Assisting reproduction of infertile men carrying a Robertsonian translocation.

BACKGROUND: In order to provide better genetic counselling for Robertsonian translocation carriers, the meiotic segregation of chromosomes 13, 14 and 21 from six infertile (13;14) and (14;21) Robertsonian translocation carriers was examined. METHODS: Dual-colour fluorescence in-situ hybridization analysis using locus-specific probes was carried out on spermatozoa of translocation carriers. Spermatozoa from six proven fertile subjects were analysed using the same probes as controls. RESULTS: We observed that the frequencies of unbalanced spermatozoa were similar in the (13;14) translocation carriers (9.0, 10.0 and 12.9%) and in the (14;21) translocation carriers (8.7, 7.2 and 7.0%). These frequencies were significantly increased compared with the control population (P < 0.05). CONCLUSIONS: This high frequency might justify the use of preimplantation genetic diagnosis in these patients where the translocation is usually associated with infertility, requiring intracytoplasmic sperm injection, as it might improve the outcome of the assisted reproduction technique.     

Sperm segregation analysis of a (13;22) Robertsonian translocation carrier by FISH: a comparison of locus-specific probe and whole chromosome painting

BACKGROUND: The t(13;22) Robertsonian translocation constitutes a rare form of rearrangement between acrocentric human chromosomes. Most of the meiotic segregation studies of human Robertsonian translocations have been performed on common t(13;14) and t(14;21) translocations. Analysis of the chromosomal constitution in sperm of Robertsonian translocation carriers is of great interest for assessing the risk of unbalanced forms and adapting genetic counselling. In the present study, we present the first meiotic segregation study of a t(13;22) Robertsonian translocation in human sperm. METHODS: A total of 11 787 sperm nuclei were scored using two distinct FISH labelling techniques, i.e. the locus-specific probes (LSI) method and the whole chromosome painting (WCP) technique. RESULTS: The frequency of normal or balanced sperm resulting from alternate meiotic segregation was 86%. Incidences of unbalanced complements resulting from adjacent segregation modes were 12.79% and 14.36% in LSI and WCP assays, respectively. No significant excess of nullisomy or disomy for the affected chromosomes was observed. CONCLUSIONS: Similar results in segregation were obtained with the two techniques, demonstrating the efficiency of the two strategies for the direct segregation analysis of Roberstsonian translocations. The results obtained indicated a moderate meiotic production of imbalance. This study shows that the rare Robertsonian translocation (13;22) displays a similar distribution of balanced and unbalanced sperm patterns as the common Robertsonian translocations previously studied. This suggests that the behaviour of acrocentric chromosomes was similar in all cases of centric fusion.     

Study of two markers of apoptosis and meiotic segregation in ejaculated sperm of chromosomal translocation carrier patients

BACKGROUND: To try to explain the infertility of chromosomal translocation carrier patients, we compared the expression of two markers of apoptosis in the sperm of patients and of fertile donors, and we studied the meiotic segregation in the ejaculated sperm of these translocation carriers. METHODS: Twenty semen samples of translocation carriers, [reciprocal (n=14) and Robertsonian translocations (n=6)], were compared with the semen samples of donors (n=20). Different tests were applied: annexin V binding assay; terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling (TUNEL); and fluorescence in situ hybridization (FISH). RESULTS: The annexin V binding assay in sperm of patients with chromosomal translocation (n=17) showed a significantly increased proportion of sperm with externalized phosphatidylserine (PS) than in the control group (n=20, P相似文献   

Meiotic segregation patterns and ICSI pregnancy outcome of a rare (13;21) Robertsonian translocation carrier: a case report

t(13;21) is an uncommon Robertsonian translocation (RT) with limited information in the literature. Hereby, we assessed the meiotic segregation and interchromosomal effect (ICE) in sperm nuclei from a t(13;21) carrier. The pregnancy outcome following ICSI was also included as reference for physicians and patients. Dual-colour fluorescent in situ hybridization (FISH) was carried out to analyse the segregation pattern of chromosomes 13 and 21, while triple-colour FISH was used to investigate the possible concurrence of ICE. With respect to chromosomal constitutions of 13 and 21, 88.39% of the spermatozoa were normal or balanced due to alternative segregations, and 11.08% showed nullisomy or disomy as a result of adjacent segregations. However, for chromosome 18 and sex chromosomes, the proportion of normal haploids was 98.79%. The rate of disomy was not significantly higher than the controls for either chromosome 18 or X/Y. The rare t(13;21) case exhibited a similar pattern of meiotic segregation as in the common RTs. ICEs were not observed in the current case.     

Meiotic segregation in spermatozoa of a 45,XY,-14,der(18)t(14;18)(q11;p11.3) translocation carrier: a case report

A 35-year-old male was found to have a 45,XY,-14,der(18)t(14;18)(q11;p11.3) karyotype during the investigations for a couple with infertility for 8 years. Two sperm samples were obtained and analysed in triple fluorescence in situ hybridization (FISH) with the D18Z1 and LSI IGH/BCL2 probes. The frequency of gametes exhibiting a normal or balanced chromosomal equipment was 87.26 and 90.97% in samples 1 and 2, respectively. No statistically significant difference was found between the results of meiotic segregation of both samples. These proportions are close to those observed among Robertsonian translocation carriers. They can probably be explained by the formation of trivalent in cis configuration during meiosis I between the derivative chromosome and the normal chromosomes 14 and 18, as in Robertsonian translocation carriers. These results suggest that the configuration adopted at pachytene strongly determines the segregation mode that will be preferentially followed during anaphase I.     

Meiotic segregation of rare Robertsonian translocations: sperm analysis of three t(14q;22q) cases

BACKGROUND: The t(14;22) remains one of the rare Robertsonian translocations observed in human, with an occurrence estimated at 1.2%. Three cases of rare Robertsonian translocation t(14;22) were investigated for meiotic segregation in sperm samples from male carriers using the fluorescent in situ hybridization (FISH) procedure. The three carriers included two men with an abnormal semen analysis and one with normal semen parameters. METHODS: Both locus-specific probes and whole-chromosome painting probes, specific for chromosomes 14 and 22, were used in this study. The number of spermatozoa scored for each probe set ranged from 3279 to 10,024. RESULTS: In the three carriers, similar frequencies, ranging from 78.53 to 81.76%, were found for normal and balanced spermatozoa resulting from alternate segregation. The total proportion of unbalanced spermatozoa resulting from adjacent modes of segregation ranged from 17.59 to 20.94%. CONCLUSION: This finding confirmed the predominance of alternate segregation over other segregation types in all Robertsonian translocations and indicates a higher production of imbalances in the t(14;22) than in most of the Robertsonian translocations previously analysed. This could be related to the variable location of breakpoints in Robertsonian translocations. This breakpoint diversity could also play a role in the differences in reproductive status observed in male carriers of Robertsonian translocations.     

Analysis using fish of sperm and embryos from two carriers of rare rob(13;21) and rob(15;22) robertsonian translocation undergoing PGD

The majority of fluorescence in situ hybridization (FISH) studies on the meiotic segregation of Robertsonian translocations focus on the most common types, rob(13; 14) and rob(14; 21). Here we report the first study for carriers of rare Robertsonian translocations rob(13; 21) and rob(15; 22) combining analysis of meiotic segregation in sperm and blastomeres following pre-implantation genetic diagnosis (PGD). Dual-colour FISH was applied to nuclei from spermatozoa and blastomeres from PGD embryos using two subterminal contig probes for each translocation, and a second round with probes for chromosomes 16 and 18. Patient 1 had a rob(13; 21) and patient 2 had a rob(15; 22), and 86.3% and 87.5% of gametes respectively were consistent with meiotic segregation resulting in a normal or balanced chromosome complement. Analysis of embryos showed that for patient 1 and 2 respectively, 25% and 46% were balanced, and of the unbalanced embryos, 50% and 31% were mosaic or chaotic. Our patients with a rob(13; 21) and rob(15; 22) were found to have a similar meiotic segregation pattern to that for male carriers of the common Robertsonian translocations. The observed rate in unbalanced embryos being mosaic or chaotic may result in an increased risk of chromosomal abnormalities. Our results may help to improve the genetic counseling for carriers of rare Robertsonian translocations.     

Chromosomal segregation in spermatozoa of 14 Robertsonian translocation carriers

Male carriers of Robertsonian (Rob) translocations can havefertility problems associated with low sperm counts and abnormalsperm morphology. In this study, spermatozoa from 14 Rob translocationcarriers, seven der(13;14), two der(13;15), two der(14;15),two der(14;21) and one der(21;22), were tested by fluorescencein-situ hybridization (FISH) for the chromosomes involved, tostudy meiotic segregation behaviour. It was shown that in eachtype of Rob translocation, meiotic segregation behaviour issimilar, comparable and occurs non-randomly. Most of the spermatozoaresults from alternate segregation (range: 76–89.47%).There is, however, still much unbalanced spermatozoa resultingfrom adjacent segregation mode (range: 10.24–23.41%).These data provide useful information for genetic counsellingpurposes. Moreover, aneuploidy for chromosomes 13,18, 21, Xand Y was studied in five patients and suggested an inter-chromosomaleffect.     

Sperm FISH studies in seven male carriers of Robertsonian translocation t(13;14)(q10;q10)

BACKGROUND: Robertsonian translocation t(13;14) is one of the most common structural reorganization in humans, but meiotic segregation studies in these carriers are still limited. The segregation pattern of the chromosomes involved, the possible influence of the translocated chromosomes on the synapsis and disjunction of other chromosome pairs [interchromosomal effects (ICE)] and the rates of unbalanced spermatozoa produced still deserve attention, not only to obtain a better characterization of the meiotic behaviour of this reorganization, but also to offer carrier couples accurate genetic counselling. METHODS: Multicolour fluorescence in-situ hybridization was used to analyse the segregation of chromosomes 13 and 14 and the possible occurrence of ICE (on chromosomes 18, 21, 22, X and Y) in seven male carriers of a t(13;14)(q10;q10). RESULTS AND CONCLUSIONS: The individuals analysed showed a homogeneous segregation pattern, with a clear predominance of alternate segregations resulting in the production of normal/balanced spermatozoa (83-88.23%). A significant increase in the disomy rates for the sex chromosomes, which could be considered as a positive ICE, was observed in two of the carriers analysed.     

Rare Robertsonian translocations and meiotic behaviour: sperm FISH analysis of t(13;15) and t(14;15) translocations: a case report

t(13;15) and t(14;15) are two rare Robertsonian translocations. Meiotic segregation was studied in four males heterozygous for the rare Robertsonian translocations t(13;15) and t(14;15). Both locus-specific probes (LSPs) and whole chromosome painting (WCP) probes, specific to chromosomes 13, 14 and 15, were used in this study. The number of spermatozoa scored for each carrier ranged from 891 to 5000. The frequencies of normal and balanced sperm resulting from the alternate mode of segregation ranged from 77.6 to 92.8%, confirming the prevalence of alternate segregation over other segregation modes in all Robertsonian translocations. The incidences of unbalanced complements ranged from 6.7 to 20.4%, with a significant excess of disomy rates over the complementary frequencies of nullisomy. This variability might reflect differences in the location of breakpoints in translocated chromosomes, leading to the variable production of unbalanced gametes and the variable alterations of semen parameters in Robertsonian translocation carriers.     

运用全染色体探针FISH技术进行罗伯逊易位携带者胚胎植入前诊断

目的运用全染色体探针荧光原位杂交技术(fluorescent in situ hybridization ,FISH)对14/21罗伯逊易位携带者的胚胎进行植入前遗传学诊断,达到优生目的.方法应用FITC、Texas标记的21/14全染色体探针对14/21罗伯逊易位携带者的胚胎活检后的单个卵裂球进行遗传学检测,选择正常信号或携带者信号核型胚胎进行移植.结果常规单精子显微注射-胚胎移植(intracytoplasmic sperm injection,ICSI)后获取胚胎12个,活检优质胚胎11个,杂交后有信号胚胎7个,其中1个正常胚胎,1个携带者胚胎;1个14单体胚胎,1个21单体胚胎,1个21三体胚胎,另2个胚胎活检后均为21号二体,无14号信号.结论运用14/21全染色体探针荧光原位杂交技术对14/21罗伯逊易位携带者的胚胎进行植入前遗传学诊断是能够将正常核型胚胎挑选出来进行移植,从而达到优生目的.     

Correlation between fluorescence in-situ hybridization analyses and in-vitro development to blastocyst stage of embryos from Robertsonian translocation (13;14) carriers

BACKGROUND: Little is known about the extent and timing of selection against the embryos that are carriers of unbalanced translocations. METHODS: Fluorescence in-situ hybridization (FISH) with probes for chromosomes 13, 14 and 18 was performed, mostly on day 3, on 69 human embryos which were then allowed to develop further in culture to day 5, from five carriers of Robertsonian translocation (RT) t(13;14). RESULTS: Twelve normal/balanced blastocysts were replaced in seven consecutive cycles (day 5). Three cycles resulted in clinical pregnancies. The proportion of blastocysts displaying a normal/balanced karyotype was 56%, while only the 20% of blocked embryos were normal/balanced (chi(2): P < 0.05). All the embryos analysed on day 5, except one, displayed mosaicism. The percentages of diploid cells for chromosomes 13 and 14 were significantly lower than for chromosome 18 (chromosome 13: 49.0 +/- 28.0; chromosome 14: 53.0 +/- 31.8; chromosome 18: 75.7 +/- 20.4; Mann-Whitney test: P < 0.01). The embryos displaying vertical line 62% of diploid cells for at least two of the three chromosomes analysed, more frequently reached the blastocyst stage (blocked embryos: blastocysts chromosome 13: 43.1 +/- 30.3, 64.9 +/- 29.0; chromosome 18: 64.9 +/- 29.0, 83.0 +/- 12.9; Mann-Whitney test: P < 0.01). CONCLUSIONS: Normal/balanced embryos developed better but the proportion of abnormal blastocysts was still high. Preimplantation genetic diagnosis is recommended to select normal/balanced embryos from RT t(13;14) carriers.     

A balanced complex chromosomal rearrangement (BCCR) in a family with reproductive failure

Balanced complex chromosomal rearrangements are very rare events in the human population. Translocations involving three or more chromosomes frequently lead to a severe reproductive impairment secondary to meiotic disturbance in males and to chromosomal imbalance in gametes of females. We report a new familial case of complex chromosome anomaly involving chromosomes 13, 14 and 22. Cytogenetic investigations showed a complex chromosomal chromosome rearrangement involving: (i) a Robertsonian translocation between chromosomes 13 and 14; and (ii) a reciprocal translocation between the long arms of chromosome 14 and the long arm of chromosome 22. The aetiology of the translocation was characterized by conventional fluorescence in-situ hybridization (FISH) studies and routine R- and G-banding (RTBG and GBTG) combined with alpha and beta satellite centromeric FISH probes. Predicted configuration of the hexavalent at pachytene stage of meiosis was used to consider the modes of segregation; only two configurations resulted in a normal or balanced gamete karyotype. Reproductive management and genetic counselling are discussed.     

Two cases of Robertsonian translocations in oligozoospermic males and their consequences for pregnancies induced by intracytoplasmic sperm injection

Two case histories are presented documenting structural chromosome abnormalities in infertile males. The abnormalities were detected only after application of intracytoplasmic sperm injection (ICSI) was repeatedly unsuccessful or resulted in an abnormal pregnancy. A mosaic Robertsonian translocation 45,XY,der(13;13)(q10; q10)/46,XY,t(13;13)(p10;p10), der(13p;13p) incompatible with normal offspring was found in a male with extreme oligozoospermia after three subsequent ICSI treatments were unsuccessful and one had resulted in a spontaneous abortion. A second case involved a Robertsonian translocation 45,XY,der(13;14)(q10;q10) which was detected in a male with extreme oligozoospermia after ultrasound abnormalities were found in an ICSI-induced twin pregnancy. Amniocentesis showed an unbalanced 46,XY,+13,der(13;14)(q10;q10) karyotype in one twin and a Robertsonian 45,XX,der(13;14)(q10;q10) karyotype in the other twin. Chromosome analysis of males with abnormal sperm characteristics is advised prior to ICSI.      

10, 15 reciprocal translocation in an infertile man: ultrastructural and fluorescence in-situ hybridization sperm study: case report

BACKGROUND: Peculiar sperm defects are described in a sterile man heterozygous for a balanced translocation t(10;15) (q26;q12). As this structural reorganization was absent in the parents, the translocation must have appeared de novo in the present patient. METHODS: Spermatozoa were analysed under light and transmission electron microscopy (TEM). Fluorescence in-situ hybridization (FISH) was performed on the lymphocyte karyotype. Aneuploidy frequencies of chromosomes 18, X and Y in sperm nuclei, not involved in the translocation, were investigated using three-colour FISH. Dual- colour FISH was used to evaluate segregation of chromosomes 10, 15 in decondensed sperm nuclei. Moreover, three-colour FISH, using telomeric probes for chromosomes 10, 15 was performed in order to distinguish balanced and unbalanced gametes. RESULTS AND CONCLUSIONS: Overall, structural characteristics indicate general immaturity of the germinal cells. FISH sperm analysis detected an increase in chromosome 18 disomy (0.81%) suggesting an interchromosomal effect. A high frequency of diploidies, particularly 18,18,X,X and 18,18,X,Y, was also found. FISH segregation analysis for chromosomes 10, 15 indicated that 32.8% were balanced gametes, whereas 68.2% were unbalanced. Taken together, these data demonstrate in a male carrier of a reciprocal translocation t(10;15) the presence of diffuse ultrastructural sperm alterations and a high frequency of sperm aneuploidies. The existence of a correlation among these factors is proposed.     

精子荧光原位杂交分析男性染色体相互易位携带者的减数分裂分离

目的探讨用精子荧光原位杂交（fluorescence in situ hybridization，HSH）分析男性染色体相互易位携带者减数分裂的分离。方法对4例男性染色体相互易位携带者的精子通过化学方法解聚，在2条易位染色体的断裂点两侧的4个染色体区域中分别选用位于其中3个区域的3个位点探针，进行多色FISH，分析精子染色体组成并推断其分离类型。结果4例染色体相互易位携带者的核型分别为46，XY，t（2；18）（p16；q23）、46，XY，t（4；6）（q34；q21）、46，XY，t（8；13）（q23；q21）和46，XY，t（4；5）（4q31；5q13），其分离结果是：对位分离精子占27．1％～49．4％，邻位-1分离精子占26．9％～37．6％，邻位-2精子占2．7％～15．7％，3：1分离精子占8．6％～33．7％，减数分裂Ⅱ不分离精子占0．2％～1．9％，4：0或二倍体精子占0．1％～0．4％。结论不同的男性染色体相互易位携带者减数分裂的分离结果可能不同，对其行精子FISH分析有助于提供更准确的遗传咨询和行胚胎植入前遗传学诊断的预后估计。     

Translocation t(13;14) in nine generations with a case of translocation homozygosity

The Robertsonian translocation 5(13;14)(p11;q11) was studied in three families with probable common ancestry in Eastern Finland. In the largest family the translocation has segregated through at least nine generations. The same family also included a female who was homozygous for t(13;14). No clear-cut effect of the translocation on fertility could be demonstrated and only one case of trisomy 13 was recorded in the offspring of t(13;14) carriers. The results are discussed, with implication for human chromosomal evolution.     

Successful PGD cycles for mosaic Robertsonian translocation carriers provide insights into the mechanism of formation of the derivative chromosomes

Preimplantation genetic diagnosis (PGD) has been carried out for two couples with different mosaic Robertsonian translocations. Two PGD cycles for a mosaic 13;13 homologous Robertsonian translocation carrier resulted in the birth of a healthy child in each cycle, illustrating the importance of scanning G‐banded preparations from homologous Robertsonian carriers for the presence of a normal cell line. One couple was referred for PGD because the male partner carried a mosaic 14;15 Robertsonian translocation with a normal cell line. A single PGD cycle resulted in the birth of a healthy child. Follow‐up studies and extended FISH analysis of the carrier's lymphocytes detected three cell lines, two carrying different 14;15 Robertsonian chromosomes and one normal cell line. The two 14;15 Robertsonian chromosomes had different breakpoints in the proximal short arm regions. We suggest that the presence of the D15Z1 polymorphism on the short arm of one chromosome 14 mediated the post‐zygotic formation of the two different Robertsonian chromosomes. © 2013 Wiley Periodicals, Inc.     

Molecular cytogenetic studies of Xq critical regions in premature ovarian failure patients

BACKGROUND: Premature ovarian failure (POF) is defined as amenorrhoea for more than 6 months, occurring before the age of 40, with an FSH serum level higher than 40 mIU/ml. Cytogenetically visible rearrangements of the X chromosome are associated with POF. Our hypothesis was that cryptic Xq chromosomal rearrangements could be an important etiological contributor of POF. METHODS: Ninety POF women were recruited and compared to 20 control women. Peripheral blood samples were collected and metaphase chromosomes were prepared using standard cytogenetic methods. To detect Xq chromosomal micro-rearrangements, fluorescence in situ hybridization (FISH) analysis was performed using a selection of 30 bacterial artificial chromosome (BAC) and P1 artificial chromosome clones, spanning Xq13-q27. We further localized the translocation breakpoints by FISH with additional BAC clones. RESULTS: Chromosomal abnormalities were identified in 8.8% of our 90 patients [one triple X, three large Xq deletions 46,X,del(X)(q22.3), 46,X,del(X)(q21.2) and 46,X,del(X)(q21.32), two balanced X;autosome translocations 46,X,t(X;1) (q21.1;q32) and 46,X,t(X;9)(q21.31;q21.2) and two Robertsonian translocations 45,XX,der(15;22)(q10;q10) and 45,XX,der(14;21)(q10;q10)]. The two Xq translocation breakpoints were among a cluster of repetitive elements without any known genes. FISH analysis did not reveal any Xq chromosomal micro-rearrangement. CONCLUSIONS: Karyotyping is definitely helpful in the evaluation of POF patients. No submicroscopic chromosomal rearrangements affecting Xq region were identified. Further analysis using DNA microarrays should help delineate Xq regions involved in POF.     

Chromosome analysis of human spermatozoa from an oligoasthenozoospermic carrier for a 13;14 Robertsonian translocation by their injection into mouse oocytes

We present a case of a 46,XY der(13;14) Robertsonian translocation carrier whose spermatozoa were karyotyped after injection into mouse oocytes. Fresh semen samples as well as recovered samples were used. There was no significant difference in the survival rate of mouse oocytes (fresh: 78.1% versus frozen: 81.7%), activation rate (fresh: 84.0% versus frozen: 90.6%), fertilization rate (fresh: 72.0% versus thawing of frozen: 76.5%) between fresh or frozen spermatozoa. Metaphase chromosome spreads from 45 spermatozoa were analysed. The frequency of spermatozoa that were chromosomally unbalanced with respect to the translocation was 8.9%, and the frequency of abnormalities unrelated to translocation was 4.4%. An excess of spermatozoa with balanced chromosomes was observed: compared with normal, 23 (51.1%) versus 16 (35.6%) respectively; but this segregation difference was not statistically significant (chi(2) = 0.9, P > 0.3). After genetic counselling with the carrier and his partner, intracytoplasmic sperm injection treatment was performed. Healthy female and male infants were delivered at 36 weeks gestation via a Caesarean section. Both babies were carriers for the balanced Robertsonian translocations detected for prenatal diagnosis at 16 weeks gestation. The present study demonstrates that patients can be given further information about the proportion of the spermatozoa which carry a chromosomal abnormality.