Heteromorphic variants of chromosome 9.

BACKGROUND: Heterochromatic variants of pericentromere of chromosome 9 are reported and discussed since decades concerning their detailed structure and clinical meaning. However, detailed studies are scarce. Thus, here we provide the largest ever done molecular cytogenetic research based on >300 chromosome 9 heteromorphism carriers. RESULTS: In this study, 334 carriers of heterochromatic variants of chromosome 9 were included, being 192 patients from Western Europe and the remainder from Easter-European origin. A 3-color-fluorescence in situ hybridization (FISH) probe-set directed against for 9p12 to 9q13~21.1 (9het-mix) and 8 different locus-specific probes were applied for their characterization. The 9het-mix enables the characterization of 21 of the yet known 24 chromosome 9 heteromorphic patterns. In this study, 17 different variants were detected including five yet unreported; the most frequent were pericentric inversions (49.4%) followed by 9qh-variants (23.9%), variants of 9ph (11.4%), cenh (8.2%), and dicentric- (3.8%) and duplication-variants (3.3%). For reasons of simplicity, a new short nomenclature for the yet reported 24 heteromorphic patterns of chromosome 9 is suggested. Six breakpoints involved in four of the 24 variants could be narrowed down using locus-specific probes. CONCLUSIONS: Based on this largest study ever done in carriers of chromosome 9 heteromorphisms, three of the 24 detailed variants were more frequently observed in Western than in Eastern Europe. Besides, there is no clear evidence that infertility is linked to any of the 24 chromosome 9 heteromorphic variants.

A new multicolor fluorescence in situ hybridization probe set directed against human heterochromatin: HCM-FISH.

A new multicolor fluorescence in situ hybridization (mFISH) probe set is presented, and its possible applications are highlighted in 25 clinical cases. The so-called heterochromatin-M-FISH (HCM-FISH) probe set enables a one-step characterization of the large heterochromatic regions within the human genome. HCM-FISH closes a gap in the now available mFISH probe sets, as those do not normally cover the acrocentric short arms; the large pericentric regions of chromosomes 1, 9, and 16; as well as the band Yq12. Still, these regions can be involved in different kinds of chromosomal rearrangements such as translocations, insertions, inversions, amplifications, and marker chromosome formations. Here, examples are given for all these kinds of chromosomal aberrations, detected as constitutional rearrangements in clinical cases. Application perspectives of the probe set in tumors as well as in evolutionary cytogenetic studies are given.

Biomarkers for genome instability in some genetic disorders: a pilot study.

CONTEXT: The study of genome integrity in some genetic disorders has diagnostic and prognostic importance because of the evident relationship between genome instability and both DNA repair deficiencies and cancer predisposition. OBJECTIVE: The objective was to compare the chromosomal and DNA damage responses in lymphocytes from patients with Nijmegen breakage syndrome (NBS), Fanconi anemia (FA) and Williams-Beuren syndrome (WBS) to find additional biomarkers of genome instability. METHODS: The cytogenetic approaches were combined with the alkaline Comet assay to estimate genome integrity in cultured or freshly isolated and H(2)O(2)-treated lymphocytes. RESULTS: Basal frequencies of chromosome aberrations were significantly increased in NBS/FA probands and NBS heterozygous carriers. The NBS diagnosis was confirmed by detecting site-specific rearrangements, while the mitomycin C (MMC)-stress test was highly positive in a FA patient. Among patients with suspected WBS, 12 individuals had a 7q11.23 microdeletion. In the Comet assay, genome instability was revealed in all three disorders, impaired capacity to repair oxidative damage being observed in NBS and WBS in contrast to FA and controls. CONCLUSION: The results indicate that the estimates of DNA damage response may be proposed as efficient biomarkers for detecting and characterizing genome instability in the genetic disorders under study.

Chromosomal instability at the 7q11.23 region impacts on DNA-damage response in lymphocytes from Williams-Beuren syndrome patients.

Williams-Beuren syndrome (WBS) is the chromosomal disorder arising from a hemizygous microdeletion at 7q11.23. The present study was focused on a comparative investigation of genomic integrity in WBS patients by use of cytogenetic methods and the alkaline comet assay. Lymphocytes of whole peripheral blood were cultured and metaphases were examined for frequency and spectrum of chromosome aberrations. A WBS-related microdeletion was detected by means of the FISH (fluorescence in situ hybridization) technique. The blood samples from patients who were carriers of this microdeletion, were tested in the comet assay. For this purpose, freshly collected lymphocytes were exposed to hydrogen peroxide (100µM, 1min, 4°C). The frequencies of endogenous and exogenous DNA damage, and the kinetics and efficiency of DNA repair were measured during three subsequent hours of incubation. Comparison of the two data sets in this group of patients demonstrated a slightly elevated average frequency of chromosome aberrations, significantly increased levels of endogenous and H(2)O(2)-induced DNA damage, and somewhat impaired DNA repair. The relationship between an abnormal DNA-damage response and the 7q11.23 hemizygous microdeletion was confirmed experimentally when comparing the comet assay data in FISH-positive and FISH-negative lymphocytes from WBS-suspected patients. Briefly, our results indicate the impact of chromosomal instability within this region on susceptibility towards DNA damage, which may contribute to pathogenesis of this disease. It was shown also that the comet assay, as well as an experimental design proposed here, seem to be useful tools for estimating genome integrity in WBS patients.

Paternally derived der(7)t(Y;7)(p11.1 approximately 11.2;p22.3)dn in a mosaic case with Turner syndrome.

An unusual mosaic karyotype was detected in a 6-year-old female patient with clinical diagnosis of Turner syndrome (TS). Cytogenetic and molecular cytogenetic studies revealed besides a cell line with 45,X a second cell line where the short arm of the Y-chromosome was translocated onto the short arm of a chromosome 7; karyotype: 45,X,der(7)t(Y;7)(p11.1 approximately 11.2;p22.3)/45,X. To delineate the mechanisms of rearrangement and karyotypic evolution in this case, further studies were performed. A maternal origin of the X-chromosome and biparental origin of both chromosomes 7 were determined by microsatellite analysis. Furthermore, using parental-origin-determination fluorescence in situ hybridization (pod-FISH) it could be established that the derivative chromosome 7 was of paternal origin. Overall, this is to the best of our knowledge the first report of such a complex mosaic TS karyotype.

Pallister-Killian syndrome: rapid decrease of isochromosome 12p frequency during amniocyte subculturing. Conclusion for strategy of prenatal cytogenetic diagnostics.

Pallister-Killian syndrome (PKS) is characterized cytogenetically by mosaic tetrasomy of chromosome 12p. Routine prenatal diagnosis of PKS is still complicated because of the difficulties of discriminating between the supernumerary isochromosome 12p and the duplication 21q and because of the variable level of mosaicism. The frequency of cells with an extra metacentric chromosome i(12)(p10) is usually determined by tissue-limited or tissue-specific mosaicism. We demonstrated a decrease of the abnormal clone with extra i(12p) in the amniotic fluid cells of the PKS fetus during amniocyte subculturing. The rapid loss of the i(12p) in the course of amniocyte subculturing should be the focus of attention during prenatal karyotyping. This is especially necessary for cultures with slow growth, which require further interpretation of the result during cytogenetic diagnosis of PKS.