Acrocentric chromosome disomy is increased in spermatozoa from fathers of Turner syndrome patients

The aim of the present study was to investigate whether there was an increase of aneuploidy in the sperm from fathers of Turner syndrome patients of paternal origin who, in a previous study, showed an elevated incidence of XY meiotic nondisjunction. Sperm disomy frequencies for chromosomes 4, 13, 18, 21 and 22 were assessed by fluorescence in situ hybridisation in four of these individuals. As a group, the Turner syndrome fathers showed a general increase in disomy frequencies for chromosomes 13, 21 and 22, with a statistically significant increase in disomy frequencies for chromosomes 13 and 22 in one of the fathers and for chromosome 21 in two of them. Data from a previous work carried out by us in two fathers of Down syndrome patients of paternal origin also revealed increased sperm disomy frequencies for chromosomes 13, 21 and 22. Pooled as one group, these six fathers of aneuploid offspring of paternal origin had a statistically significant increase in the frequency of nondisjunction for these chromosomes with respect to control individuals. Our findings indicate that there may be an association between fathering aneuploid offspring and increased frequencies of aneuploid spermatozoa. Such increases do not seem to be restricted to the chromosome pair responsible for the aneuploid offspring. Acrocentric chromosomes and other chromosome pairs that usually show only one chiasma during meiosis seem to be more susceptible to malsegregation.     

Chromosome 21 disomy in the spermatozoa of the fathers of children with trisomy 21, in a population with a high prevalence of Down syndrome: increased incidence in cases of paternal origin.

Between April 1991 and December 1994, epidemiological studies detected a population with a high prevalence of Down syndrome in El Vallès, Spain. Parallel double studies were carried out to determine the parental and the meiotic origins of the trisomy 21, by use of DNA polymorphisms, and to establish the incidence of disomy 21 in the spermatozoa of the fathers of affected children, by use of multicolor FISH. Results show that the overall incidence of chromosome 21 disomy in the fathers of affected children was not significantly different from that in the control population (0.31% vs. 0.37%). However, analysis of individual data demonstrates that two cases (DP-4 and DP-5) with significant increases of disomy 21 (0. 75% and 0.78% vs. 0.37%) correspond to the fathers of the two individuals with Down syndrome of paternal origin. DP-5 also had a significant increase of sex-chromosome disomies (0.69% vs. 0.37%) and of diploid spermatozoa (1.13% vs. 0.24%).     

Nondisjunction of human acrocentric chromosomes: studies of 432 trisomic fetuses and liveborns

The present report summarizes molecular studies on the parent and meiotic stage of origin of the additional chromosome in 432 fetuses or liveborns with an additional chromosome 13, 14, 15, 21, or 22. Our studies suggest that there is little variation in the origin of nondisjunction among the five acrocentric trisomies and that there is no association between the origin of nondisjunction and the likelihood of survival to term of the trisomic conceptus. The proportion of cases of paternal origin was similar among the five trisomies: 12% for trisomy 13, 17% for trisomy 14, 12% for trisomy 15, 9% for trisomy 21, and 11% for trisomy 22. The stage of nondisjunction was also similar among the five trisomies, with the majority of cases of maternal origin being due to nondisjunction at meiosis I, whereas for paternally derived cases, nondisjuction occurred primarily at meiosis II.     

Parental origin of the extra chromosome in prenatally diagnosed fetal trisomy 21

Trisomy 21 (Down syndrome) is one of the most common chromosomal abnormalities. Of cases of free trisomy 21 causing Down syndrome, about 95% result from nondisjunction during meiosis, and about 5% are due to mitotic errors in somatic cells. Previous studies using DNA polymorphisms of chromosome 21 showed that paternal origin of trisomy 21 occurred in only 6.7% of cases. However, these studies were conducted in liveborn trisomy 21-affected infants, and the possible impact of fetal death was not taken into account. Using nine distinct DNA polymorphisms, we tested 110 families with a prenatally diagnosed trisomy 21 fetus. Of the 102 informative cases, parental origin was maternal in 91 cases (89.2%) and paternal in 11 (10.8%). This percentage differs significantly from the 7.0% observed in previous studies (P<0.001). In order to test the influence of genomic parental imprinting, we determined the origin of the extra chromosome 21 in relation to different factors: advanced maternal age, maternal serum human chorionic gonadotropin (hormone of placental origin), severity of the disease, gestational age at diagnosis and fetal gender. We found that the increased frequency of paternal origin of nondisjunction in trisomy 21-affected fetuses cannot obviously be explained by factors leading to selective loss of paternal origin fetuses.     

Molecular studies of trisomy 18.

We have determined the parental origin of 50 cases of trisomy 18. In 48 cases the additional chromosome was maternal in origin, and in 2 cases it was paternal in origin. Seven cases, including both those with an additional paternal chromosome, appeared to be the result of postzygotic error. In contrast to the situation in nondisjunction involving chromosomes 21 and X, there was no evidence for nullochiasmate nondisjunction.     

Trisomy 15 with loss of the paternal 15 as a cause of Prader-Willi syndrome due to maternal disomy.

Uniparental disomy has recently been recognized to cause human disorders, including Prader-Willi syndrome (PWS). We describe a particularly instructive case which raises important issues concerning the mechanisms producing uniparental disomy and whose evaluation provides evidence that trisomy may precede uniparental disomy in a fetus. Chorionic villus sampling performed for advanced maternal age revealed trisomy 15 in all direct and cultured cells, though the fetus appeared normal. Chromosome analysis of amniocytes obtained at 15 wk was normal in over 100 cells studied. The child was hypotonic at birth, and high-resolution banding failed to reveal the deletion of 15q11-13, a deletion which is found in 50%-70% of patients with PWS. Over time, typical features of PWS developed. Molecular genetic analysis using probes for chromosome 15 revealed maternal disomy. Maternal nondisjunction with fertilization of a disomic egg by a normal sperm, followed by loss of the paternal 15, is a likely cause of confined placental mosaicism and uniparental disomy in this case of PWS, and advanced maternal age may be a predisposing factor.     

Trisomy 21: Association between reduced recombination and nondisjunction

To assess the association between recombination and nondisjunction of chromosome 21, we analyzed cytogenetic and DNA markers in 104 trisomy 21 individuals and their parents. Our DNA marker studies of parental origin were informative in 100 cases, with the overwhelming majority (94) being maternal in origin. This value is significantly higher than the 75%-80% maternal nondisjunction rate typically observed in cytogenetic studies of trisomy 21 and illustrates the increased accuracy of the molecular approach. Using the maternally derived cases and probing at 19 polymorphic sites on chromosome 21, we created a genetic map that spans most of the long arm of chromosome 21. The map was significantly shorter than the normal female linkage map, indicating that absence of pairing and/or recombination contributes to nondisjunction in a substantial proportion of cases of trisomy 21.     

Aneuploidy in human spermatozoa

We reviewed the frequency and distribution of disomy in spermatozoa obtained by multicolor-FISH analysis on decondensed sperm nuclei in (a) healthy men, (b) fathers of aneuploid offspring of paternal origin and (c) individuals with Klinefelter syndrome and XYY males. In series of healthy men, disomy per autosome is approximately 0.1% but may range from 0.03 (chromosome 8) to 0.47 (chromosome 22). The great majority of authors find that chromosome 21 (0.18%) and the sex chromosomes (0.27%) have significantly elevated frequencies of disomy although these findings are not universal. The total disomy in FISH studies is 2.26% and the estimated aneuploidy (2× disomy) is 4.5%, more than double that seen in sperm karyotypes (1.8%). Increased disomy levels of low orders of magnitude have been reported in spermatozoa of some normal men (stable variants) and in men who have fathered children with Down, Turner and Klinefelter syndromes. These findings suggest that men with a moderately elevated aneuploidy rate may be at a higher risk of fathering paternally derived aneuploid pregnancies. Among lifestyle factors, smoking, alcohol and caffeine have been studied extensively but the compounding effects of the 3 are difficult to separate because they are common lifestyle behaviors. Increases in sex chromosome abnormalities, some autosomal disomies, and in the number of diploid spermatozoa are general features in 47,XXY and 47,XYY males. Aneuploidy of the sex chromosomes is more frequent than aneuploidy of any of the autosomes not only in normal control individuals, but also in patients with sex chromosome abnormalities and fathers of paternally derived Klinefelter, Turner and Down syndromes.     

Nondisjunction in human sperm: comparison of frequencies in acrocentric chromosomes.

Acrocentric chromosomes may be particularly predisposed to nondisjunction because of the frequency of trisomy for these chromosomes in human spontaneous abortions and liveborns. Studies of aneuploidy in human sperm have provided data on only a few acrocentric chromosomes, with evidence that chromosome 21 has a significantly increased frequency of disomy. To determine whether other acrocentric chromosomes have a higher frequency of nondisjunction or if chromosome 21 is anomalous, disomy frequencies for chromosomes 13 and 22 were studied by fluorescence in situ hybridization (FISH) analysis of 51,043 sperm nuclei from five normal men for whom the frequency of disomy for chromosomes 15 and 21 was known. The mean frequency of disomy for chromosome 13 (0.19%) did not differ significantly from that for other autosomes; however, the frequency of disomy 22 (1.21%) was significantly elevated (P < 0.001, Mantel-Haenszel chi(2) test). The G-group chromosomes (Nos. 21 and 22) also showed a significantly increased frequency of disomy (0. 75%) compared to acrocentric D-group chromosomes (viz., chromosomes 13 and 15; 0.15%) (P < 0.001, Mantel-Haenszel chi(2) test) and other autosomes (chromosomes 1, 2, 4, 9, 12, 13, 15, 16, 18, and 20; 0. 13%) studied in the same men (P < 0.001, Mantel-Haenszel chi(2) test).     

Chromosome spatial order in human cells: evidence for early origin and faithful propagation

We have investigated the origin and nature of chromosome spatial order in human cells by analyzing and comparing chromosome distribution patterns of normal cells with cells showing specific chromosome numerical anomalies known to arise early in development. Results show that all chromosomes in normal diploid cells, triploid cells and in cells exhibiting nondisjunction trisomy 21 are incorporated into a single, radial array (rosette) throughout mitosis. Analysis of cells using fluorescence in situ hybridization, digital imaging and computer-assisted image analysis suggests that chromosomes within rosettes are segregated into tandemly linked “haploid sets” containing 23 chromosomes each. In cells exhibiting nondisjunction trisomy 21, the distribution of chromosome 21 homologs in rosettes was such that two of the three homologs were closely juxtaposed, a pattern consistent with our current understanding of the mechanism of chromosomal nondisjunction. Rosettes of cells derived from triploid individuals contained chromosomes segregated into three, tandemly linked haploid sets in which chromosome spatial order was preserved, but with chromosome positional order in one haploid set inverted with respect to the other two sets. The spatial separation of homologs in triploid cells was chromosome specific, providing evidence that chromosomes occupy preferred positions within the haploid sets. Since both triploidy and nondisjunction trisomy 21 are chromosome numerical anomalies that arise extremely early in development (e.g., during meiosis or during the first few mitoses), our results support the idea that normal and abnormal chromosome distribution patterns in mitotic human cells are established early in development, and are propagated faithfully by mitosis throughout development and into adult life. Furthermore, our observations suggest that segregation of chromosome homologs into two haploid sets in normal diploid cells is a remnant of fertilization and, in normal diploid cells, reflects segregation of maternal and paternal chromosomes. Received: 19 January 1998; in revised form: 28 May 1998 / Accepted: 30 June 1998     

The meiotic stage of nondisjunction in trisomy 21: Determination by using DNA polymorphisms

We have studied DNA polymorphisms at loci in the pericentromeric region on the long arm of chromosome 21 in 200 families with trisomy 21, in order to determine the meiotic origin of nondisjunction. Maintenance of heterozygosity for parental markers in the individual with trisomy 21 was interpreted as resulting from a meiosis I error, while reduction to homozygosity was attributed to a meiosis II error. Nondisjunction was paternal in 9 cases and was maternal in 188 cases, as reported earlier. Among the 188 maternal cases, nondisjunction occurred in meiosis I in 128 cases and in meiosis II in 38 cases; in 22 cases the DNA markers used were uninformative. Therefore meiosis I was responsible for 77.1% and meiosis II for 22.9% of maternal nondisjunction. Among the 9 paternal nondisjunction cases the error occurred in meiosis I in 2 cases (22.2%) and in meiosis II in 7 (77.8%) cases. Since there was no significant difference in the distribution of maternal ages between maternal I error versus maternal II error, it is unlikely that an error at a particular of maternal ages between maternal I error versus maternal II error, it is unlikely that an error at a particular meiotic stage contributes significantly to the increasing incidence of Down syndrome with advancing maternal age. Although the DNA polymorphisms used were at loci which map close to the centromere, it is likely that rare errors in meiotic-origin assignments may have occurred because of a small number of crossovers between the markers and the centromere.(ABSTRACT TRUNCATED AT 250 WORDS)     

Analysis of sex chromosome aneuploidy in sperm from fathers of Turner syndrome patients

Numerical sex chromosome abnormalities were analyzed in sperm from four fathers of Turner syndrome patients of paternal origin to determine whether there was an increased frequency of sex chromosome aneuploidy and to elucidate whether meiotic malsegregation mechanisms could be involved in the origin of Turner syndrome. Determination of the parental origin of the single X chromosome (maternal in all four cases) and exclusion of X and Y mosaicism were carried out by polymerase chain reaction amplification of five X chromosome polymorphisms and three Y chromosome segments. A total of 45,299 sperm nuclei from Turner fathers and 85,423 sperm nuclei from eight control donors was analyzed by three-color fluorescence in situ hybridization. The four patients showed a significant increase in the percentages of XY sperm (mean 0.22%; range 0.20% to 0.22%) compared with control donors (mean 0.11%; range 0.06% to 0.18%). These results suggest that the four individuals have an increased frequency of nondisjunctional errors in meiosis I, resulting in the production of an increased proportion of XY spermatozoa and of sperm lacking a sex chromosome. Received: 24 November 1998 / Accepted: 2 February 1999     

Origin of the extra chromosome in trisomy 21

Summary Of 61 families of children with trisomy 21, polymorphism of chromosome 21 elucidating the origin of the extra chromosome was found in 42. Nondisjunction was of paternal origin in 8 cases (19.04%) and the anomaly occurred with equal frequency during the first and second meiotic divisions. Maternal nondisjunction was demonstrated in 34 cases (80.95%), in which nondisjunction occurred by far the most often during the first meiotic division (29 cases).These results are in agreement with data from the literature, and suggest the existence of at least two different causes for chromosomal nondisjunction, the first being the same in both sexes and occurring in both meiotic divisions and the second specifically limited to the first meiotic division in the mother.Attachée de Recherche au CNRSAttachée de Recherche à l'INSERM     

QF-PCR examination of parental and meiotic origin of trisomy 21 in Central and Eastern Europe.

Study of parental/meiotic origin of free trisomy 21 in nuclear families from Russia (70 cases), Ukraine (32 cases), and 22 from Germany revealed maternal nondisjunction in 77.3% (Germany), 93.8% (Ukraine), and 91.4% (Russia), paternal origin in 13.6%, 6.2%, and 8.6%, respectively. Maternal meiosis I errors were found in 84.4% (Ukraine), 77.1% (Russia), paternal origin in 3.1% (Ukraine), 2.9% (Russia). Maternal meiosis II errors occurred in 9.4% and 14.3% and paternal in 3.1% and 5.7% in Ukraine and Russia, respectively. No significant differences were found in maternal/paternal origin among Ukraine, Russia, Germany, and published data from other European regions.     

Aneuploidy in human sperm: the use of multicolor FISH to test various theories of nondisjunction.

While it is known that all chromosomes are susceptible to meiotic nondisjunction, it is not clear whether all chromosomes display the same frequency of nondisjunction. By use of multicolor FISH and chromosome-specific probes, the frequency of disomy in human sperm was determined for chromosomes 1, 2, 4, 9, 12, 15, 16, 18, 20, and 21, and the sex chromosomes. A minimum of 10,000 sperm nuclei were scored from each of five healthy, chromosomally normal donors for every chromosome studied, giving a total of 418,931 sperm nuclei. The mean frequencies of disomy obtained were 0.09% for chromosome 1; 0.08% for chromosome 2; 0.11% for chromosome 4; 0.14% for chromosome 9; 0.16% for chromosome 12; 0.11% for chromosomes 15, 16, and 18; 0.12% for chromosome 20; 0.29% for chromosome 21; and 0.43% for the sex chromosomes. Data for chromosomes 1, 12, 15, and 18, and the sex chromosomes have been published elsewhere. When the mean frequencies of disomy were compared, the sex chromosomes and chromosome 21 had significantly higher frequencies of disomy than that of any other autosome studied. These results corroborate the pooled data obtained from human sperm karyotypes and suggest that the sex chromosome bivalent and the chromosome 21 bivalent are more susceptible to nondisjunction during spermatogenesis. From these findings, theories proposed to explain the variable incidence of nondisjunction can be supported or discarded as improbable.     

Increased rate of nondisjunction in sex cells derived from low-quality semen

The relationship between chromosomal nondisjunction and semen quality was studied in two groups of males who differ highly in their semen quality: 12 individuals with low-quality semen caused by varicocele, and 8 subjects with high-quality semen, selected from sperm donors for in vitro fertilization. Chromosomal nondisjunction was inferred from the rate of disomy found in mature sperm cells. To determine the rate of disomy, we applied fluorescence in situ hybridization using satellite-specific probes for chromosomes 1, 15, 18, X and Y. In sperm cells of males with low-quality semen, the mean rate of disomy for each of the autosomes and of hetero-disomy for the sex chromosomes (XY) was significantly higher than that observed in the high-quality semen samples: more than 15-fold higher for chromosomes 1 and 15, and 7-fold higher for chromosomes 18 and XY. Yet, the homo-disomy rate for each of the sex chromosomes (XX and YY) was almost the same in both types of semen. The large discrepancy between the low- and high-quality semen in the rate of sex chromosome hetero-disomy versus the similar rate of homo-disomy strongly suggests that the abnormal chromosomal segregation in meiocytes of males with low-quality semen resulted from chromosomal nondisjunction at the first meiotic division. The results indicate that men showing poor semen quality are at an increased risk for meiotic nondisjunction, similar to women at the end of their reproductive years. Received: 30 June 1997 / Accepted: 17 September 1997     

Centromeric genotyping and direct analysis of nondisjunction in humans: Down syndrome

In species with chiasmate meioses, alterations in genetic recombination are an important correlate of nondisjunction. In general, these alterations fall into one of two categories: either homologous chromosomes fail to pair and/or recombine at meiosis I, or they are united by chiasmata that are suboptimally positioned. Recent studies of human nondisjunction suggest that these relationships apply to our species as well. However, methodological limitations in human genetic mapping have made it difficult to determine whether the important determinant(s) in human nondisjunction is absent recombination, altered recombination, or both. In the present report, we describe somatic cell hybrid studies of chromosome 21 nondisjunction aimed at overcoming this limitation. By using hybrids to “capture” individual chromosomes 21 of the proband and parent of origin of trisomy, it is possible to identify complementary recombinant meiotic products, and thereby to uncover crossovers that cannot be detected by conventional mapping methods. In the present report, we summarize studies of 23 cases. Our results indicate that recombination in proximal 21q is infrequent in trisomy-generating meioses and that, in a proportion of the meioses, recombination does not occur anywhere on 21q. Thus, our observations indicate that failure to recombine is responsible for a proportion of trisomy 21 cases. Received: 12 January 1997; in revised form: 16 February 1998 / Accepted: 19 February 1998     

Human sperm chromosome complements in chemotherapy patients and infertile men

Our studies of human sperm karyotypes and interphase sperm analyzed by fluorescence in situ hybridization (FISH) have both yielded estimates of disomy frequencies of approximately 0.1% per chromosome with an overall aneuploidy frequency in human sperm of approximately 5%–6%. However, the distribution of aneuploidy in sperm is not even, as our data from sperm karyotypes and multicolour FISH analyses both demonstrate a significant increase in the frequency of aneuploidy for chromosome 21 and the sex chromosomes. We have studied men at increased risk of sperm chromosomal abnormalities including cancer patients and infertility patients. Testicular cancer patients were studied before and 2–13 years after chemotherapy (CT) with BEP (bleomycin, etoposide, cisplatin). Sperm karyotype analysis on 788 sperm demonstrated no significant difference in the frequency of numerical or structural chromosomal abnormalities post-CT vs pre-CT. Similarly, multicolour FISH analysis for chromosomes 1, 12, XX, YY and XY in 161,097 sperm did not detect any significant differences in the frequencies of disomy before and after treatment. However, recent evidence has suggested a significant increase in the frequency of disomy and diploidy during CT. We have found that infertile men, who would be candidates for intracytoplasmic sperm injection, have an increased frequency of chromosomally abnormal sperm karyotypes. Also, FISH analysis for chromosomes 1, 12, 13, 21, XX, YY and XY in 255,613 sperm demonstrated a significant increase in chromosomes 1, 13, 21, and XY disomy in infertile men compared with control donors. Received: 4 July 1998; in revised form: 7 September 1998 / Accepted: 8 September 1998     

Frequency of XY sperm increases with age in fathers of boys with Klinefelter syndrome

With increasing availability of drugs for impotence and advanced reproductive technologies for the treatment of subfertility, more men are fathering children at advanced ages. We conducted a study of the chromosomal content of sperm of healthy men aged 24-57 years to (a) determine whether father's age was associated with increasing frequencies of aneuploid sperm including XY, disomy X, disomy Y, disomy 21, and sperm diploidy, and (b) examine the association between the frequencies of disomy 21 and sex-chromosomal aneuploidies. The study group consisted of 38 fathers of boys with Klinefelter syndrome (47, XXY) recruited nationwide, and sperm aneuploidy was assessed using multicolor X-Y-21 sperm FISH ( approximately 10,000 sperm per donor). Paternal age was significantly correlated with the sex ratio of sperm (Y/X; P=.006) and with the frequency of XY sperm (P=.02), with a clear trend with age by decades (P<.006). Compared with fathers in their 20s (who had an average frequency of 7.5 XY sperm per 10,000), the frequencies of XY sperm were 10% higher among fathers in their 30s, 31% higher among those in their 40s, and 160% higher among those in their 50s (95% CI 69%-300%). However, there was no evidence for age effects on frequencies of sperm carrying nullisomy sex; disomies X, Y, or 21; or meiosis I or II diploidies. The frequencies of disomy 21 sperm were significantly associated with sex-chromosomal aneuploidy (P=.04)-in particular, with disomy X (P=.004), but disomy 21 sperm did not preferentially carry either sex chromosome. These findings suggest that older fathers produce higher frequencies of XY sperm, which may place them at higher risk of fathering boys with Klinefelter syndrome, and that age effects on sperm aneuploidy are chromosome specific.     

Isolation of chromosome-21-specific DNA probes and their use in the analysis of nondisjunction in Down syndrome

Summary Thirteen single-copy, chromosome-21-specific DNA probes were isolated from a recombinant library made from flow-sorted chromosome 21 DNA and regionally mapped using a panel of somatic cell hybrids. Five probes mapped in the 21q21-q22.1 region, six to the 21q22.1-qter region, and one to each of the regions 21q22.1-q22.2 and 21q22.3. Two of these probes, one of which maps in the critical region for Down syndrome, have recently been shown to be expressed at high levels in Down syndrome brain tissue (Stefani et al. 1988). Following preliminary screening for restriction fragment lenght polymorphisms (RFLPs), five polymorphisms were discovered with four of the chromosome 21 DNA probes. A frequent MspI polymorphism detected by one of the probes was used in conjunction with four previously described polymorphic chromosome 21 probes to analyse the origin of nondisjunction in 33 families with a child or fetus with trisomy 21. The parental origin of the additional chromosome 21 was determined in 12 cases: in 9 (75%) of these it was derived from the mother and in the other 3 cases (25%) it was of paternal origin. Cytogenetic analysis of Q-banding heteromorphisms was informative in three of five families tested, and in each case the RFLP results were confirmed. The meiotic stage of nondisjunction was defined with confidence in five families, the results being obtained with pericentromeric RFLP or cytogenetic markers. Recombination between two nondisjoined chromosomes was demonstrated in one family and is consistent with the view that a lack of recombination between chromosome 21 homologues or failure of their conjunction is not the invariable cause of trisomy 21.