Chromosome and cytome analysis of the somatic cells of nuclear-chemical production workers with incorporated 239Pu

The analysis of plutonium production factors has been carried out by using two methodical approaches: assessment of chromosomal aberrations level in routine and G-banded metaphases and molecular-cytogenetic investigation of aneugenic/clastogenic damages in cytokinesis-block binuclear lymphocytes by FISH with centromere specific DNA probes. The obtaining data point out for the first time about both aneugenic and clastogenic influences of incorporated 239Pu with activity range from 0.37 to 6.95 kBq. Correlation analysis of chromosome aberrations with cytome abnormalities allowed finding significant connection between number parameters of metaphase and interphase approaches. The results of this study support the suggestion that aberrant chromosomes are involved preferable in aneugenic events. The FISH technique in binucleated cytokinesis-blocked lymphocytes allows extending of detecting spectrum of chromosome damages and glance of aneugenic mechanisms. Correlations between metaphase and interphase-FISH results point out a high sensitivity of FISH cytome assay, which could be used as an independent test for detection both clastogenic and aneugenic environment influences.     

Isolation and molecular cytogenetic characteristics of the alpha-satellite DNA of human chromosome 6

In this work a molecular-cytogenetic characteristic has been given to the alpha-satellite DNA of chromosome 6. Using different restrictases made it possible to evaluate structural heterogeneity of the obtained subgroup of the alphoid DNA. Polymorphic restriction fragments of DNA have been found which can be used in determining linkage groups.     

The most frequent chromosomal abnormalities in karyotypes of patients with reproductive problems

Results of cytogenetic and molecular-cytogenetic inspection of 210 matrimonial pairs with the problems of reproduction are presented. Different types of chromosomal aberrations have been detected in the karyotypes of the patients in 46 (10.95%) cases. Such structural chromosomal aberrations as pericentric inversions, Robertsonian translocations, balanced reciprocal translocations, and marker chromosomes as well prevailed the numerical chromosomal aberrations (89.13% and 10.87% cases accordingly). In the general group of the inspected patients there were 19 cases (4.52%) characterized by the low level of X and Y chromosome mosaicism. The authors suppose that the patients with the exposed chromosomal abnormalities need the differentiated approach at their treatment.     

Molecular-cytogenetic identification of partial trisomy of a short arm of chromosome 8

We present de novo diagnosed case of partial trisomy of short arm of chromosome 8 with psyhomotoric delay and microanomalies. Inverted duplication of short arm of chromosome 8 was identified using molecular-cytogenetic method. This case is compared with literature data on the same cases. The further intensive study of such cases is necessary to delineate this chromosomal syndrome     

16q subtelomeric deletion in proband with congenital malformations and mental retardation

We present a female child with mild mental retardation and congenital malformations. After fluorescence in situ hybridization (FISH) we found only abnormal karyotype in all cells. We used rapid FISH and original DNA probes--PAC62.10.1 and PAC20.19.N, specific for segments of chromosome 16q24. Karyotype of proband 46,XX.ish del(16)(q24.2:) (PAC20.19.N,PAC62.10.1-). Parent karyotypes are normal. This case may suggest the presence of clinical picture 16q- with defined clinical polymorphism at small telomeric loss, and also its necessary of the use of molecular-cytogenetic techniques in genetic departments.     

Comparative chromosome painting

The review of the data on comparative chromosomal painting in mammals is presented. The development of new molecular-cytogenetic methods has resulted in the accumulation of the detailed information on homology of chromosomal segments of more than 50 species from 11 orders. In this review, modern methods of obtaining painting probes are considered in detail, and the basic tendencies of karyotype evolution in different taxa are discussed. Putative karyotypes of the ancestors of primates, carnivores, and placental mammals are considered.     

Fluorescence <Emphasis Type="Italic">in situ</Emphasis> hybridisation

Fluorescence in situ hybridisation (FISH) is a rapid and reliable technique for chromosomal investigations that is used for a wide variety of cytogenetic purposes at present. This molecular-cytogenetic method has been developed continuously for many years. As a consequence, various modifications with different kinds of fluorescently labelled probes have been introduced to optimise the detection of DNA and RNA sequences. This review articlepaper presents the general principles of in situ hybridisation, probe labelling and examples of proper use of different kinds of probes. In addition, some newer FISH methods and their usefulness in human molecular cytogenetics are described.     

Partial 5p monosomy or trisomy in 11 patients from a family with a t(5;15)(p13.3;p12) translocation

A family with six alive patients with partial monosomy 5p and five with partial trisomy 5p due to a t(5;15)(p13.3;p12) translocation is reported. The translocation was present in four generations with eight balanced carriers. This is the first molecular-cytogenetic and clinical study with both syndromes present in the same family. Using fluorescence in situ hybridization (FISH) with bacterial artificial chromosome (BAC) probes, the breakpoint was mapped to 5p13.3, in the interval corresponding to the BAC clone RP11-1079N14, thereof resulting a 5pter-5p13.3 deletion or duplication of ~32 Mb. These chromosome imbalances can be considered pure, since the other imbalance produced involving chromosome 15p has no phenotypic effect. The presence of several individuals with 5p monosomy and 5p trisomy in the same family is valuable for a better delineation of both syndromes.     

The role of ultrasonic and cytogenetic markers in prenatal diagnostics

As a result of leadthrough 288 invasive manipulations with the use of cytogenetic and molecular-cytogenetic methods we have found 16 different disorders in the fetus karyotype of the expectant mothers of high risk groups. For the most part the Down syndrome and the Shereshevsky-Therner syndrome were detected among aneuploidies. The maximal amount of anomalous karyotypes (28.6%) was detected at the pregnants with congenital malformations in the fetus. The proofs of the increased frequencies of some chromosome homologue heteromorphisms were got in the group of patients where the so-called "soft markers" of aneuploidies or their combination with biochemical markers were used as supposition for the invasive procedure. The echographic screening proved to be the most informative method among the different approaches to the formation of the groups of the high genetic risk.     

Genetic factors of male infertility. The role of complex examination in the case of spermatogenesis disturbances

Data about some genetic factors of male infertility are presented, and methods, which may be used for its diagnosing, are studied. Among genetic factors the following are distinguished: changes in the level of genes (mutations), chromosomes (chromosomal aberrations), and total DNA (chromatin dispersion and DNA fragmentation). As well as standard cytogenetic methods of investigation there are a number of molecular-cytogenetic methods like FISH (fluorescent in situ hybridization), TUNEL (Terminal uridine deoxynucleotidyl transferase dUTP nick end labeling), SCSA (sperm chromatin structural assay), SCGE (single cell gel electrophoresis), and SCD (sperm chromatin dispersion). The thorough study of the sperm of infertile men on several levels of organization let us assess the informativity of each method separately and together, and also develop an optimal algorithm of diagnostics with the aim to choose further tactics in the treatment of male infertility.     

Fluorescence in situ hybridization to chromosomes as a tool to understand human and primate genome evolution

For the last 15 years molecular cytogenetic techniques have been extensively used to study primate evolution. Molecular probes were helpful to distinguish mammalian chromosomes and chromosome segments on the basis of their DNA content rather than solely on morphological features such as banding patterns. Various landmark rearrangements have been identified for most of the nodes in primate phylogeny while chromosome banding still provides helpful reference maps. Fluorescence in situ hybridization (FISH) techniques were used with probes of different complexity including chromosome painting probes, probes derived from chromosome sub-regions and in the size of a single gene. Since more recently, in silico techniques have been applied to trace down evolutionarily derived chromosome rearrangements by searching the human and mouse genome sequence databases. More detailed breakpoint analyses of chromosome rearrangements that occurred during higher primate evolution also gave some insights into the molecular changes in chromosome rearrangements that occurred in evolution. Hardly any "fusion genes" as known from chromosome rearrangements in cancer cells or dramatic "position effects" of genes transferred to new sites in primate genomes have been reported yet. Most breakpoint regions have been identified within gene poor areas rich in repetitive elements and/or low copy repeats (segmental duplications). The progress in various molecular and molecular-cytogenetic approaches including the recently launched chimpanzee genome project suggests that these new tools will have a significant impact on the further understanding of human genome evolution.     

The Xq22 inversion breakpoint interrupted a novel Ras-like GTPase gene in a patient with Duchenne muscular dystrophy and profound mental retardation

A male patient with profound mental retardation, athetosis, nystagmus, and severe congenital hypotonia (Duchenne muscular dystrophy [DMD]) was previously shown to carry a pericentric inversion of the X chromosome, 46,Y,inv(X)(p21.2q22.2). His mother carried this inversion on one X allele. The patient's condition was originally misdiagnosed as cerebral palsy, and only later was it diagnosed as DMD. Because the DMD gene is located at Xp21.2, which is one breakpoint of the inv(X), and because its defects are rarely associated with severe mental retardation, the other clinical features of this patient were deemed likely to be associated with the opposite breakpoint at Xq22. Our precise molecular-cytogenetic characterization of both breakpoints revealed three catastrophic genetic events that had probably influenced neuromuscular and cognitive development: deletion of part of the DMD gene at Xp21.2, duplication of the human proteolipid protein gene (PLP) at Xq22.2, and disruption of a novel gene. The latter sequence, showing a high degree of homology to the Sec4 gene of yeast, encoded a putative small guanine-protein, Ras-like GTPase that we have termed "RLGP." Immunocytochemistry located RLGP at mitochondria. We speculate that disruption of RLGP was responsible for the patient's profound mental retardation.     

Structural unbalanced chromosome rearrangements resolved by comparative genomic hybridization.

The identification of unbalanced structural chromosome rearrangements using conventional cytogenetic techniques depends on recognition of the unknown material from its banding pattern. Even with optimally banded chromosomes, when large chromosome segments are involved, cytogeneticists may not always be able to determine the origin of extrachromosomal material and supernumerary chromosomes. We report here on the application of comparative genomic hybridization (CGH), a new molecular-cytogenetic assay capable of detecting chromosomal gains and losses, to six clinical samples suspected of harboring unbalanced structural chromosome abnormalities. CGH provided essential information on the nature of the unbalanced aberration investigated in five of the six samples. This approach has proved its ability to resolve complex karyotypes and to provide information when metaphase chromosomes are not available. In cases where metaphase chromosome spreads were available, confirmation of CGH results was easily obtained by fluorescence in situ hybridization (FISH) using specific probes. Thus the combined use of CGH and FISH provided an efficient method for resolving the origin of aberrant chromosomal material unidentified by conventional cytogenetic analysis.     

Mapping of <Emphasis Type="Italic">Hieracium</Emphasis> (Asteraceae) chromosomes with genus-specific satDNA elements derived from next-generation sequencing data

The highly repetitive DNA fraction of the eukaryotic genome is considered a mobile, rapidly changing entity, thus reflecting trajectories of short-term evolutionary change. It consists of several large classes in which transposable elements and satellite DNA (satDNA) predominate. Despite a growing awareness of its structure and functional significance, the evolutionary dynamics of repetitive elements and, particularly, satDNA remain poorly characterized. Next-generation sequencing (NGS) has opened up new possibilities for high-throughput genome analysis. Here, we applied satDNA repeatome elements derived from NGS data as probes for fluorescence in situ hybridization to characterize the karyotypes of three diploid hawkweed species of the predominantly polyploid apomictic genus Hieracium, namely H. intybaceum, H. prenanthoides and H. alpinum. Three cluster-distributed, genus-specific satDNA elements that are not present in the sister genus Pilosella were identified; notably, one element spans the functional centromeres. Each of the investigated diploids possessed a species-specific assortment of detected repeats. Their utilization as molecular-cytogenetic markers, in combination with ribosomal DNA loci, allowed for the development of a system to identify the individual chromosomes of the Hieracium species, thus providing a basis for the future investigation of karyotype evolution in diploid hawkweeds and for exploring satDNA dynamics in hybrids and apomicts of allopolyploid origin.     

Specific Genomic Regions Are Differentially Affected by Copy Number Alterations across Distinct Cancer Types, in Aggregated Cytogenetic Data

Background

Regional genomic copy number alterations (CNA) are observed in the vast majority of cancers. Besides specifically targeting well-known, canonical oncogenes, CNAs may also play more subtle roles in terms of modulating genetic potential and broad gene expression patterns of developing tumors. Any significant differences in the overall CNA patterns between different cancer types may thus point towards specific biological mechanisms acting in those cancers. In addition, differences among CNA profiles may prove valuable for cancer classifications beyond existing annotation systems.

Principal Findings

We have analyzed molecular-cytogenetic data from 25579 tumors samples, which were classified into 160 cancer types according to the International Classification of Disease (ICD) coding system. When correcting for differences in the overall CNA frequencies between cancer types, related cancers were often found to cluster together according to similarities in their CNA profiles. Based on a randomization approach, distance measures from the cluster dendrograms were used to identify those specific genomic regions that contributed significantly to this signal. This approach identified 43 non-neutral genomic regions whose propensity for the occurrence of copy number alterations varied with the type of cancer at hand. Only a subset of these identified loci overlapped with previously implied, highly recurrent (hot-spot) cytogenetic imbalance regions.

Conclusions

Thus, for many genomic regions, a simple null-hypothesis of independence between cancer type and relative copy number alteration frequency can be rejected. Since a subset of these regions display relatively low overall CNA frequencies, they may point towards second-tier genomic targets that are adaptively relevant but not necessarily essential for cancer development.     

Analysis of pericentromeric chromosome 21 specific YAC clones by FISH: Identification of new markers for molecular-cytogenetic application

Fluorescence in situ hybridization (FISH) of chromosome 21 specific yeast artificial chromosome (YAC) clones after Alu-PCR (polymerase chain reaction) amplification has been used to find new region-specific DNA probes for the heterochromatic region of chromosome 21. Six overlapping YAC clones from a pericentromeric contig map (region 21cen-21q11) were analyzed. Four YAC clones were characterized as hybridizing to several chromosomal locations. They are, therefore, either chimeric or shared by different chromosomes. Two of them containing alphoid satellite DNA, are localized at the centromeric regions of chromosomes 13 and 21 (clone 243A11), and on 13cen, 21cen and 1q3 (clone 781G5); the two others are localized at both 21q11 and 13q2 (clone 759D3), and at 18p (clone 770B3). Two YACs were strongly specific for chromosome 21q11 only (clones 124A7 and 881D2). These YACs were used effectively as probes for identifications of chromosome 21 during metaphase and interphase analysis of 12 individuals, including three families with Down syndrome offspring, and 6 amniocyte samples. The location of YAC clones on 21q11 close to the centromeric region allows the application of these clones as molecular probes for the analysis of marker chromosomes with partial deletions of the long arm as well as for pre- and postnatal diagnosis of trisomy 21 when alphoid or more distal region-specific DNA probes are uninformative. Overlapping YAC clones covering human chromosome 21q may be systematically used to detect a set of band-specific DNA probes for molecular-cytogenetic application.     

Molecular-cytogenetic studies of ribosomal genes and heterochromatin reveal conserved genome organization among 11 Quercus species

Genomes of 11 Quercus species were characterized using cytogenetic (Giemsa C-banding, fluorochrome banding), molecular-cytogenetic (fluorescence in situ hybridization, FISH, to ribosomal genes) and molecular (dot-blot for ribosomal gene-copy number assessment) techniques. Ribosomal genes are the first DNA sequences to be physically mapped in oaks, and the copy number of the 18S-5.8S-26 S rRNA genes is estimated for the first time. Oak karyotypes were analysed on the basis of DAPI banding and FISH patterns; five marker chromosomes were found. In addition, chromosomal organization of ribosomal genes with respect to AT- and GC-differentiated heterochromatin was studied. Fluorochrome staining produced very similar CMA/DAPI banding patterns, and the position and number of ribosomal loci were identical for all the species studied. The 18S-5.8S-26 S rRNA genes in oak complements were represented by a major locus at the subterminal secondary constriction (SC) of the only subtelocentric chromosome pair and a minor locus at paracentromeric SC of one metacentric pair. The only 5 S rDNA locus was revealed at the paracentromeric region of the second largest metacentric pair. A striking karyotypic similarity, shown by both fluorochrome banding and FISH patterns, implies close genome relationships among oak species no matter their geographic origin (European or American) or their ecophysiology (deciduous or evergreens). Dot-blot analysis gave preliminary evidence for different copy numbers of 18S-5.8S-26 S rRNA genes in diploid genomes of Q. cerris, Q. ilex, Q. petraea, Q. pubescens and Q. robur (2700, 1300, 2200, 4000 and 2200 copies, respectively) that was correlated with the size polymorphism of the major locus. Received: 26 February 1999 / Accepted: 16 March 1999     

Molecular cytogenetics of <Emphasis Type="Italic">Bellevalia</Emphasis> (Hyacinthaceae) species occurring in Greece

Eight taxa of the genus Bellevalia (Hyacinthaceae) occurring in Greece, all with basic chromosome number x = 4, were cytologically studied using classical and molecular-cytogenetic techniques [fluorochrome banding with chromomycin A₃, fluorescence in situ hybridization (FISH) using probes of 18S–5.8S–26S and 5S ribosomal RNA genes]. Two of the examined taxa are endemic, i.e., B. brevipedicellata and B. sitiaca, both restricted to the Island of Kriti. B. hyacinthoides and B. edirnensis are Balkan endemics, and the four remaining taxa, i.e., B. dubia subsp. boissieri, B. trifoliata, B. romana, and B. ciliata, are more widely distributed Mediterranean elements. Genome size, estimated by flow cytometry, ranged from 18.59 to 53.38 pg. The results of fluorochrome banding and FISH are reported for the first time for the genus Bellevalia. Despite the morphological similarity of the chromosome complement, which is in accordance with the general “basic” Bellevalia karyotype formula, the karyotypes of the studied species are clearly distinguished by the number and position of GC-rich bands and ribosomal DNA (rDNA) loci, revealing clear interspecific differentiation among the taxa. Additionally, examination of the polyploid species B. sitiaca and B. edirnensis and populations of B. hyacinthoides and B. ciliata with different ploidy levels permits discussion about the origin of polyploids and the taxonomic relationships among the taxa.     

阿奇霉素联合辛伐他汀对慢性阻塞性肺疾病合并肺动脉高压患者肺功能的影响

目的:探讨阿奇霉素联合辛伐他汀治疗慢性阻塞性肺疾病合并肺动脉高压的临床疗效及对患者肺功能的影响。方法:选取2013年6月-2016年3月我院收治的慢性阻塞性肺疾病合并肺动脉高压患者107例,根据治疗方法不同分为对照组(49例)与实验组(58例)。对照组患者采用阿奇霉素治疗,实验组患者在对照组基础上给予辛伐他汀治疗。观察并比较两组患者的临床疗效、不良反应以及肺功能指标的变化情况。结果:实验组患者治疗有效率(87.93%)高于对照组(73.47%),差异具有统计学意义(P0.05)。与治疗前比较,两组患者治疗后1 s用力呼吸容积(FEV1)、用力肺活量(FVC)及FEV1/FVC水平均升高,差异具有统计学意义(P0.05);与对照组比较,实验组患者治疗后1 s用力呼吸容积(FEV1)、用力肺活量(FVC)及FEV1/FVC水平较高,差异具有统计学意义(P0.05)。治疗后,两组患者血清总胆固醇(TC)及三酰甘油(TG)水平均降低,差异具有统计学意义(P0.05);与对照组比较,实验组患者治疗后血清总胆固醇(TC)及三酰甘油(TG)水平较低,差异具有统计学意义(P0.05)。两组患者不良反应发生率比较,差异无统计学意义(P0.05)。结论:阿奇霉素联合辛伐他汀治疗慢性阻塞性肺疾病合并肺动脉高压的临床效果显著,不仅能够改善患者肺功能,降低血脂相关指标水平,并且安全性较高,值得临床推广应用。     

Association of arytenoid chondritis with equine lymphocyte antigens but no association with laryngeal hemiplegia, umbilical hernias and cryptorchidism

Associations were sought between ELA A1-A10 and W11 antigens and the presence of laryngeal hemiplegia, arytenoid chondritis, umbilical hernias and cryptorchidism in Thoroughbreds and/or Quarter Horses. No significant associations were detected between laryngeal hemiplegia and any ELA antigen in Thoroughbreds. The association between arytenoid chondritis and A9 was significant with a relative risk (RR) of 15.6 and aetiologic fraction (EF) of 0.80 in Thoroughbreds. There were apparent associations based on RR between A4 and A5 in Quarter Horses with umbilical hernias (RR = 7.5 and 6.1 respectively); however, these were not statistically significant. No significant associations were detected with cryptorchidism in Quarter Horses when the control population included both sexes. When only unaffected males were used as the control group, there was an apparent increase in relative risk with A6 (from RR = 1.7 to 4.3); however this was not statistically significant. Cryptorchidism in Thoroughbreds showed an increased relative risk with A5 regardless of whether the control population included males and females (RR = 4.1) or only males (RR = 4.7) but the increases were not statistically significant.