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Biomedical subjects

Uwe Claussen

Publications and source records attributed to Uwe Claussen.

At least 19 recordsLinked to original sources
PubMed2003

Somatic mosaicism for maternal uniparental disomy 15 in a girl with Prader-Willi syndrome: confirmation by cell cloning and identification of candidate downstream genes.

Although uniparental disomy often results from the postzygotic rescue of a meiotic non-disjunction event, mosaicism is usually confined to the placenta. We describe a girl with Prader-Willi syndrome (PWS) who is mosaic for normal cells and cells with maternal uniparental disomy 15 [upd(15)mat] in blood and skin. Somatic mosaicism was confirmed by cloning and genotyping of skin fibroblasts. X inactivation studies indicated that upd occurred prior to X inactivation. RNA samples from the cloned cells were used in DNA microarray experiments to study the effect of upd(15)mat on the gene expression pattern of fibroblasts. Proof of principle was obtained by detecting several chromosome 15 genes known to be imprinted. We did not obtain any evidence for novel 15q genes showing imprinted expression in fibroblasts. Differentially expressed genes on other chromosomes are candidates for downstream genes regulated by an imprinted gene and may play a role in the pathogenesis of PWS. The finding of strongly reduced mRNA levels in upd(15)mat cells of the gene encoding secretogranin II (SCG2), which is a precursor of the dopamine releasing factor secretoneurin, raises the question whether hyperphagia in patients with PWS might be due to a defect in dopamine-modulated food reward circuits.

Adult↗
PubMed2003

A multiple translocation event in a patient with hexadactyly, facial dysmorphism, mental retardation and behaviour disorder characterised comprehensively by molecular cytogenetics. Case report and review of the literature.

UNLABELLED: We report a 13-year-old female patient with multiple congenital abnormalities (microcephaly, facial dysmorphism, anteverted dysplastic ears and postaxial hexadactyly), mental retardation, and adipose-gigantism. Ultrasonography revealed no signs of a heart defect or renal abnormalities. She showed no speech development and suffered from a behavioural disorder. CNS abnormalities were excluded by cerebral MRI. Initial cytogenetic studies by Giemsa banding revealed an aberrant karyotype involving three chromosomes, t(2;4;11). By high resolution banding and multicolour fluoresence in-situ hybridisation (M-FISH, MCB), chromosome 1 was also found to be involved in the complex chromosomal aberrations, confirming the karyotype 46,XX,t(2;11;4).ish t(1;4;2;11)(q43;q21.1;p12-p13.1;p14.1). To the best of our knowledge no patient has been previously described with such a complex translocation involving 4 chromosomes. This case demonstrates that conventional chromosome banding techniques such as Giemsa banding are not always sufficient to characterise complex chromosomal abnormalities. Only by the additional utilisation of molecular cytogenetic techniques could the complexity of the present chromosomal rearrangements and the origin of the involved chromosomal material be detected. Further molecular genetic studies will be performed to clarify the chromosomal breakpoints potentially responsible for the observed clinical symptoms. CONCLUSION: This report demonstrates that multicolour-fluorescence in-situ hybridisation studies should be performed in patients with congenital abnormalities and suspected aberrant karyotypes in addition to conventional Giemsa banding.

Abnormalities, Multiple↗
PubMed2003

Human adenoma cells are highly susceptible to the genotoxic action of 4-hydroxy-2-nonenal.

Oxidative stress and resulting lipid peroxidation are important risk factors for dietary-associated colon cancer. To get a better understanding of the underlying molecular mechanisms, we need to characterise the risk potential of the key compounds, which cause DNA damage in cancer-relevant genes and especially in human target cells. Here, we investigated the genotoxic effects of 4-hydroxy-2-nonenal (HNE) and hydrogen peroxide (H(2)O(2)) in human colon cells (LT97). LT97 is a recently established cell line from a differentiated microadenoma and represents cells from frequent preneoplastic lesions of the colon. The genomic characterisation of LT97 was performed with 24-colour FISH. Genotoxicity was determined with single cell microgelelectrophoresis (Comet assay). Comet FISH was used to study the sensitivity of TP53-a crucial target gene for the transition of adenoma to carcinoma-towards HNE. Expression of glutathione S-transferases (GST), which deactivates HNE, was determined as GST activity and GSTP1 protein levels. LT97 cells were compared to primary human colon cells and to a differentiated clone of HT29. Karyotyping revealed that the LT97 cell line had a stable karyotype with only two clones, each containing a translocation t(7;17) and one aberrant chromosome 1. The Comet assay experiments showed that both HNE and H(2)O(2) were clearly genotoxic in the different human colon cells. HNE was more genotoxic in LT97 than in HT29clone19A and primary human colon cells. After HNE incubation, TP53 migrated more efficiently into the comet tail than the global DNA, which suggests a higher susceptibility of the TP53 gene to HNE. GST expression was significantly lower in LT97 than in HT29clone19A cells, which could explain the higher genotoxicity of HNE in the colon adenoma cells. In conclusion, the LT97 is a relevant model for studying genotoxicity of colon cancer risk factors since colon adenoma are common preneoplastic lesions occurring in advanced age.

Adenoma↗
PubMed2003

Ten years follow up of a boy with a complex chromosomal rearrangement: going from a > 5 to 15-breakpoint CCR.

A moderately mentally retarded 10-year-old boy of very short stature was found initially to have a complex chromosomal rearrangement (CCR) involving chromosome 1, 2, 3, 4, and 8. A balanced twelve-breakpoint CCR was suggested after extensive investigations including subtelomere FISH, whole chromosome paints, comparative genomic hybridization (CGH), multicolor FISH (MFISH), and spectral karyotyping (SKY). SKY and MFISH gave slightly discrepant results. For further clarification of the karyotype, multicolor banding (MCB) analysis and FISH with region-specific YAC probes were done. This allowed clarification of a sixteen-fragment CCR to be made, the most complex constitutional chromosomal rearrangement reported so far. Remarkably, two 'secondary' insertions originated from the interior of a 'primary' insertion by an excision/duplication event. The randomness of the fragments and the complexity of the derivative chromosomes suggest that this CCR is the result of a single meiotic event, e.g., faulty repair of a five-chromosome knot.

Child↗
PubMed2003

First patient with trisomy 21 accompanied by an additional der(4)(:p11 --> q11:) plus partial uniparental disomy 4p15-16.

We report on a rare additional numerical chromosomal aberration in a child with Down syndrome due to free trisomy 21. The karyotype showed 48,XY,+21,+mar after GTG banding, with the marker present in 80% of cells. The supernumerary marker chromosome (SMC) was as small as approximately one-third of 18p, and with the recently developed centromere-specific multi-color fluorescence in situ hybridization (cenM-FISH) technique, it was shown that the SMC was a derivative chromosome 4. The SMC was not specifically stained by arm-specific probes for chromosome 4; thus, it has been described as der(4)(:p11 --> q11:). Microsatellite analysis resulted in a partial maternal uniparental isodisomy (UPD) for chromosome 4p15-16 and a maternal origin for two chromosomes 21. Until now only two similar cases have been described in the literature, but without clarifying the origin of the SMC and without looking for an additional UPD. This is the only reported case of a UPD 4p in a liveborn child.

Chromosome Aberrations↗
PubMed2003

Synovial fibroblasts and synovial macrophages from patients with rheumatoid arthritis and other inflammatory joint diseases show chromosomal aberrations.

Chromosomal aberrations were investigated in nuclei extracted from synovial tissue and first-passage synovial fibroblasts (P-1 SFB, 98% enrichment) or macrophages (P-1 Mphi) from patients with rheumatoid arthritis (n=10). The findings were compared with those in other rheumatic diseases (osteoarthritis, n=14; reactive arthritis, n=1), as well as with those in chronic obstructive pulmonary disease (n=8). Controls were paired peripheral blood lymphocytes from arthritic patients, synovial tissue or SFB/Mphi from joint trauma/normals (n=9), and peripheral blood monocytes from normal donors (n=10). GTG banding of metaphase chromosomes and interphase fluorescence in situ hybridization with centromere-specific probes were used. Comparable chromosomal aberrations were observed in synovial tissue and P-1 SFB of patients with rheumatoid arthritis, osteoarthritis, and reactive arthritis (polysomy 7 and aneusomies of chromosomes 4, 8, 9, 12, and 18). Notably, aneusomies of chromosomes 4, 6, 7, 8, 9, 11, 12, and/or X were also detected in P-1 synovial Mphi from rheumatoid arthritis (90% of the cases), osteoarthritis (93%), and reactive arthritis (1/1), as well as bronchial Mphi from chronic obstructive pulmonary disease (25%). No aberrations were detected in paired peripheral blood lymphocytes (except for one osteoarthritis case with a karyotype 45,X[10]/46,XX[17]), or in peripheral blood monocytes and synovial tissue of normals/joint trauma. Because Mphi aberrations were common to chronic joint and pulmonary disease, chronic inflammatory stress may induce chromosomal aberrations with potential functional relevance in local mesenchymal cells and infiltrating leukocytes in an organ-independent fashion.

Adult↗
PubMed2003

Residual chromosomal damage after radiochemotherapy with and without amifostine detected by 24-color FISH.

BACKGROUND: Amifostine is a radioprotective drug applied to reduce acute radiation toxicity during a course of conventionally fractionated radiotherapy. In the present study, amifostine was used in patients undergoing adjuvant radiochemotherapy for rectal cancer. It was described previously that additional application of amifostine led to less acute skin and bowel toxicity. The present study was aimed to determine whether amifostine has an influence on the amount of residual chromosomal damage. MATERIAL AND METHODS: Peripheral lymphocytes of twelve rectal cancer patients who had undergone postoperative radiochemotherapy 2-3 years ago were investigated for residual chromosomal damage using 24-color fluorescence in situ hybridization (24-color FISH). All twelve patients had received a total dose of 55.8 Gy in conventional fractionation of 1.8 Gy and a 120-h continuous infusion of 5-fluorouracil (5-FU) chemotherapy (1,000 mg/m(2) per day) in the 1st and 5th week of irradiation. Seven out of twelve patients had been given additional amifostine on chemotherapy days (500 mg total dose as short i.v. infusion immediately prior to the daily radiation fraction). Cultivation of lymphocytes and 24-color FISH were performed according to standard protocols. 100 metaphases per patient were analyzed for chromosomal aberrations in a blind study. RESULTS: Analysis of the average number of breaks per mitosis (B/M) revealed an increased amount of residual chromosomal damage in the group treated with amifostine (0.65 B/M [0.32-0.97]) as well as in those treated without amifostine (0.76 B/M [0.31-1.25]). Also the average number of cells containing aberrations per 100 analyzed metaphases was similar (with amifostine: 22.1 [13-32] vs. 24.4 [13-35] without amifostine). The aberration types, occurring as simple translocations, reciprocal translocations, breaks, dicentrics, inversions, rings and complex chromosomal rearrangements, did not show any specific accumulation in one or the other group either. CONCLUSION: While there was a significant amifostine-mediated clinical amelioration of normal tissue toxicity, the comparison of residual chromosomal damage 2-3 years after completion of radiochemotherapy was characterized by a high interindividual variation, and no equivalent difference could be detected between the two groups.

Aged↗
PubMed2003

Fluorescence in situ hybridization (FISH) on human chromosomes using photoprobe biotin-labeled probes.

Fluorescence in situ hybridization (FISH) on human chromosomes in meta- and interphase is a well-established technique in clinical and tumor cytogenetics and for studies of evolution and interphase architecture. Many different protocols for labeling the DNA probes used for FISH have been published. Here we describe for the first time the successful use of Photoprobe biotin-labeled DNA probes in FISH experiments. Yeast artificial chromosome (YAC) and whole chromosome painting (wcp) probes were tested.

Biotin↗
PubMed2003

Highly complex karyotypic changes in acute myelogenous leukemia: a case report.

We report on a patient with a clinically diagnosed acute myelogenous leukemia (AML) with so far unrecorded complex translocations. As GTG-banding was not able to resolve all karyotypic changes, spectral karyotyping (SKY) and multiplex-fluorescence in situ hybridization (M-FISH) were performed for comparison. Both methods gave nearly identical results, however, they were unable to characterize all involved chromosomal breakpoints in detail. Thus, multicolor banding (MCB) technique was applied and its results were confirmed for two large derivative chromosomes by microdissection and reverse painting. Using this battery of molecular cytogenetic approaches the karyotype of this AML case could be described as 40 approximately 44,XY,der(1)t(1;5;8;20) (1qter-->1p12::5q14.3-->5q15 or 5q15-->5q14.3::8p11.2-->8p23.? 3::20p11.1-->20p13),del(2)(q12),der(3)t(3;6),der(5)t(5;18) (5p15.33-->5q11::18q21.3-->18q23),del(6),-8,der(9)t(9;17;15),der(10)t(3;10),del(11)(q24),-15,-16,del(17),der(18) t(8;18;5;2;20)(8q24.3-->8q24.2 or 8q24.2-->8q24.3::18p11.22-->18q21.3::5q14.3-->5q11::2q32-->2q12::20q13.2-->20q13.33), der(20)t(1;20;18)(1p36.33-->1p31.3-22.3::20p11.1-->20q11.2 or 20q11.2-->20p11.1::18p11.22-->18p11.32).

Chromosome Banding↗
PubMed2003

Detailed Hylobates lar karyotype defined by 25-color FISH and multicolor banding.

A comprehensive and detailed comparative chromosome map of the white-handed gibbon (Hylobates lar = HLA) has been established by hybridizing the recently developed complete human multicolor banding (MCB) probe set on metaphase chromosomes of a male HLA lymphoblastoid cell line. Thus, it was possible to precisely determine the breakpoints and distribution plus orientation of specific DNA-regions in this cytogenetically highly rearranged species compared to Homo sapiens (HSA). In general, the obtained results are in concordance with previous molecular-cytogenetic studies. In this study all 71 breakpoints present in HLA compared to HSA could be determined exactly. This study is a valuable complement to our knowledge on the phylogeny of huminoid chromosomes.

Animals↗
PubMed2002

The DNA-based structure of human chromosome 5 in interphase.

In contrast to those of metaphase chromosomes, the shape, length, and architecture of human interphase chromosomes are not well understood. This is mainly due to technical problems in the visualization of interphase chromosomes in total and of their substructures. We analyzed the structure of chromosomes in interphase nuclei through use of high-resolution multicolor banding (MCB), which paints the total shape of chromosomes and creates a DNA-mediated, chromosome-region-specific, pseudocolored banding pattern at high resolution. A microdissection-derived human chromosome 5-specific MCB probe mixture was hybridized to human lymphocyte interphase nuclei harvested for routine chromosome analysis, as well as to interphase nuclei from HeLa cells arrested at different phases of the cell cycle. The length of the axis of interphase chromosome 5 was determined, and the shape and MCB pattern were compared with those of metaphase chromosomes. We show that, in lymphocytes, the length of the axis of interphase chromosome 5 is comparable to that of a metaphase chromosome at 600-band resolution. Consequently, the concept of chromosome condensation during mitosis has to be reassessed. In addition, chromosome 5 in interphase is not as straight as metaphase chromosomes, being bent and/or folded. The shape and banding pattern of interphase chromosome 5 of lymphocytes and HeLa cells are similar to those of the corresponding metaphase chromosomes at all stages of the cell cycle. The MCB pattern also allows the detection and characterization of chromosome aberrations. This may be of fundamental importance in establishing chromosome analyses in nondividing cells.

Cell Cycle↗
PubMed2002

Heterogenic molecular basis for loss of ABL1-BCR transcription: deletions in der(9)t(9;22) and variants of standard t(9;22) in BCR-ABL1-positive chronic myeloid leukemia.

The objective of this study was to characterize the ABL1-BCR fusion gene in 76 BCR-ABL1-positive chronic myeloid leukemia (CML) patients regarding expression as well as genomic status, to assess the frequency of ABL1-BCR gene deletion in these patients, which has been reported to be an adverse prognostic factor in Philadelphia chromosome-positive CML. Patients were analyzed for ABL1-BCR 1b-b3 and/or 1b-b4 transcription by RT-PCR analysis. ABL1-BCR gene status was analyzed by FISH in 16 CML patients with no ABL1-BCR transcript. FISH revealed a partial or total deletion of the ABL1-BCR gene in 9/16 and localized the 5' portion of ABL1 and the 3' portion of BCR at separated loci in 5/16 patients. The latter FISH pattern resulted from a nonreciprocal translocation in two and a complex translocation in three individuals. In 2/16 patients, FISH could not exclude an intact ABL1-BCR fusion gene. Thus, most CML patients without ABL1-BCR transcript could be characterized cytogenetically to belong to two major subgroups: a silent ABL1-BCR gene was attributed to a deletion in der(9)t(9;22) in 56% of the investigated patients or to variants of a standard t(9;22) (approximately 31%). Conversely, none of the 50 patients with an ABL1-BCR transcript exhibited a variant t(9;22) in GTG-banding analysis. Thus, genomic aberrations such as deletions or complex genomic rearrangements are the basic and most frequent cause for ABL1-BCR RNA negativity in CML. The heterogeneity of the underlying molecular mechanisms may explain divergent clinical implications described for patients with an ABL1-BCR deletion and those with no ABL1-BCR transcript.

Chromosome Deletion↗
PubMed2002

Is 24-color FISH detection of in-vitro radiation-induced chromosomal aberrations suited to determine individual intrinsic radiosensitivity?

BACKGROUND: Reliable determination of intrinsic radiosensitivity in individual patients is a serious need in radiation oncology. Chromosomal aberrations are sensitive indicators of a previous exposure to ionizing irradiation. Former molecular cytogenetic studies showed that such aberrations as an equivalent of intrinsic radiosensitivity can be detected by fluorescence in-situ hybridization (FISH) techniques using whole chromosome painting (wcp) probes. However, only one up to three randomly chosen wcp probes have been applied for such approaches until now. As a random distribution of chromosomal rearrangements along the chromosomes is up to now still controversial, the power of the 24-color FISH approach should be elucidated in the present study. METHODS AND MATERIAL: Lymphocytes derived from lymphoblastoid cell lines of one patient with Nijmegen breakage syndrome (NBS homozygote) and of two NBS heterozygotes and peripheral blood lymphocytes of two controls were analyzed. Samples of each patient/control were irradiated in vitro with 0.0 Gy, 0.7 Gy or 2.0 Gy prior to cultivation. Chromosomal aberrations were analyzed in detail and quantified by means of 24-color FISH as an expression of the individual intrinsic radiosensitivity. RESULTS: 24-color FISH analyses were done in a total of 1,674 metaphases. After in-vitro irradiation, 21% (0.7 Gy) or 57% (2.0 Gy) of the controls' cells, 15% (0.7 Gy) or 53% (2.0 Gy) of the heterozygotes' cells and 54% (0.7 Gy) or 79% (2.0 Gy) of the homozygote's cells contained aberrations. The highest average rates of breaks per mitosis [B/M] (0.7 Gy: 1.80 B/M, 2.0 Gy: 4.03 B/M) and complex chromosomal rearrangements [CCR] (0.7 Gy: 0.20 CCR/M, 2.0 Gy: 0.47 CCR/M) were observed in the NBS patient. Moreover, the proportion of different aberration types after irradiation showed a distinct increase in the rate of CCR combined with a decrease in dicentrics in the NBS homozygote. CONCLUSION: To come to a more complete picture of radiation-induced aberrations and to detect and quantify genetically determined intrinsic radiosensitivity, a 24-color FISH approach using all human chromosome painting probes has been successfully applied on cytogenetic preparation lymphocytes. The controls and NBS heterozygotes were clearly distinguished from the NBS homozygote subject.

Adult↗
PubMed2002

Homologous sequences at human chromosome 9 bands p12 and q13-21.1 are involved in different patterns of pericentric rearrangements.

A thorough study of the heterochromatin organisation in the pericentromeric region and the proximal long (q) and short (p) arms of human chromosome 9 (HSA 9) revealed homology between 9p12 and 9q13-21.1, two regions that are usually not distinguishable by molecular cytogenetic techniques. Furthermore, the chromosomal regions 9p12 and 9q13-21.1 showed some level of homology with the short arms of the human acrocentric chromosomes. We studied five normal controls and 51 clinical cases: 48 with chromosome 9 heteromorphisms, one with an exceptionally large inversion and two with an additional derivative chromosome 9. Using fluorescence in situ hybridisation (FISH) with three differentially labelled chromosome 9-specific probes we were able to distinguish 12 heteromorphic patterns in addition to the most frequent pattern (defined as normal). In addition, we studied one inversion 9 case with the recently described multicolour banding (MCB) technique. Our results, and previously published findings, suggest several hotspots for recombination in the pericentromeric heterochromatin of HSA 9. They also demonstrate that constitutional inversions affecting the pericentromeric region of chromosome 9 carry breakpoints located preferentially in 9p12 or 9q13-21.1 and less frequently in 9q12.

Chromosome Aberrations↗