ai Genetics & IVF Institute of Obstetrics and Gynecology Hospital Fudan University. Briefly, a single spermatozoon was injected within 4 h after the follicular aspiration; the fertilization state of the embryo was observed 16–18 h after ICSI and then cultured for 5–6 days in IVF lab to develop to the blastocyst stage. For embryos at the blastocyst stage, three to ten cells were removed from the trophectoderm cells, and then, the biopsied cells were placed into polymerase chain reaction (PCR) tubes containing the alkaline denaturation buffer for cell lysis. Whole genomic amplification (WGA) was performed according to the multiple displacement amplification method. Finally, isothermal DNA amplification with phi 29 DNA polymerase was performed (Repli-g single cell kit, QIAGEN GmbH, Hilden, Germany) in accordance with the manufacturer’s protocol at 30 °C for 8 h, and then, the reaction was stopped by incubation at 65 °C for 3 min.
aSNP
SNP genotyping of WGA products were performed through Illumina Human Cyto-12 microarray according to the Infinium chip procedure, as described previously [22], with each bead chip containing approximately 300,000 SNPs. Briefly, the WGA product was amplified again in an isothermal reaction overnight prior to fragmentation. After isopropanol precipitation and resuspension, the samples were hybridized to a bead chip overnight for about 20 h, followed by an automated extension staining process. After staining, the bead chips were scanned on an iScan reader. Afterwards, genotype call rate and molecular karyotype were analyzed automatically using the BlueFuse®-Multi software (Illumina, Inc. San Diego, USA), while the molecular karyotypes were subsequently used to identify the chromosomal copy number variations, including aneuploidy and segmental anomalies. All the SNP data were analyzed and reported by two genetic technicians independently and verified by a senior genetic director in our laboratory. Embryos without any observed unbalance were described as euploid.
Statistical analysis
Data were expressed as mean ± standard deviation or as percentages. All statistical procedures were conducted using the SPSS version 17.0 software (SPSS, Chicago, IL). Differences between the frequency distributions of segregation products were compared using chi-square (χ2) or Fisher’s exact test. Moreover, quantitative clinical characteristics, including carrier’s age and embryo numbers, were compared with the two-sample t test. Additionally, stratified analysis was also carried out to evaluate the potential interaction by age. All p values were presented upon two-sided test, and the level of p < 0.05 was considered statistically significant.
Results
Clinical characteristics
A total of 532 translocation carriers were involved in this study. A total of 10,673 oocyte-cumulus complexes were retrieved in 767 controlled ovarian hyperstimulation (COH) cycles, and 8954 metaphase II oocytes were performed with routine ICSI. The results suggested that the D3 embryo rate and blastocyst formation rate were higher in male carriers than in female carriers (p = 0.001, OR (95%CI) = 1.23 (1.09–1.38); p = 0.004, OR (95%CI) = 1.16 (1.05–1.28), respectively). In addition, trophectoderm biopsy was carried out on 3042 blastocysts, among which 3030 were successfully diagnosed with aSNP, yielding the diagnosis rate of 99.70%. Of all the diagnosed embryos, 874 (28.8%) were identified as euploid, and the rate was markedly higher in male carriers than in female carriers (p = 0.035, OR (95%CI) = 1.19 (1.01–1.39)). There were no statistical differences in other clinical parameters between male and female carriers (Table 1).
Table 1.
Clinical characteristics and results of reciprocal translocation carriers undergoing PGT
| Parameter | Total | Male carriers | Female carries | p value | OR (95%CI) |
|---|---|---|---|---|---|
| Patients | 532 | 239 | 293 | ||
| COH cycles | 767 | 325 | 442 | ||
| Female age (years) | 30.9 ± 4.0 | 31.5 ± 4.5 | 31.0 ± 3.9 | NS | NS |
| Male age (years) | 32.8 ± 5.5 | 32.7 ± 5.2 | 33.0 ± 5.6 | NS | NS |
| Retrieved oocytes | 10,673 | 4485 (13.8 ± 7.2) | 6188 (14.0 ± 6.6) | NS | NS |
| Injected oocytes | 8954 | 3738 (11.5 ± 6.0) | 5216 (11.8 ± 5.8) | NS | NS |
| 2-pronuclei zygotes | 7726 | 3218 (9.9 ± 5.5) | 4508 (10.2 ± 5.2) | NS | NS |
| Fertilization rate (%) | 7726 | 3218 (86.09%) | 4508 (86.43%) | NS | NS |
| D3 embryos | 6334 | 2665 (8.2 ± 4.8) | 3594 (8.1 ± 4.5) | NS | NS |
| D3 embryo rate (%) | 6334 | 2665 (82.82%) | 3594 (79.72%) | 0.001 | 1.23 (1.09–1.38) |
| Biopsied blastocyst | 3042 | 1351 (4.2 ± 3.0) | 1691 (3.8 ± 3.3) | NS | NS |
| Blastocyst formation rate (%) | 3042 | 1351 (50.69%) | 1691 (47.05%) | 0.005 | 1.16 (1.05–1.28) |
| Diagnosed blastocyst | 3030 | 1346 (4.2 ± 3.0) | 1684 (3.8 ± 3.3) | NS | NS |
| Euploid embryo rate (%) | 874 | 413 (30.68%) | 458 (27.20%) | 0.035 | 1.19 (1.01–1.39) |
| ET cycles | 443 | 200 | 243 | ||
| Positive HCG (%) | 223 | 98 (49.0%) | 125 (51.4) | NS | NS |
| Clinical pregnancies (%) | 193 | 85 (42.5%) | 108 (44.4%) | NS | NS |
| Spontaneous abortions (%) | 21 | 7 (3.5%) | 14 (5.7%) | NS | NS |
| Deliveries (%) | 32 | 12 (6.0%) | 20 (8.2%) | NS | NS |
| Ongoing pregnancies (%) | 134 | 60 (30.0%) | 74 (30.5%) | NS | NS |
Sperm parameters
Typically, parameters, including sperm concentration, sperm progressive motility, total sperm count, and total progressively motile sperm count, were lower in male carriers than those in the spouses of female translocation carriers (p = 1.67 × 10−4, p = 8.17 × 10−4, p = 0.004, p = 0.001, respectively) (Supplementary Table 1), while no statistical difference was observed in semen volume. In addition, the sperm parameters were compared between male carriers undergoing sperm FISH and all male carriers, and no significant difference was discovered (Supplementary Table 2).
Sperm FISH analysis
A total of 20,000 spermatozoa from each of 40 male carriers enrolled from PGT patients were performed with the sperm FISH test to evaluate the proportion of all aspects of segregation products of quadrivalent structure. For each patient, 500 unselected spermatozoa were analyzed respectively. The karyotypes, FISH probes used, and the results of segregation products in sperm are shown in Table 2. Figure 1 shows the result of sperm FISH of one reciprocal translocation carrier with the karyotype of 46,XY,t(2;8)(q31;q24.1). The rate of alternate segregation was the highest (37.85% ± 7.69, ranging from 20.2 to 52.5%), followed by adjacent-1 (27.70% ± 8.88, ranging from 11.8 to 45.5%), 3:1 (19.57% ± 9.69, ranging from 4.3 to 38.3%), and adjacent-2 (10.84% ± 6.98, ranging from 1.8 to 30.1%), while 4:0/other segregation was the lowest (6.07% ± 5.73, ranging from 0.3 to 31.3%).
Table 2.
Karyotypes of the patients and the results of sperm FISH in this study
| Patient | Karyotype | Alternate | Adjacent-1 | Adjacent-2 | 3:1 | 4:0/others | FISH probes |
|---|---|---|---|---|---|---|---|
| Patient 1 | 46,XY,t(2;8)(q31;q24.1) | 40.70% | 24.10% | 3.60% | 25.30% | 6.30% | Tel 8q SR, Tel 2q SG,CEP 8 SA |
| Patient 2 | 46,XY,t(5;21)(q22;q11.2) | 39.30% | 12.20% | 14.50% | 25.50% | 8.50% | Tel 21q SR, Tel 5q SG, Tel 5p SA |
| Patient 3 | 46,XY,t(7;18)(p14;q11.2) | 35.10% | 28.60% | 18.80% | 15.60% | 2.00% | Tel 7p SR, Tel 18q SG,CEP 7 SA |
| Patient 4 | 46,XY,t(8;15)(q24.1;q22.1) | 45.20% | 36.10% | 3.90% | 12.50% | 2.30% | Tel 15q SR, Tel 8q SG, CEP 8 SA |
| Patient 5 | 46,XY,t(11;22)(q25;q13.1) | 27.40% | 17.60% | 5.40% | 38.30% | 11.40% | Tel 11q SR, Tel 22q SG, CEP 11 SA |
| Patient 6 | 46,XY,t(3;6)(q27;q15) | 42.40% | 26.80% | 7.60% | 21.10% | 2.10% | Tel 3q SR, Tel 6q SG,CEP 6 SA |
| Patient 7 | 46,XY,t(3;11)(p14;q22) | 39.40% | 24.00% | 9.90% | 25.00% | 1.70% | Tel 3p SR, Tel 11q SG, CEP 11 SA |
| Patient 8 | 46,XY,t(4;5)(q35;q31) | 29.50% | 17.80% | 22.20% | 16.50% | 14.00% | Tel 4q SR, Tel 5q SG, CEP 4 SA |
| Patient 9 | 46,XY,t(10;18)(q26;q21.1) | 36.00% | 38.00% | 10.30% | 13.60% | 2.20% | Tel 18q SR, Tel 10q SG, CEP 10 SA |
| Patient 10 | 46,XY,t(1;4)(q43;p13) | 41.10% | 29.80% | 11.30% | 16.70% | 1.20% | Tel 1q SR, Tel 4p SG, CEP 4 SA |
| Patient 11 | 46,XY,t(9;17)(p24;q11.2) | 41.30% | 30.20% | 11.40% | 16.00% | 1.20% | Tel 17q SR, Tel 9p SG, CEP 9 SA |
| Patient 12 | 46,XY,t(5;15)(p15.2;q11.2) | 44.60% | 26.60% | 8.20% | 16.50% | 4.10% | Tel 15q SR, Tel 5p SG, CEP 15 SA |
| Patient 13 | 46,XY,t(1;5)(p13;q30) | 22.20% | 15.10% | 18.60% | 32.30% | 11.80% | Tel 5q SR, Tel 1p SG, Tel 5p SA |
| Patient 14 | 46,XY,t(11;17)(q13;q25) | 39.80% | 31.20% | 8.40% | 18.50% | 2.10% | Tel 1p SR, Tel 17q SG, CEP 11 SA |
| Patient 15 | 46,XY,t(4;21)(q31.3;q22.3) | 30.60% | 31.40% | 18.30% | 17.90% | 1.70% | Tel 4q SR, Tel 21q SG, CEP 4 SA |
| Patient 16 | 46,XY,t(2;18)(q35;q21.1) | 37.30% | 32.20% | 4.10% | 22.40% | 4.10% | Tel 2q SR, Tel 18q SG, CEP 18 SA |
| Patient 17 | 46,XY,t(3;11)(q29;p13) | 34.20% | 21.10% | 6.60% | 32.40% | 5.80% | Tel 3q SR, Tel 11p SG,CEP 11 SA |
| Patient 18 | 46,XY,t(5;13)(p15.3;q22) | 30.20% | 11.80% | 16.80% | 35.70% | 5.60% | Tel 5q SR, Tel 13q SG, Tel 5p SA |
| Patient 19 | 46,XY,t(11;19)(p13;q11) | 52.30% | 12.70% | 21.40% | 13.40% | 0.30% | Tel 19q SR, Tel 11p SG, CEP 11 SA |
| Patient 20 | 46,XY,t(13;16)(q31.1;q12.1) | 36.30% | 22.70% | 7.00% | 30.00% | 4.10% | Tel 16q SR, Tel 13q SG, CEP 16SA |
| Patient 21 | 46,XY,t(5;7)(q35;q32) | 43.20% | 28.30% | 5.10% | 19.70% | 3.70% | Tel 7q SR, Tel 5q SG, CEP 7 SA |
| Patient 22 | 46,XY,t(4;5)(q31.3;q33.3) | 24.70% | 14.40% | 28.30% | 14.30% | 18.40% | Tel 4q SR, Tel 5q SG, CEP 4 SA |
| Patient 23 | 46,XY,t(2;10)(p16;p14) | 46.30% | 26.70% | 14.40% | 10.30% | 2.40% | Tel 2p SR, Tel 10p SG,CEP 10 SA |
| Patient 24 | 46,XY,t(14;16)(q22;q22) | 38.10% | 18.40% | 9.30% | 31.70% | 2.60% | Tel 16q SR, Tel 14q SG, CEP 16 SA |
| Patient 25 | 46,XY,t(2;6)(p21;q23) | 32.60% | 27.50% | 12.90% | 22.30% | 4.80% | Tel 6q SR, Tel 2p SG, CEP 6 SA |
| Patient 26 | 46,XY,t(2;13)(p21;q14) | 32.20% | 31.60% | 8.80% | 17.90% | 9.50% | Tel 13q SR, Tel 2p SG, CEP 2 SA |
| Patient 27 | 46,XY,t(10;21)(q11.22;q11.2) | 32.80% | 17.20% | 15.80% | 31.50% | 2.80% | Tel 21q SR, Tel 10q SG, CEP 10 SA |
| Patient 28 | 46,XY,t(1;4)(p32;p16) | 38.10% | 19.40% | 3.20% | 35.80% | 3.50% | Tel 4p SR, Tel 1p SG, CEP 4 SA |
| Patient 29 | 46,XY,t(2;18)(q32;q22) | 37.30% | 23.80% | 6.20% | 29.40% | 3.40% | Tel 18q SR, Tel 2q SG, CEP 18 SA |
| Patient 30 | 46,XY,t(8;11)(p23.3;q22.3) | 44.60% | 38.60% | 7.20% | 4.60% | 4.90% | Tel 11q SR, Tel 8p SG, CEP 11 SA |
| Patient 31 | 46,XY,t(3;5)(q27;q31) | 49.00% | 39.40% | 1.8% | 4.50% | 5.3% | Tel 3q SR, Tel 5q SG, CEP 3 SA |
| Patient 32 | 46,xy,t(4;8)(q21;p22) | 20.20% | 12.00% | 30.10% | 6.40% | 31.30% | Tel 4q SR, Tel 8p SG, CEP 4 SA |
| Patient 33 | 46,XY,t(9;15)(p24;q22) | 26.90% | 31.70% | 7.00% | 21.80% | 12.60% | Tel 15q SR, Tel 9p SG, CEP9 SA |
| Patient 34 | 46,XY,t(4;8)(q21;p23) | 41.20% | 24.70% | 14.40% | 9.60% | 10.10% | Tel 4q SR, Tel 8p SG, CEP 4 SA |
| Patient 35 | 46,XY,t(7;18)(q32;q12.2) | 51.20% | 39.80% | 2.30% | 4.30% | 2.40% | Tel 18q SR, Tel 7q SG, CEP18 SA |
| Patient 36 | 46,XY,t(4;7)(q31.3;q32) | 40.10% | 39.50% | 4.00% | 9.90% | 6.50% | Tel 4q SR, Tel 7q SG, CEP 7 SA |
| Patient 37 | 46,XY,t(6;12)(q25;q15) | 40.80% | 45.50% | 2.00% | 7.20% | 4.40% | Tel 12q SR, Tel 6q SG, CEP 6 SA |
| Patient 38 | 46,XY,t(6;21)(q14;q22) | 52.50% | 19.80% | 16.20% | 4.40% | 7.10% | Tel 21q SR, Tel 6q SG, CEP 6 SA |
| Patient 39 | 46,XY,t(3;14)(q26.3;q23) | 37.40% | 15.40% | 8.90% | 29.80% | 8.70% | Tel 3q SR, Tel 14q SG, CEP 3 SA |
| Patient 40 | 46,XY,t(6;18)(q23;q23) | 39.90% | 24.40% | 7.50% | 22.20% | 6.00% | Tel 6q SR, Tel 18q SG, CEP 6 SA |
FISH, fluorescent in situ hybridization; SO, spectrum red; SG, spectrum green; SA, spectrum aqua; CEP, centromeric probe; LSI, locus specific probe; Tel, subtelomeric probe. All probes were from Abbott Molecular, Inc.
Fig. 1.
Sperm FISH in one reciprocal translocation carrier, 46,XY,t(2;8)(q31;q24.1). The five types of unbalanced sperm cells: a alternate segregation pattern; b adjacent-1 segregation pattern; c adjacent-2 segregation pattern; d 3:1 segregation pattern; e 4:0 segregation pattern
Embryo meiotic segregation analysis
The meiotic segregation pattern was determined by means of the molecular karyotype of embryos. Specifically, a total of 2503 embryos were produced by the 2:2 segregation pattern, among which alternate was the most common pattern (1252/2503, 50.0%), followed by adjacent-1 (909/2503, 36.3%) and adjacent-2 (342/2503, 13.7%). In addition, the frequencies of 3:1 and 4:0/other segregation patterns were 6.7% (202/3030) and 10.7% (325/3030), respectively.
When comparing the meiotic segregation products of quadrivalent structure between male sperm FISH and male embryos, we found that the percentages of alternate and adjacent-1 products in male carriers were evidently higher than those in sperm (p = 2.48 × 10−5, OR (95%CI) = 1.27 (1.14–1.42); p = 2.88 × 10−10, OR (95%CI) = 1.46 (1.30–1.64), respectively), while those of adjacent-2 and 3:1 products were lower (p = 0.003, OR (95%CI) = 0.74 (0.61–0.90); p = 5.28 × 10−44, OR (95%CI) = 0.19 (0.14–0.24), respectively) (Table 3). Through stratified analysis by age, the distributions of alternate and 3:1 products were roughly similar to the whole, while the statistical difference of adjacent and 4:0/others was not shown in elder carriers.
Table 3.
Comparison of meiotic segregation products of quadrivalent structure between sperm FISH and male embryos
| Segregation products | Sperm FISH, n (%) | Embryos of male carriers, n (%) | p value | OR (95%CI) |
|---|---|---|---|---|
| Overall | 20,000 | 1346 | ||
| Alternate | 7568 (37.85%) | 587 (43.61%) | p = 2.48 × 10−5 | 1.27 (1.14–1.42) |
| Adjacent-1 | 5140 (25.70%) | 451 (33.51%) | p = 2.88 × 10−10 | 1.46 (1.30–1.64) |
| Adjacent-2 | 2168 (10.84%) | 111 (8.25%) | p = 0.003 | 0.74 (0.61–0.90) |
| 3:1 | 3910 (19.55%) | 58 (4.31%) | p = 5.28 × 10−44 | 0.19 (0.14–0.24) |
| 4:0/others | 1214 (6.07%) | 139 (10.33%) | p = 5.50 × 10−10 | 1.78 (1.48–2.14) |
| < 35 years | ||||
| Alternate | 6256 (37.92%) | 516 (43.25%) | p = 9.78 × 10−5 | 1.27 (1.12–1.43) |
| Adjacent-1 | 4374 (26.51%) | 411 (34.45%) | p = 2.47 × 10−9 | 1.46 (1.29–1.65) |
| Adjacent-2 | 1706 (10.34%) | 97 (8.13%) | p = 0.015 | 0.77 (0.62–0.95) |
| 3:1 | 3220 (19.52%) | 46 (3.86%) | p = 2.58 × 10−41 | 0.17 (0.12–0.22) |
| 4:0/others | 944 (5.72%) | 123 (10.31%) | p = 1.28 × 10−10 | 1.90 (1.55–2.31) |
| ≥ 35 years | ||||
| Alternate | 1314 (37.54%) | 71 (46.41%) | p = 0.027 | 1.44 (1.04–1.99) |
| Adjacent-1 | 767 (21.91%) | 40 (26.14%) | p = NS | 1.26 (0.87–1.83) |
| Adjacent-2 | 463 (13.23%) | 14 (9.15%) | p = NS | 0.66 (0.38–1.16) |
| 3:1 | 686 (19.62%) | 12 (7.84%) | p = 2.94 × 10−4 | 0.35 (0.19–0.63) |
| 4:0/others | 270 (7.71%) | 16 (10.46%) | p = NS | 1.40 (0.82–2.38) |
When conducting the comparison of segregation products in embryos between male and female carriers, consistent results were observed. The incidences of alternate and adjacent-1 products in male carriers were higher in comparison with female carriers (p = 0.022, OR (95%CI) = 1.19 (1.03–1.37); p = 1.66 × 10−4, OR (95%CI) = 1.35 (1.15–1.58), respectively); on the contrary, those of adjacent-2 and 3:1 products were lower in male carriers than in female carriers (p = 2.26 × 10−6, OR (95%CI) = 0.57 (0.45–0.72); p = 3.30 × 10−6, OR (95%CI) = 0.48 (0.35–0.66), respectively) (Table 4). Stratified analysis by age revealed that the distribution was also significantly different in young carriers except for 4:0/other segregation, but the difference was not obvious in elder carriers.
Table 4.
Comparison of meiotic segregation products of quadrivalent structure between female and male carriers
| Segregation products | Embryos of female carriers, n (%) | Embryos of male carriers, n (%) | p value | OR (95%CI) |
|---|---|---|---|---|
| Overall | 1684 | 1346 | ||
| Alternate | 665 (39.49%) | 587 (43.61%) | p = 0.022 | 1.19 (1.03–1.37) |
| Adjacent-1 | 458 (27.20%) | 451 (33.51%) | p = 1.66 × 10−4 | 1.35 (1.15–1.58) |
| Adjacent-2 | 231 (13.72%) | 111 (8.25%) | p = 2.26 × 10−6 | 0.57 (0.45–0.72) |
| 3:1 | 144 (8.55%) | 58 (4.31%) | p = 3.30 × 10−6 | 0.48 (0.35–0.66) |
| 4:0/others | 186 (11.05%) | 139 (10.33%) | NS | 0.93 (0.74–1.17) |
| < 35 years | ||||
| Alternate | 557 (39.09%) | 516 (43.25%) | p = 0.031 | 1.19 (1.02–1.39) |
| Adjacent-1 | 402 (28.21%) | 411 (34.45%) | p = 5.89 × 10−4 | 1.34 (1.13–1.58) |
| Adjacent-2 | 205 (14.39%) | 97 (8.13%) | p = 6.04 × 10−7 | 0.53 (0.41–0.68) |
| 3:1 | 123 (8.63%) | 46 (3.86%) | p = 7.33 × 10−7 | 0.43 (0.30–0.60) |
| 4:0/others | 138 (9.68%) | 123 (10.31%) | NS | 1.07 (0.83–1.39) |
| ≥ 35 years | ||||
| Alternate | 108 (41.70%) | 71 (46.41%) | NS | 1.21 (0.81–1.81) |
| Adjacent-1 | 56 (21.62%) | 40 (26.14%) | NS | 1.28 (0.81–2.05) |
| Adjacent-2 | 26 (10.4%) | 14 (9.15%) | NS | 0.93 (0.46–1.79) |
| 3:1 | 21 (8.11%) | 12 (7.84%) | NS | 0.97 (0.46–2.02) |
| 4:0/others | 48 (18.53%) | 16 (10.46%) | p = 0.029 | 0.51 (0.28–0.94) |
S, not statistically significant
However, by comparing the meiotic segregation products between male sperm FISH and female embryos, no statistical differences were seen in both alternate and adjacent-1 products (Table 5). Figure 2 shows the comparison of alternate segregation product frequency of quadrivalent structure between embryos and sperm in reciprocal translocation carriers.
Table 5.
Comparison of meiotic segregation products of quadrivalent structure between male sperm FISH and female embryos
| Segregation products | Sperm FISH, n (%) | Embryos of female carriers, n (%) | p value | OR (95%CI) |
|---|---|---|---|---|
| Overall | 20,000 | 1684 | ||
| Alternate | 7568 (37.84%) | 665 (39.49%) | NS | 1.07 (0.97–1.19) |
| Adjacent-1 | 5140 (25.70%) | 458 (27.20%) | NS | 1.08 (0.97–1.21) |
| Adjacent-2 | 2168 (10.84%) | 231 (13.72%) | p = 3.00 × 10−4 | 1.31 (1.13–1.51) |
| 3:1 | 3910 (19.55%) | 144 (8.55%) | p = 1.02 × 10−28 | 0.39 (0.32–0.46) |
| 4:0/others | 1214 (6.07%) | 186 (11.05%) | p = 1.48 × 10−15 | 1.92 (1.63–2.26) |
| < 35 years | ||||
| Alternate | 6256 (37.92%) | 557 (39.09%) | NS | 1.05 (0.94–1.17) |
| Adjacent-1 | 4374 (26.51%) | 402 (28.21%) | NS | 1.09 (0.97–1.23) |
| Adjacent-2 | 1706 (10.34%) | 205 (14.39%) | p = 2.05 × 10−6 | 1.46 (1.25–1.70) |
| 3:1 | 3220 (19.52%) | 123 (8.63%) | p = 4.51 × 10−24 | 0.39 (0.32–0.47) |
| 4:0/others | 944 (5.72%) | 138 (9.68%) | p = 1.37 × 10−9 | 1.77 (1.47–2.13) |
| ≥ 35 years | ||||
| Alternate | 1314 (37.54%) | 108 (41.70%) | NS | 1.19 (0.92–1.54) |
| Adjacent-1 | 767 (21.91%) | 56 (21.62%) | NS | 0.98 (0.72–1.34) |
| Adjacent-2 | 463 (13.23%) | 26 (10.4%) | NS | 0.73 (0.48–1.11) |
| 3:1 | 686 (19.62%) | 21 (8.11%) | p = 4.95 × 10−6 | 0.36 (0.23–0.57) |
| 4:0/others | 270 (7.71%) | 48 (18.53%) | p = 1.57 × 10−9 | 2.72 (1.94–3.81) |
NS, not statistically significant
Fig. 2.
The comparison of alternate segregation products of quadrivalent structure between reciprocal translocation carriers and sperm FISH. The rate of alternate segregation products which presents normal/balanced chromosomes in embryos of male carriers was significant higher than that in the whole sperm and embryos of female carriers, whereas no difference was found between the whole sperm and female carriers
Discussion
Among the male carriers undergoing PGT treatment, a total of 20,000 spermatozoa from 40 male carriers were analyzed by FISH, to evaluate the proportions of all aspects of segregation products of quadrivalent structure. Sperm FISH using appropriate probes is a useful technique to predict the rate of each class of segregation modes in ejaculated spermatozoa from translocation carriers; meanwhile, the FISH technology is easy to perform and allows for the count of hundreds of sperm cells. In this study, the molecular karyotypes by SNP-array platform of biopsied embryos were analyzed to identify the chromosomal copy number variations, and then, meiotic segregation patterns were subsequently classified. Although numerous meiotic segregation products could be generated, a prevalence of alternate products which contained normal/balanced chromosomes was observed in both sperm and embryo samples in our study, followed by adjacent-1 products. These results were consistent with most previous studies in that homologous centromeres have an increased tendency to migrate to the opposite poles in cells [23–26].
To figure out whether the morphologically normal spermatozoa under microscopic selection by conventional ICSI have a higher rate of normal/balanced chromosomes than that of the whole sperm in reciprocal translocation carriers, the comparison of the segregation products of quadrivalent structure among male sperm and embryos from carriers was performed. We found that the percentages of alternate and adjacent-1 products in the embryos of male carriers were remarkably higher than those in sperm FISH; in contrast, those of adjacent-2 and 3:1 products were lower. Taking into account the possible confounder, we compared the segregation products through stratified analysis by carriers’ age and found that the distribution was roughly similar to the whole. In addition, consistent results were obtained when comparing the embryo products between male and female carriers. To be specific, the incidences of alternate and adjacent-1 products in male carriers were higher than those in female carriers, while those of adjacent-2 and 3:1 products were lower. However, such differences could not be observed in alternate or adjacent-1 products when comparing the segregation products between male sperm FISH and female embryos. For male carriers, the sperm was selected through the ICSI process, while for female carriers, all the eggs were used for fertilization without selection. Then, the comparison of embryos with balanced chromosome contents between male and female could be used to evaluate the effect of ICSI. In addition, the results of sperm FISH obtained from unselected sperm and the comparison of sperm FISH with female embryos with no statistically significant differences can also be used to evaluate the effect of ICSI. Taken together, the above results suggested that the morphologically normal spermatozoon selected by conventional ICSI in male translocation carriers had a higher rate of balanced chromosomes. Notably, the process of ICSI might help to exclude those physiologically abnormal spermatozoa in male carriers, but not to exclude the physiologically abnormal oocytes in female carriers, which could explain this phenomenon. By contrast, Chelli et al. reported that ICSI could not improve the selection of normal/balanced spermatozoa by comparing with the whole sperm in reciprocal translocation carriers [13]. In addition, Casutto et al. and Celik-Ozenco et al. showed that sperm FISH could not reveal the relationship between sperm morphology and sperm chromosomal content [27, 28]. But the two studies had a limited sample size and analyzed only a small number of sperm; thus, selection bias was inevitable. In addition, some other researchers confirmed that MSOME during ICSI procedure could help deselect the physiologically abnormal spermatozoa, and in patients with multiple failures or severe oligozoospermia, microinjection of motile spermatozoa strictly selected by high-power light microscopy can improve the IVF/ICSI pregnancy rates [14–16].
Moreover, in terms of the clinical characteristics, we found that D3 embryo rate and blastocyst formation rate in male carriers were 1.23 and 1.16 times higher than those in female carriers, respectively. Meanwhile, the euploid rate of all the diagnosed blastocysts was higher in male carriers (1.19 times) than that in female carriers. These clinical outcomes were consistent with published data, which contained considerable samples [29, 30]. Although sperm quality is reported to impact embryo development, and the low-quality sperm may lead to a low fertilization rate, slow growth rate, and poor implantation rate during transfer [31, 32], there is no significant difference between the fertilization rate and clinical pregnancy rate among male and female carriers. Our results suggest that ICSI may help to reduce spermatozoa with unbalanced chromosome content in male carriers, thereby increasing the euploid embryo rate. Besides, it is well known that translocation will impair male’s fecundity by decreasing the sperm quantity and quality [33]. Our results showed that sperm parameters, including sperm concentration, sperm progressive motility, total sperm count, and total progressively motile sperm count, were remarkably decreased in male carriers than those in the counterparts of female translocation carriers, which was consistent with the reported results [34, 35]. In contrast, by comparing the sperm parameters of male carriers who received FISH with those of all male carriers, no significant difference was found, indicating that the sperm samples tested with FISH could represent the male carrier population and the possible deviation could be ignored.
One limitation of this study was that the sperm FISH test could not evaluate the abnormalities of other non-translocation chromosomes. The FISH method could not provide an overview of a sperm karyotype; it only allows the analysis of a limited number of chromosomes that were recognized by the probes; the content of additional chromosome numerical abnormalities in sperm could not be evaluated accurately. Previous studies revealed that the interchromosomal effect (ICE) might disturb meiosis in the proper pairing and disjunction of other chromosomes, which could lead to additional chromosome numerical abnormalities unrelated to translocation [36, 37], although other studies suggested that an ICE was either absent or negligible [11, 38]. To date, there is no accessible way at present could overcome such limitation. In addition, our study has some obvious advantages. This is the first report to investigate whether the morphologically normal spermatozoa selected by conventional ICSI have a higher rate of normal/balanced chromosomes than that of the whole sperm by comprehensively analyzing the rates of alternate segregation products between embryos and sperm. Moreover, the large sample size marks another advantage of our research.
In summary, our results indicate that ICSI can help to increase the selection of spermatozoa with normal/balanced chromosome contents and then improve the euploid embryo rate. Such data can provide a better insight into the clinical difference between male and female reciprocal translocation carriers. In addition, these results help to predict the possibility of acquiring euploid embryos and to provide more accurate genetic counseling for each pair of translocation carriers receiving PGT.
Supplementary information
(DOC 50 kb).
(DOC 46 kb).
Acknowledgments
We express deepest gratitude to all the families enrolled in our research. We also thank the staff of the Genetics Laboratory and IVF Laboratory for their invaluable efforts and contribution relating to the experiment.
Author contributions
C.L, S.Z, and X.S designed the research and wrote the manuscript; C.L, S.Z, SJ.Z, J.W, M.X, J.Z, J.F, Y.S X.S, and C.L executed the research and data collection (S.Z, M.X, C.L, and M.X performed the analysis; S.Z, SJ.Z, and J.Z performed the microarray experiments and SJ.Z performed the FISH experiments; J.F and Y.S performed the intra-cytoplasmic sperm injection and blastocyst biopsy experiments; J.W, X.S, and C.L collected the cases). X.S and C.C directed the critical discussion of the manuscript. All authors approved the final manuscript.
Funding
The research was supported by the Science and Technology Innovation Action Plan Program of Shanghai (18411953800) and Shanghai Shen Kang Hospital Development Centre Municipal Hospital New Frontier Technology Joint Project (SHDC12017105).
Compliance with ethical standards
The study protocol was approved by the Ethics Committee of Human Subject Research of the Obstetrics and Gynecology Hospital, Fudan University.
Conflict of interest
The authors declare that they have no conflict of interest.
Informed consent
Informed consents were obtained from all individual participants included in the study.
Footnotes
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Caixia Lei and Shuo Zhang contributed equally to this work.
References
- 1.Palermo G, Joris H, Devroey P, Van Steirteghem AC. Pregnancies after intracytoplasmic injection of single spermatozoon into an oocyte. Lancet. 1992;340(8810):17–18. doi: 10.1016/0140-6736(92)92425-F. [DOI] [PubMed] [Google Scholar]
- 2.Menkveld R, Holleboom CA, Rhemrev JP. Measurement and significance of sperm morphology. Asian J Androl. 2011;13(1):59–68. doi: 10.1038/aja.2010.67. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Clementini E, Palka C, Iezzi I, Stuppia L, Guanciali-Franchi P, Tiboni GM. Prevalence of chromosomal abnormalities in 2078 infertile couples referred for assisted reproductive techniques. Hum Reprod. 2005;20(2):437–442. doi: 10.1093/humrep/deh626. [DOI] [PubMed] [Google Scholar]
- 4.Zhang HG, Wang RX, Li LL, Sun WT, Zhang HY, Liu RZ. Male carriers of balanced reciprocal translocations in Northeast China: sperm count, reproductive performance, and genetic counseling. Genet Mol Res. 2015;14(4):18792–18798. doi: 10.4238/2015.December.28.28. [DOI] [PubMed] [Google Scholar]
- 5.Ananthapur V, Avvari S, Veena K, Sujatha M, Jyothy A. Non-Robertsonian translocation t (2;11) is associated with infertility in an oligospermic man. Andrologia. 2014;46(4):453–455. doi: 10.1111/and.12120. [DOI] [PubMed] [Google Scholar]
- 6.Fiorentino F, Spizzichino L, Bono S, Biricik A, Kokkali G, Rienzi L, Ubaldi FM, Iammarrone E, Gordon A, Pantos K. PGD for reciprocal and Robertsonian translocations using array comparative genomic hybridization. Hum Reprod. 2011;26(7):1925–1935. doi: 10.1093/humrep/der082. [DOI] [PubMed] [Google Scholar]
- 7.Zhang Y, Zhu S, Wu J, Liu S, Sun X. Quadrivalent asymmetry in reciprocal translocation carriers predicts meiotic segregation patterns in cleavage stage embryos. Reprod BioMed Online. 2014;29(4):490–498. doi: 10.1016/j.rbmo.2014.06.010. [DOI] [PubMed] [Google Scholar]
- 8.Scriven PN, Handyside AH, Ogilvie CM. Chromosome translocations: segregation modes and strategies for preimplantation genetic diagnosis. Prenat Diagn. 1998;18(13):1437–1449. doi: 10.1002/(SICI)1097-0223(199812)18:13<1437::AID-PD497>3.0.CO;2-P. [DOI] [PubMed] [Google Scholar]
- 9.Vozdova M, Oracova E, Horinova V, Rubes J. Sperm fluorescence in situ hybridization study of meiotic segregation and an interchromosomal effect in carriers of t(11;18) Hum Reprod. 2008;23(3):581–588. doi: 10.1093/humrep/dem345. [DOI] [PubMed] [Google Scholar]
- 10.Kekesi A, Erdei E, Torok M, Dravucz S, Toth A. Segregation of chromosomes in spermatozoa of four Hungarian translocation carriers. Fertil Steril. 2007;88(1):212–215. doi: 10.1016/j.fertnstert.2006.11.097. [DOI] [PubMed] [Google Scholar]
- 11.Xie Y, Xu Y, Wang J, Miao B, Zeng Y, Ding C, Gao J, Zhou C. Preliminary analysis of numerical chromosome abnormalities in reciprocal and Robertsonian translocation preimplantation genetic diagnosis cases with 24-chromosomal analysis with an aCGH/SNP microarray. J Assist Reprod Genet. 2018;35(1):177–186. doi: 10.1007/s10815-017-1045-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Chow J, Yeung W, Lee V, Lau E, Ng E. Evaluation of preimplantation genetic testing for chromosomal structural rearrangement by a commonly used next generation sequencing workflow. Eur J Obstet Gynecol Reprod Biol. 2018;224:66–73. doi: 10.1016/j.ejogrb.2018.03.013. [DOI] [PubMed] [Google Scholar]
- 13.Chelli MH, Ferfouri F, Boitrelle F, Albert M, Molina-Gomes D, Selva J, Vialard F. High-magnification sperm selection does not decrease the aneuploidy rate in patients who are heterozygous for reciprocal translocations. J Assist Reprod Genet. 2013;30(4):525–530. doi: 10.1007/s10815-013-9959-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 14.Wilding M, Coppola G, di Matteo L, Palagiano A, Fusco E, Dale B. Intracytoplasmic injection of morphologically selected spermatozoa (IMSI) improves outcome after assisted reproduction by deselecting physiologically poor quality spermatozoa. J Assist Reprod Genet. 2011;28(3):253–262. doi: 10.1007/s10815-010-9505-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 15.Bartoov B, Berkovitz A, Eltes F, Kogosovsky A, Yagoda A, Lederman H, Artzi S, Gross M, Barak Y. Pregnancy rates are higher with intracytoplasmic morphologically selected sperm injection than with conventional intracytoplasmic injection. Fertil Steril. 2003;80(6):1413–1419. doi: 10.1016/j.fertnstert.2003.05.016. [DOI] [PubMed] [Google Scholar]
- 16.Antinori M, Licata E, Dani G, Cerusico F, Versaci C, D'Angelo D, et al. Intracytoplasmic morphologically selected sperm injection: a prospective randomized trial. Reprod BioMed Online. 2008;16(6):835–841. doi: 10.1016/S1472-6483(10)60150-2. [DOI] [PubMed] [Google Scholar]
- 17.Calogero AE, Burrello N, De Palma A, Barone N, D'Agata R, Vicari E. Sperm aneuploidy in infertile men. Reprod BioMed Online. 2003;6(3):310–317. doi: 10.1016/S1472-6483(10)61850-0. [DOI] [PubMed] [Google Scholar]
- 18.Chantot-Bastaraud S, Ravel C, Berthaut I, McElreavey K, Bouchard P, Mandelbaum J, Siffroi JP. Sperm-FISH analysis in a pericentric chromosome 1 inversion, 46,XY,inv(1)(p22q42), associated with infertility. Mol Hum Reprod. 2007;13(1):55–59. doi: 10.1093/molehr/gal094. [DOI] [PubMed] [Google Scholar]
- 19.Benet J, Oliver-Bonet M, Cifuentes P, Templado C, Navarro J. Segregation of chromosomes in sperm of reciprocal translocation carriers: a review. Cytogenet Genome Res. 2005;111(3–4):281–290. doi: 10.1159/000086901. [DOI] [PubMed] [Google Scholar]
- 20.Anton E, Vidal F, Blanco J. Role of sperm FISH studies in the genetic reproductive advice of structural reorganization carriers. Hum Reprod. 2007;22(8):2088–2092. doi: 10.1093/humrep/dem152. [DOI] [PubMed] [Google Scholar]
- 21.Martin RH. Cytogenetic determinants of male fertility. Hum Reprod Update. 2008;14(4):379–390. doi: 10.1093/humupd/dmn017. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 22.Fan JB, Hu S, Craumer W, Barker D. BeadArray-based solutions for enabling the promise of pharmacogenomics. Biotechniques. 2005;39(10 Suppl):S583–S588. doi: 10.2144/000112047. [DOI] [PubMed] [Google Scholar]
- 23.Anton E, Vidal F, Blanco J. Reciprocal translocations: tracing their meiotic behavior. Genet Med. 2008;10(10):730–738. doi: 10.1097/GIM.0b013e318187760f. [DOI] [PubMed] [Google Scholar]
- 24.Lledo B, Ortiz JA, Morales R, Ten J, de la Fuente PE, Garcia-Ochoa C, et al. The paternal effect of chromosome translocation carriers observed from meiotic segregation in embryos. Hum Reprod. 2010;25(7):1843–1848. doi: 10.1093/humrep/deq111. [DOI] [PubMed] [Google Scholar]
- 25.Ko DS, Cho JW, Park SY, Kim JY, Koong MK, Song IO, Kang IS, Lim CK. Clinical outcomes of preimplantation genetic diagnosis (PGD) and analysis of meiotic segregation modes in reciprocal translocation carriers. Am J Med Genet A. 2010;152A(6):1428–1433. doi: 10.1002/ajmg.a.33368. [DOI] [PubMed] [Google Scholar]
- 26.Yakut T, Ercelen N, Acar H, Kimya Y, Egeli U. Meiotic segregation analysis of reciprocal translocations both in sperms and blastomeres. Am J Med Genet A. 2006;140(10):1074–1082. doi: 10.1002/ajmg.a.31215. [DOI] [PubMed] [Google Scholar]
- 27.Cassuto NG, Le Foll N, Chantot-Bastaraud S, Balet R, Bouret D, Rouen A, et al. Sperm fluorescence in situ hybridization study in nine men carrying a Robertsonian or a reciprocal translocation: relationship between segregation modes and high-magnification sperm morphology examination. Fertil Steril. 2011;96(4):826–832. doi: 10.1016/j.fertnstert.2011.07.1143. [DOI] [PubMed] [Google Scholar]
- 28.Celik-Ozenci C, Jakab A, Kovacs T, Catalanotti J, Demir R, Bray-Ward P, Ward D, Huszar G. Sperm selection for ICSI: shape properties do not predict the absence or presence of numerical chromosomal aberrations. Hum Reprod. 2004;19(9):2052–2059. doi: 10.1093/humrep/deh361. [DOI] [PubMed] [Google Scholar]
- 29.Keymolen K, Staessen C, Verpoest W, Liebaers I, Bonduelle M. Preimplantation genetic diagnosis in female and male carriers of reciprocal translocations: clinical outcome until delivery of 312 cycles. Eur J Hum Genet. 2012;20(4):376–380. doi: 10.1038/ejhg.2011.208. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 30.Ko DS, Cho JW, Lee HS, Kim JY, Kang IS, Yang KM, Lim CK. Preimplantation genetic diagnosis outcomes and meiotic segregation analysis of robertsonian translocation carriers. Fertil Steril. 2013;99(5):1369–1376. doi: 10.1016/j.fertnstert.2012.12.010. [DOI] [PubMed] [Google Scholar]
- 31.Ishikawa T, Shiotani M, Izumi Y, Hashimoto H, Kokeguchi S, Goto S, Fujisawa M. Fertilization and pregnancy using cryopreserved testicular sperm for intracytoplasmic sperm injection with azoospermia. Fertil Steril. 2009;92(1):174–179. doi: 10.1016/j.fertnstert.2008.04.051. [DOI] [PubMed] [Google Scholar]
- 32.Caseiro AL, Regalo A, Pereira E, Esteves T, Fernandes F, Carvalho J. Implication of sperm chromosomal abnormalities in recurrent abortion and multiple implantation failure. Reprod BioMed Online. 2015;31(4):481–485. doi: 10.1016/j.rbmo.2015.07.001. [DOI] [PubMed] [Google Scholar]
- 33.Vozdova M, Kasikova K, Oracova E, Prinosilova P, Rybar R, Horinova V, et al. The effect of the swim-up and hyaluronan-binding methods on the frequency of abnormal spermatozoa detected by FISH and SCSA in carriers of balanced chromosomal translocations. Hum Reprod. 2012;27(3):930–937. doi: 10.1093/humrep/der445. [DOI] [PubMed] [Google Scholar]
- 34.Yin B, Zhu Y, Wu T, Shen S, Zeng Y, Liang D. Clinical outcomes for couples containing a reciprocal chromosome translocation carrier without preimplantation genetic diagnosis. Int J Gynaecol Obstet. 2017;136(3):304–308. doi: 10.1002/ijgo.12062. [DOI] [PubMed] [Google Scholar]
- 35.Baumgartner A, Van Hummelen P, Lowe XR, Adler ID, Wyrobek AJ. Numerical and structural chromosomal abnormalities detected in human sperm with a combination of multicolor FISH assays. Environ Mol Mutagen. 1999;33(1):49–58. doi: 10.1002/(SICI)1098-2280(1999)33:1<49::AID-EM6>3.0.CO;2-F. [DOI] [PubMed] [Google Scholar]
- 36.Alfarawati S, Fragouli E, Colls P, Wells D. Embryos of robertsonian translocation carriers exhibit a mitotic interchromosomal effect that enhances genetic instability during early development. PLoS Genet. 2012;8(10):e1003025. doi: 10.1371/journal.pgen.1003025. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 37.Zhang S, Lei C, Wu J, Sun H, Zhou J, Zhu S, Wu J, Fu J, Sun Y, Lu D, Sun X, Zhang Y. Analysis of segregation patterns of quadrivalent structures and the effect on genome stability during meiosis in reciprocal translocation carriers. Hum Reprod. 2018;33(4):757–767. doi: 10.1093/humrep/dey036. [DOI] [PubMed] [Google Scholar]
- 38.Estop AM, Cieply K, Munne S, Surti U, Wakim A, Feingold E. Is there an interchromosomal effect in reciprocal translocation carriers? Sperm FISH studies. Hum Genet. 2000;106(5):517–524. doi: 10.1007/s004390000275. [DOI] [PubMed] [Google Scholar]
Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Supplementary Materials
(DOC 50 kb).
(DOC 46 kb).