Trisomy 18: studies of the parent and cell division of origin and the effect of aberrant recombination on nondisjunction.

We have studied the mechanism of origin of 63 cases of trisomy 18. In 2 the additional chromosome was paternal in origin, and in the remaining 61 it was maternal in origin. Both paternal cases were attributable to a postzygotic mitotic (PZM) error. Among the 54 maternal cases for which the cell division of error was established, only 16 were attributable to an error at the first meiotic division (mat MI), whereas no fewer than 35 were due to an error at the second meiotic division (mat MII), the remaining 3 being the result of a PZM error involving the maternal chromosome 18. A standard map of chromosome 18 was constructed and compared with the nondisjunctional map. Approximately one-third of the mat MI errors were associated with complete absence of recombination, whereas in the remaining two-thirds and in all the mat MII errors recombination in the nondisjoined chromosomes appeared to be normal. All the maternal errors were associated with an increased maternal age, although this reached significance only for the mat MII category of nondisjunction. Our observations on chromosome 18 are compared with those on other chromosomes for which there are comparable data.     

Molecular studies of the parental origin and nature of human X isochromosomes

X-chromosome restriction fragment length polymorphisms were used to determine the parental origin of the isochromosome in nine individuals with an i(Xq) or idic(Xq). We were able to specify the parental source of eight of the nine isochromosomes, with six being maternal and two paternal in origin. In two cases, one i(Xq) and one idic(Xq), we used Xq markers to determine the level of heterozygosity in the isochromosome. Each was homozygous at all tested loci, suggesting that each originated from a single X chromosome and not from an exchange of material between two X's.     

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.     

Molecular studies of parental origin and mosaicism in 45,X conceptuses

Summary The present report summarizes molecular studies of parental origin and sex chromosome mosaicism in forty-one 45,X conceptuses, consisting of 29 spontaneous abortions and 12 liveborn individuals with Turner syndrome. Our studies indicate that most 45,X conceptuses have a single, maternally derived X chromosome, regardless of whether the conceptus is liveborn or spontaneously aborted. In studies of mosaicism, our identification of X- and Y-chromosome mosaics among 45,X spontaneous abortions indicates that mosaicism does not ensure survival to term of 45,X fetuses. However, the incidence of sex chromosmome mosaicism is substantially higher in liveborn than in aborted 45,X conceptuses, indicating that the presence of a second cell line increases the likelihood of survival to term.     

Genetics and biology of human ovarian teratomas. II. Molecular analysis of origin of nondisjunction and gene-centromere mapping of chromosome I markers.

Chromosomal heteromorphisms and DNA polymorphisms have been utilized to identify the mechanisms that lead to formation of human ovarian teratomas and to construct a gene-centromere map of chromosome 1 by using those teratomas that arise by meiotic nondisjunction. Of 61 genetically informative ovarian teratomas, 21.3% arose by nondisjunction at meiosis I, and 39.3% arose by meiosis II nondisjunction. Eight polymorphic marker loci on chromosome 1p and one marker on 1q were used to estimate a gene-centromere map. The results show clear linkage of the most proximal 1p marker (NRAS) and the most proximal 1q marker (D1S61) to the centromere at a distance of 14 cM and 20 cM, respectively. Estimated gene-centromere distances suggest that, while recombination occurs normally in ovarian teratomas arising by meiosis II errors, ovarian teratomas arising by meiosis I nondisjunction have altered patterns of recombination. Furthermore, the estimated map demonstrates clear evidence of chiasma interference. Our results suggest that ovarian teratomas can provide a rapid method for mapping genes relative to the centromere.     

Cloned DNA probes regionally mapped to human chromosome 21 and their use in determining the origin of nondisjunction.

A number of unique sequence recombinant DNA clones were isolated from a recombinant DNA library constructed from DNA enriched for chromosome 21 by flow sorting. Of these, five were mapped to chromosome 21 using a somatic cell hybrid. Regional mapping of these probes and of a probe previously assigned to chromosome 21, was carried out with the aid of chromosome 21 rearrangements using both chromosome sorting and a somatic cell hybrid. Three probes were shown to be located on either side of the breakpoint 21q21.2. Two of the probes were shown to identify restriction fragment length polymorphisms (RFLPs) with high rare-allele frequencies (0.46 and 0.43). A Bgl II RFLP revealed the parental origin of non-disjunction in three of ten families with Down's syndrome.     

New insights into human nondisjunction of chromosome 21 in oocytes

Nondisjunction of chromosome 21 is the leading cause of Down syndrome. Two risk factors for maternal nondisjunction of chromosome 21 are increased maternal age and altered recombination. In order to provide further insight on mechanisms underlying nondisjunction, we examined the association between these two well established risk factors for chromosome 21 nondisjunction. In our approach, short tandem repeat markers along chromosome 21 were genotyped in DNA collected from individuals with free trisomy 21 and their parents. This information was used to determine the origin of the nondisjunction error and the maternal recombination profile. We analyzed 615 maternal meiosis I and 253 maternal meiosis II cases stratified by maternal age. The examination of meiosis II errors, the first of its type, suggests that the presence of a single exchange within the pericentromeric region of 21q interacts with maternal age-related risk factors. This observation could be explained in two general ways: 1) a pericentromeric exchange initiates or exacerbates the susceptibility to maternal age risk factors or 2) a pericentromeric exchange protects the bivalent against age-related risk factors allowing proper segregation of homologues at meiosis I, but not segregation of sisters at meiosis II. In contrast, analysis of maternal meiosis I errors indicates that a single telomeric exchange imposes the same risk for nondisjunction, irrespective of the age of the oocyte. Our results emphasize the fact that human nondisjunction is a multifactorial trait that must be dissected into its component parts to identify specific associated risk factors.     

Molecular investigation of the parental origin of a de novo unbalanced translocation 13/18

We report a patient with a de novo translocation 13/18, identified by high-resolution banding. The breakpoints were ascertained by fluorescence in situ hybridisation with whole chromosome 13 and 18 paints. Short tandem repeat typing demonstrated the aberration to be of combined maternal/paternal origin and thereby confirmed its de novo and postzygotic formation. Thus, a gonadal mosaic in one of the parents resulting in a higher recurrence risk could be excluded. Received: 1 September 1996 / Revised: 3 November 1996     

Retrospective study of the parental origin of the extra chromosome in trisomy 18 (Edwards syndrome)

The parental origin of the extra chromosome in trisomy 18 was traced in 30 informative families using highly polymorphic (CA) repeats mapped on the long arm of chromosome 18. Proband DNA was recovered from slides of chromosome preparations in 28 cases and from paraffin-embedded tissues in two cases. The extra chromosome was found to be of maternal origin in 26 cases (86.7%), and paternal origin in 4 cases (13.3%).     

Increased nondisjunction of chromosome 21 with age in human peripheral lymphocytes

Fluorescence in situ hybridization (FISH) on binucleated cells with chromosome-specific DNA probes provides a convenient way to visualize reciprocal segregation patterns in daughter nuclei, and overcomes most problems related to the artefactual loss or gain of chromosomes that flaw chromosome preparations. In this study, FISH was employed to evaluate age- and sex-effects on spontaneous malsegregation, nondisjunction and loss of chromosome 21 in human lymphocytes after the first division in culture. A total of 68 healthy nonsmokers and nondrinkers of alcohol (37 males and 31 females) were grouped by age as Group I (0-10 years), Group II (20-30 years), Group III (40-50 years) and Group IV (60-70 years), with at least seven subjects per group and sex. FISH with a pericentric chromosome 21 specific DNA probe was carried out on binucleated lymphocytes, cytokinesis-blocked by cytochalasin B (6 microg/ml for 26 h) at 44 h after initiation of cultures.Linear regression analyses demonstrated a significant age-related increase in the frequency of micronuclei without chromosome 21 (MN-21)(r=0.73, p<0.001 in females; r=0.69, p<0.001 in males) in all binucleated cells, with a steeper slope in females (0.1758) than in males (0. 1241). Analysis using the 2x2 chi-square (chi(2)) test on the frequencies of MN-21 showed significant age-related differences in both males and females, except males in Group III and Group IV (p>0. 05). A significant sex-related difference was found only in subjects over 60 years (p<0.05), with females having more MN-21 (12.57 per thousand vs. 8.43 per thousand) than males.Loss of chromosome 21, occurring at mean levels of 0.38 per thousand in all binucleated cells and 0.24 per thousand in binucleated cells containing four FISH signals, was shown not to be age- or sex-related. A positive age-related increase in nondisjunction of chromosome 21 was shown in males (r=0.50, p<0.01), females (r=0.61, p<0.001) and all subjects (r=0.55, p<0.001) by linear regression analysis. An age effect was found only between children and adults (p<0.01 for females, p<0.05     

Trisomy of chromosome 18 in the baboon (Papio hamadryas anubis)

Trisomy 18 is usually a lethal chromosomal abnormality and is the second most common autosomal trisomy in humans, with an incidence of 1:8000 live births. It is commonly associated with abnormalities of the lower and upper extremities, having the frequency of 95% and 65%, respectively. A newborn female olive baboon (Papio hamadryas anubis) was diagnosed with intrauterine growth retardation and severe arthrogryposis-like congenital joint deformities. Cytogenetic analysis including G-banding and fluorescence in situ hybridization (FISH) revealed that the congenital abnormalities were associated with chromosomal mosaicism for trisomy 18. Genetic analysis with microsatellites from chromosome 18 confirmed the maternal origin of the extra chromosome 18. This is the first report of trisomy 18 in the baboon, which may be a promising animal model of human disease.     

Trisomy 21 (Down syndrome): studying nondisjunction and meiotic recombination by using cytogenetic and molecular polymorphisms that span chromosome 21.

By combining molecular and cytogenetic techniques, we demonstrated the feasibility and desirability of a comprehensive approach to analysis of nondisjunction for chromosome 21. We analyzed the parental origin and stage of meiotic errors resulting in trisomy 21 in each of five families by successfully using cytogenetic heteromorphisms and DNA polymorphisms. The 16 DNA fragments used to detect polymorphisms spanned the length of the long arm and detected recombinational events on nondisjoined chromosomes in both maternal meiosis I and maternal meiosis II errors. The meiotic stage at which errors occurred was determined by sandwiching the centromere between cytogenetic heteromorphisms on 21p and an informative haplotype constructed using two polymorphic DNA probes that map to 21q just below the centromere. This study illustrates the necessity of combining cytogenetic polymorphisms on 21p with DNA polymorphisms spanning 21q to determine (1) the source and stage of meiotic errors that lead to trisomy 21 and (2) whether an association exists between nondisjunction and meiotic recombination.     

Equal parental origin of chromosome 22 losses in human sporadic meningioma: no evidence for genomic imprinting.

Inactivation of tumor suppressor genes can occur either by mutation at the gene locus or by loss of part or all of the chromosome region containing the gene. The latter is most frequently detected by DNA markers as loss of heterozygosity in the tumor tissue. In several reports, the paternal homologue was preferentially retained in embryonal tumors associated with loss of particular chromosomal regions, suggesting genomic imprinting of the corresponding tumor suppressor loci. To explore the generality of these findings and the possible role of genomic imprinting in adult tumors of the nervous system, we have determined the parental origin of chromosome 22 loss in sporadic meningioma. Of nine cases studied, five tumors retained the maternally derived chromosome 22 homologue while four retained the paternally derived chromosome 22. Thus, in contrast to the embryonal tumors, the meningioma locus on chromosome 22 is inactivated by random mutation in sporadic adult meningiomas.     

Trisomy 7 and sex chromosome loss in human brain tissue

Short-term cultures of nonneoplastic brain tissue from 11 patients, seven of whom had a malignant brain tumor, were cytogenetically examined. In only a single case was a wholly normal chromosome complement detected; the remaining ten cases exhibited mosaicism with clonal numerical aberrations found alongside cells carrying a normal karyotype. The abnormal clones were characterized by trisomy 7, the loss of the Y chromosome in men and an X chromosome in women, or by combinations thereof. No structural aberrations were present. Our findings demonstrate that although -Y, -X, and +7 have in the past repeatedly been associated with brain tumors, these changes presumably reflect normal in vivo organ mosaicism and, thus, should not be accepted as neoplasia-specific in this context.     

Somatic cell hybrid deletion map of human chromosome 18.

The creation of a physical map of chromosome 18 will be useful for the eventual identification of specific chromosomal regions that are critical in the occurrence of Edwards syndrome, the 18q- syndrome, and the 18p- syndrome. To begin the investigation of these syndromes, a physical map has been constructed to order random DNA fragments to specific portions of chromosome 18. A set of somatic cell hybrids that retain deletions or translocations involving chromosome 18 has been isolated and characterized. Over 200 lambda phage from a chromosome 18-specific library have been localized to 11 distinct regions of chromosome 18 using the chromosomal breakpoints present in the somatic cell hybrids.     

Nondisjunction of chromosome 21: comparisons of cytogenetic and molecular studies of the meiotic stage and parent of origin.

In the present report, we summarize studies aimed at examining the reliability of chromosome heteromorphisms in analyses of chromosome 21 nondisjunction. We used two cytogenetic approaches--fluorescent in situ hybridization (FISH) to repetitive sequences on 21p and traditional Q-banding--to distinguish chromosome 21 homologues and then compared the results of these studies with those obtained by DNA markers. Using a conservative scoring system for Q-banding and FISH heteromorphisms, we were able to specify the parental origin of trisomy in 10% of cases; in contrast, DNA marker studies were informative for parental origin in almost all cases. The results of the molecular and cytogenetic studies of parental origin concurred in all cases in which assignments were made independently using both techniques. However, in 4 of 13 cases in which the molecular studies contributed to the interpretation of the cytogenetic findings, the two results did not agree with respect to the meiotic stage of nondisjunction. A relatively high frequency of crossing-over on either the short arm or proximal long arm of chromosome 21 could explain these results and may be a mechanism leading to nondisjunction.     

PCR protocol to detect parental origin and hidden mosaicism inSex chromosome aneuploidies

In this study, we report an accurate method to determine the parental origin of sex chromosome aneuploidies or polyploidies and to detect low percentage mosaicisms. We have amplified by polymerase chain reaction (PCR) five polymorphic markers along the X chromosome (DXS1283E, DYS II, DMD49, AR and DXS52) and three markers along the Y chromosome (SRY, DYZ3 and DYZ1). False-negative results were discarded by the simultaneous amplification of Y markers and of internal controls. We have applied this protocol to a series of 14 Turner syndrome patients with a 45,X karyotype. We have detected sex chromosome mosaicisms in two patients. The parental origin of the syndrome has been determined in the other 12 patients.