G-banding Research Articles

Cytogenetic evaluation of patients with clinical spectrum of Turner syndrome

AIM:The objective of this study was to correlate the genotype, of female patients, withshort stature and primary amenorrhea.MATERIALS AND METHODS:One hundred and forty-six subjects were recruited during 2005-2012. Microscopic and automated karyotyping analyses were done by using chromosomes isolated from the lymphocytes using Giemsa banding (GTG) to identify chromosome abnormalities.RESULTS:A total of 146 clinically suspected Turner syndrome (TS) subjects were recruited for the study, of which, 61 patients were identified to have chromosome abnormalities. The chromosomal abnormalities detected were as follows: Monosomy X (n = 19, 13.01%), triple X syndrome (n = 4, 2.7%), mosaic TS (n = 12, 8.21%), XY gonadal dysgenesis (n = 13, 8.9%), and structural abnormalities including X chromosome (n = 15, 10.27%) and one patient each with autosomal changes involving 9qh inversion and translocation of chromosomes 12 and 14.CONCLUSION:Karyotype abnormalities accounting for 46% in this study emphasize the need for karyotype testing in cases of short stature with primary amenorrhea.

Amelioration of melatonin on oxidative stress and genotoxic effects induced by cisplatin in vitro

In this study, we made an effort to evaluate the possible protective actions of melatonin on cisplatin-induced oxidative damage in mice brain homogenate and genotoxic effects in human lymphocytes under in vitro conditions. The tissue homogenate was divided into three parts. The first portion was kept as control treated with dimethyl sulphoxide (DMSO) (group 1) while the second and third portion were treated with 24 µg/g tissue cisplatin alone (group 2) and 24 µg/g tissue cisplatin in combination with 3 mM melatonin (group 3), respectively. We measured the oxidative stress biomarkers such as lipid peroxidation, 8-hydroxy 2′ deoxyguanosine (8-OHdG) and antioxidant parameters such as reduced glutathione, superoxide dismutase, glutathione peroxidase, and glutathione reductase in brain homogenate. Likewise peripheral venous blood was collected from healthy donors and human lymphocyte culture was done using karyotyping medium. Cultures were divided into three groups. Group 1 was the control i.e. lymphocytes treated with DMSO 5 µg/mL. In group 2, lymphocytes were treated with 2 µg/mL cisplatin and group 3 with a combination of 2 µg/mL cisplatin and 0.3 mM melatonin. Incubation of tissue homogenates with cisplatin elevated the malondialdehyde and 8-OHdG levels which were then reversed by melatonin. Reduction in antioxidant parameters with respect to corresponding controls were also restored by melatonin treatment. Furthermore, supplementation of melatonin was found to modulate the chromosome damage elicited by cisplatin which was determined using Giemsa (GTG) banding and karyotyping. These findings suggest that melatonin improves the cellular function and helps them to survive in the belligerent environment created by free radicals.

Clinical and laboratory features of pediatric acute myeloid leukemia with inversion of chromosome 16

To evaluate the clinical and laboratory features of pediatric inv(16) acute myeloid leukemia (AML) retrospectively. Dual color fluorescence in situ hybridization (D-FISH) using a LSI CBFβ inv(16) break apart probe labeled by Spectrum red and Spectrum green was performed in 15 acute myeloid leukemia cases, including 13 cases with or without abnormal eosinophils but with positive core binding factor β (CBFβ)-MYH11 fusion transcript detected by RT-PCR, and 2 cases with trisomy 8 (+8). The results were compared with the morphology, immunophenotype, karyotype and RT-PCR. Morphologically, 12 cases were diagnosed as M(4)EO, 2 as M(4), and 1 as M(2a). Immunophenotypically, all 13 AML cases with inv(16) showed positive expression of CD(13) and CD(33), but without the expression of any lymphoid lineage antigens. Karyotyping analysis with G-banding detected inv(16) in 10 AML cases, including 9 M(4)EO cases and 1 M(2a), but only 5 positive cases were detected using R-banding technique. Among them, 2 cases had simultaneous +8 and trisomy22 (+22), one had +22 only in addition to inv(16). D-FISH revealed a CBFβ-MYH11 rearrangement in 13 cases of AML with positive RT-PCR results, and the mean positive rate of cell detection was 55.15% (range 37.0% - 86.0%). The complete remission rate (CR) and median survival period in this series of inv(16) AML were 81.5%and 11 months, respectively, of whom, 8 cases were still in CR. Relapse and karyotypic evolution were seen in case 5 with +8, +22 in addition to inv(16). AML with inv(16) is a special subtype. Most cases belong to M(4)EO. Its prognosis is good in general, but it seems to be an unfavorable feature for AML with inv(16) and +8, +22 simultaneously, especially with karyotypic evolution. For detection of inv(16), G-banding technique is evidently superior to R-banding technique. D-FISH combined with RT-PCR are more sensitive and reliable than chromosome banding analysis.

Standardized karyotype and idiogram of Thai’s native cattle, Bos indicus (Artiodactyla, Bovidae) by convention staining, G-banding, C-banding and NOR-banding techniques

Cytogenetics of Thai’s native cattle ( Bos indicus ) at Rajamangala University of Technology Isan, Surin Campus, was studied. Blood samples were taken from two males and two females. After standard whole blood lymphocyte culture at 37 O C for 72 hours in presence of colchicine, the metaphase spreads were performed on microscopic slide and air-dried. Cytogenetic techniques including conventional staining, G-, C- and NOR-bandings were applied to the chromosomes. The diploid chromosomes number was 2 n =60, the fundamental numbers (NF) were 61 in male and 62 in female. All autosomes were acrocentric with 18 large, 20 medium and 20 small acrocentric chromosomes. The X chromosome was a large submetacentric and the Y was the smallest acrocentric. From G-banding, each chromosome pair appeared with clearly differentiated. C-banding shown C-positive at centromeric region of all chromosomes. NOR-banding staining for nuclear organizer regions (NORs) showed 6 NORs represented at telomeric end of chromosomes. The karyotype formula of Thai’s native cattle was as follows: 2 n (diploid) 60 = L a 18 +M a 20 +S a 20 + sex chromosomes

Acute megakaryoblastic leukemia and severe pulmonary fibrosis in a child with down syndrome: Successful treatment with ultra low-dose cytarabine using GATA1 mutation to monitor minimal residual disease

Acute megakaryoblastic leukemia and severe pulmonary fibrosis in a child with down syndrome: Successful treatment with ultra low-dose cytarabine using GATA1 mutation to monitor minimal residual disease

Detection of NPM1 gene mutation in acute myeloid leukemia

Objective To analyze the frequency of NPM1 mutation in de novo acute myeloid leukemia （AML） patients and the relationship between NPM1 mutation and chromosome alterations, as well as FAB subgroups, and to analyze the mutation type. Methods A total of 99 de novo AML patients from 2004 to 2010 in China-Japan Friendship Hospital were studied.Genomic DNA was amplified by polymerase chain reaction （PCR）, denaturing polyacrylamide gel electrophoresis （PAGE） and capillary electrophoresis were used to detect the mutation of NPM1 gene in 99 AML patients, and karyotyping was performed in 72 AML patients by G banding techniques.DNA sequences analysis of NPM1 mutation was performed on 10 patients.Chi-square test was used to compare the frequencies of NPM1 mutation among the different subgroups, and McNemar′s test was used to compare the different rates between denaturing PAGE and capillary electrophoresis. Results The frequencies of NPM1 mutations were detected in 15% (15/99) of AML patients with capillary electrophoresis and 11%(11/99) with denaturing PAGE(χ2=2.25，P>0.05).The NPM1 was at different rates in M2(27%, 8/30), M5(32%, 6/19), M6(13%, 1/8), respectively (χ2=1.06，P>0.05), and not detected in the other subgroups.NPM1 mutation in patients with normal karyotype(26%) was more prevalent than patients with abnormal karyotype(4%)(χ2=5.61，P<0.05).All of the 10 patients were of A type (c.860_863dupTCTG).The C-terminal portion of the NPM protein by replacing the last seven amino acids(WQWRKSL) with 11 residues (CLAVEEVSLRK).Two intronic deletions were novel, one case was IVS10-18_-15delCTTT, the other was IVS10-17_-15delTTT. Conclusions NPM1 mutations represents a common genetic abnormality in AML patients, and NPM1 mutation in patients with normal karyotype is higher than patients with abnormal karyotype.Two new intronic deletion mutations are identified.(Chin J Lab Med, 2012，35：27-31) Key words: Leukemia，myeloid，acute; Nuclear proteins; Mutation; Electrophoresis，capillary; Electrophoresis，polyacrylamide gel

Polysomy 8 in Egyptian patients with de-novo acute myeloid leukemia

Background Tetrasomy, pentasomy, and hexasomy of the entire chromosome 8 (polysomy 8) are relatively rare compared with trisomy 8, which is one of the most common recurring aberrations in myeloid hematologic malignancies. Although polysomy 8 appears to adversely influence outcome, little is known about the prognostic significance of polysomy 8 in patients with acute myeloid leukemia (AML). Aim of the work The aim of the present work was to study the numerical aberrations of chromosome 8 in Egyptian patients with AML and to clarify its role as a prognostic marker. Patients and methods The present study was carried out on 76 newly diagnosed adult patients with AML. They were attending the Hematology Department of Ain Shams University Hospitals. They were subjected to conventional cytogenetic analysis using G-banding and fluorescence in-situ hybridization using a fluorophore-labeled centromeric probe for chromosome 8. Results Metaphase and interphase fluorescence in-situ hybridization for detection of polysomy 8 was successfully performed on 72 samples. Patients were divided into two groups: group A comprised patients associated with polysomy 8 (20 patients; 27.8%) and group B comprised patients without polysomy 8 (52 patients; 72.2%). Group A patients had a statistically significant younger age compared with group B (P=0.04). A statistically significant association was seen between polysomy 8 status and hepatosplenomegaly, fever, and lymph node infiltration (P=0.03, 0.05). The presence of hepatosplenomegaly, skin infiltration, lymph node infiltration, and French–American–British subtype was shown to have a significant impact on patient outcome (P=0.05, 0.03, 0.01, and 0.04, respectively). Patients with skin infiltration, CD34 expression, HLA-DR expression, high/intermediate cytogenetic risk, and presence of polysomy 8 had significantly shorter overall survival. Conclusion and recommendations From the current study we conclude that polysomy 8 occurs in 27.8% of de-novo AML cases; 35% of these cases showed tetrasomy of chromosome 8. Polysomy of chromosome 8 was associated with shortened overall survival. Further studies with a larger number of patients are recommended for proper assessment of polysomy 8, especially tetrasomy 8, among Egyptian patients with AML.

Nucleophosmin (NPM1-A) and FMS-like tyrosine kinase 3 (FLT3/ITD) mutations in acute myeloid leukaemia with a normal karyotype ( frequency and prognostic significance)

Background Treatment of acute myeloid leukaemia (AML) with a normal karyotype (NK) has always been a challenge for clinicians. This is because it affects the majority of AML patients and falls into the intermediate-risk group that needs further risk stratifications. Aim of the study The current study aimed to verify the importance and significance of analysis of NPM1-A and FLT3/ITD mutations, alone and combined, and their relation to prognostic criteria. Patients and methods Thirty-five patients with de-novo AML with a median age of 33 years (19–60) and ten healthy volunteers (the control group) were included. All participants were subjected to clinical examination, routine laboratory investigations and (for patients only) bone marrow examination and cytogenetic analysis by the G-banding technique. Using the genomic PCR (RT and conventional) technique, NPM1 type A and FLT3/ITD mutations were studied in the NK-AML patients and controls. Their impacts were evaluated alone and combined. Results After cytogenetic analysis 20/35 (57.1) patients proved to be of NK; their median age was 37.5 (19–60) years. This NK-AML group and control group were examined for NPM1 and FLT3/ITD mutations. None of the controls had either mutation. The NK group was categorized into NPM1 + or NPM1 − and FLT3 + or FLT3 −. Their frequencies were 40% (8), 60% (12) and 40% (8), 60 (12), respectively. These subgroups were assessed after induction chemotherapy for achievement of complete remission (CR), which was recorded in 62.5% of NPM1 -positive patients and 0% of NPM1 -negative patients, showing a significant difference with a P value of 0.004, whereas nonsignificant difference between FLT3 -positive and FLT3 -negative patients was recorded. The NK patients were further subdivided into four genetic subgroups: NPM+/ FLT3 −, NPM−/ FLT3 −, NPM+/ FLT3 + and NPM−/ FLT3 +, with frequencies of 15, 45, 25 and 15%, respectively. Sixty per cent of patients who achieved CR were NPM1 +/ FLT3 −. Furthermore, 100% of this subgroup achieved CR, 40% of NPM+/ FLT3 + also achieved CR, but none of the NPM−/ FLT3 − and NPM1 −/ FLT3 + patients did. Conclusion Genotypes defined by the mutational status of NPM1 , FLT3 are associated with the outcome of treatment in NK-AML patients.

A novel X‐linked disorder with developmental delay and autistic features

Genomic duplications that lead to autism and other human diseases are interesting pathological lesions since the underlying mechanism almost certainly involves dosage sensitive genes. We aim to understand a novel genomic disorder with profound phenotypic consequences, most notably global developmental delay, autism, psychosis, and anorexia nervosa. We evaluated the affected individuals, all maternally related, using childhood autism rating scale (CARS) and Vineland Adaptive scales, magnetic resonance imaging (MRI) and magnetic resonance spectroscopy (MRS) brain, electroencephalography (EEG), electromyography (EMG), muscle biopsy, high-resolution molecular karyotype arrays, Giemsa banding (G-banding) and fluorescent in situ hybridization (FISH) experiments, mitochondrial DNA (mtDNA) sequencing, X-chromosome inactivation study, global gene expression analysis on Epstein-Barr virus (EBV)-transformed lymphoblasts, and quantitative reverse-transcription polymerase chain reaction (qRT-PCR). We have identified a novel Xq12-q13.3 duplication in an extended family. Clinically normal mothers were completely skewed in favor of the normal chromosome X. Global transcriptional profiling of affected individuals and controls revealed significant alterations of genes and pathways in a pattern consistent with previous microarray studies of autism spectrum disorder patients. Moreover, expression analysis revealed copy number-dependent increased messenger RNA (mRNA) levels in affected patients compared to control individuals. A subset of differentially expressed genes was validated using qRT-PCR. Xq12-q13.3 duplication is a novel global developmental delay and autism-predisposing chromosomal aberration; pathogenesis of which may be mediated by increased dosage of genes contained in the duplication, including NLGN3, OPHN1, AR, EFNB1, TAF1, GJB1, and MED12.

Identification of PML-RARalpha rearrangement by RT-PCR without t(15;17) on G-banding and FISH in acute promyelocytic leukaemia: a case study and review of the literature

Identification of PML-RARalpha rearrangement by RT-PCR without t(15;17) on G-banding and FISH in acute promyelocytic leukaemia: a case study and review of the literature

Cytogenetics studies of 468 patients with primary amenorrhea

Objective To analyze the relationship between karyotypes and clinic features of patients with primary amenorrhea. Method Karyotype analysis of patients with primary amenorrhea was performed by using G-banding technique. Results Karyotype analysis of 468 patients with primary amenorrhea revealed that 255 patients (54. 49% ) had normal female karyotypes and 213 patients (45.51%) had abnormal karyotypes, including 143 patients with abnormal X chromosome, 4 patients with mosaic X -Y chromosome, 57 patients with 46, XY karyotype, 8 patients with abnormal autosome and one patient with Xautosome translocation. 75.52% primary amenorrhea patients with short stature had abnormal X chromosome, and all primary amenorrhea patients with deletion or break-up of Xp11. 1 - 11.4 and Xp21 - 22 were short statures. Conclusion One of the main reasons of primary amenorrhea was chromosome abnormity,especial heterosome abnormity. Karyotype analysis should be used to detect primary amenorrhea patients in regular. There might be relationship between height improvement and the abnormity of Xp11. 1 - 11.4 and Xp21 - 22. Key words: Amenorrhea/GE; Karyotyping; Phenotype

Laboratory diagnosis of acute myeloid leukemia with minimal differentiation switching to mixed phenotype acute leukemia: one case report

Objective To analyze the diagnostic process of a rare case of acute myeloid leukemia with minimal differentiation undergoing a lineage switch to mixed phenotype acute leukemia, NOS-rare types,and to investigate its difference from other acute myeloid leukemia and mixed phenotype acute leukemia. Methods Following tests were performed on the patient with switched mixed phenotype acute leukemia and three control leukemia patients ( including two acute myeloid leukemia with minimal differentiation and one mixed phenotype acute leukemia ). Cell morphology was analyzed by bone marrow smear and related cell chemical staining. Immunophenotyping of bone marrow was performed by flow cytometry ( FCM ). G-banding technique was used for karyotype analysis and RT-PCR was used for fusion gene detection. All the laboratory data of the switched patient were compared to that of three control patients in order to reveal the characteristics of such a rare phenotype switch in acute leukemia. Results Before switching, the morphology of acute myeloid leukemia with minimal differentiation demonstrated 0.82 blasts occurring in bone marrow, distinct nucleoli and absence of Auer rods. Blast cells expressed hematopoieticassociated antigens ( CD38, HLA-DR ), myeloid antigens ( CD13, CD56, CD11b ) and CD7. And these blasts were negative for MPO, CD33, CD15, CD79, CD19, CD22, cytoplasmic CD3, CD4 and CD8. After switching, 0. 42 blasts were found in bone marrow, showed eosinophilia and presence of basophile. Blast cells expressed hematopoietic-associated antigens ( CD38, HLA-DR ), myeloid antigens ( MPO, CD13 ),lymphoid antigens ( CD19, CD79a ,cytoplasmic CD3, and CD7 ). The control group showed typical morphology and immunophenotyping. No abnormal karyotype and fusion gene were detected. Conclusions It is a rare and complicated case that acute myeloid leukemia with minimal differentiation switched to mixed phenotype acute leukemia, NOS-rare types. The laboratory features, especially the change of immunophenotyping play an important role in the diagnosis. Key words: Leukemia,myeloid,acute; Leukemia,biphenotypic,acute; Immunophenotyping; Flow cytometry

Miscarriage karyotype and its relationship with maternal body mass index, age, and mode of conception

This study investigated the association between miscarriage karyotype and body mass index, maternal age, and mode of conception. Miscarriages after IVF and/or intracytoplasmic sperm injection were less frequently aneuploid; advanced maternal age was associated with an increase in aneuploid products of conception; overweight and obese women aged <35 years were less likely to have aneuploid miscarriages than women in a healthy weight range, suggesting alternate mechanisms for miscarriage in this population.

Application of array-based comparative genomic hybridization in primary amenorrhea women

Objective To explore the molecular mechanisms of primary amenorrhea by using arrayCGH technology. Methods Ten patients with primary amenorrhea and 10 female volunteers with regular menstrual cycles as healthy controls were selected. All patients and control samples were analyzed by conventional chromosome analysis (G-banding technology) and array-CGH technology, respectively. ArrayCGH was performed using Affymetrix Cytogenetic 2. 7M arrays following the manufacturer's standard protocol. Results Both the patient group and control group analyzed by conventional G-banding karyotype technology showed a negative result with a normal female karyotype: 46, XX. The result of array-CGH analysis demonstrated a microdeletion of approximately 110 000 bp located at the end of the short arm of X chromosome [46, X, del (X) (p22. 33 )] were identified in 5 patients, which was not detected in the control group. All healthy control samples by array-CGH analysis showed no pathological DNA copy number variation. Conclusions Array-CGH technology can improve the diagnosis rate of chromosomal disease at the DNA level. It is necessary to provide array-CGH for higher resolution genetic analysis of idiopathic primary amenorrhea patient who can not be identified by conventional technology. Key words: Amenorrhea; Microarray analysis; Nucleic acid hybridization

Karyomorphological and biochemical studies in Glebionis coronaria (L.) Spach and Glebionis segetum (L.) Fourreau from Italy

The karyotype structures and the composition and distribution of the heterochromatin in Glebionis coronaria and Glebionis segetum using Giemsa and fluorescent banding techniques were analysed. The species studied are diploids with 2n = 2x = 18 chromosomes. G. coronaria possesses the most symmetrical karyotype, comprising mainly metacentric chromosomes. G. segetum, with formula 12 m + 6 sm, showed a slightly less symmetrical karyotype. Giemsa C‐banding revealed little constitutive heterochromatin in both species. The presence of telomeric bands was restricted to two chromosome pairs. After staining with chromomycin A3, the chromosomes of G. coronaria and G. segetum revealed bright fluorescence at the telomeric regions of two chromosomes. No 4‐6‐diamidino‐2‐phenyl‐indole bright blocks were observed. To estimate genetic variability in the species under consideration, genotypic expression was also determined through isozyme electrophoresis of cytosolic ascorbate peroxidase, considered a genetic marker in the study of plant phylogenetics. The relationships between G. coronaria and G. segetum are discussed. Abbreviations: ASC, ascorbate; BSA, bovine serum albumin; cAPX, cytosolic ascorbate peroxidase; CMA, chromomycin A3; DAPI, 4‐6‐diamidino‐2‐phenyl‐indole; EDTA, ethylenediaminetetraacetic acid; PAGE, polyacrylamide gel electrophoresis

The evolution of gene regulation, the RNA universe, and the vexed questions of artefact and noise

In the ten years since the first sequencing of the human genome, much has been made of the need to look to gene regulation, and not gene number or DNA sequence, for the evolution of organismal diversity and complexity - an issue that rose to prominence, with the realization first, that the number of human genes is about the same as the number required to specify a nematode worm; and second, that the DNA of H. sapiens is roughly 96% identical to that of the chimpanzee. But the realization that the secret of evolution lies in changes in gene regulation considerably predates the revela tions of genomics. Allan Wilson and colleagues, in a paper published in 1974 [1], drew attention to the simple and striking fact that morphologically homogeneous frog species also have relatively homogeneous karyo types, whereas mammalian species, which are marked ly diverse morphologically, also show major differ ences in chromosome number and organization; changes in proteins, by contrast, are much the same for both groups. They concluded that genome organization, and by implication gene regulation, is more important for metazoan evolution than protein sequence (and cite earlier publications of EB Ford and Susumu Ohno for the same insight). The following year, Mary-Claire King and Wilson published a more detailed examination of the chromosomal distinctions between human and chimpanzee [2], arguing compellingly, without benefit of highthroughput anything, that changes in the organization of the genome, and not changes in protein-coding sequence, must account for the crucial differences between the two primates. In those pre-genomic days, the protein data were in large part immunological and electrophoretic; the analysis of genome reorganization depended on chromosome banding patterns (Giemsa banding, not FISH); and almost nothing was known of the mechanism of gene regulation in eukaryotes. The ground between then and now is covered in a recent review by Sean Carroll [3], who acknowledges Emile Zuckerkandl and Eric Davidson as early proponents of the importance of gene regulation in morphological evolution and charts the remarkable history of the development of ideas consequent on the discovery of the homeobox genes, with a strong emphasis on the evolution of cis-regulatory elements - that is to say, DNA binding sites for gene regulatory proteins - as the basis for morphological change. The argument is that DNA regulatory elements and the proteins that bind to them, often combinatorially, constitute regulatory networks that can evolve rapidly through changes to the regulatory elements, which are often modular, different modules binding different proteins characteristic of distinct differentiated states of a cell. The gene regulatory proteins can also change, of course, but are generally more highly conserved than their binding sites. Tuch et al. [4] have published a short and pellucid overview of the essential points and principles of this schema, in the context of recent evidence on how such regulatory circuits can become rewired in yeast. Beyond regulatory proteins

The clinical and experimental analyses of a multiple myeloma patient with derivative der(Y;1)

To investigate the clinical significance of a rare chromosome abnormality der(Y)t(Y;1) in a patient with multiple myeloma (MM). The chromosome spread was prepared after 24 h culture of bone marrow. G-banding technique was used to analyze the karyotype. Fluorescence in situ hybridization (FISH) was performed to ascertain the origin of abnormal chromosome detected by conventional karyotypic analysis. Flow cytometry was used to detect the expression of the CD38/CD138/ZAP70. Immunoelectrophore was applied to identify the type of immunoglobulin. A complex pattern of chromosome rearrangement was observed: 92,XXYY[3]/49,X,der(Y)t(Y;1)(q12;q21),t(11;14)(q13;q32),+18,+20,+21[47]/49,X,idem,del(13q22),ace[1]/98,XX,der(Y)t(Y;1) x 2,+18,+18,+20,+20,+21,+21[10]/46,XY[19]. The result was confirmed by metaphase-FISH. The type of immunoglobulin was IgD with the level of 6.24g/L. The CD38/CD138 was positive but ZAP70 was negative. Structural abnormality of chromosome Y is rare in blood malignancy. Most of them were described in myelodysplastic syndrome or myeloproliferative disorders. It is the first report of der(Y)t(Y;1) abnormality in multiple myeloma.

Comprehensive 5-Year Study of Cytogenetic Aberrations in 668 Infertile Men

The causes of male infertility are heterogeneous but more than 50% of cases have a genetic basis. Specific genetic defects have been identified in less than 20% of infertile males and, thus, most causes remain to be elucidated. The most common cytogenetic defects associated with nonobstructive azoospermia are numerical and structural chromosome abnormalities, including Klinefelter syndrome (47,XXY) and Y chromosome microdeletions. To refine the incidence and nature of chromosomal aberrations in males with infertility we reviewed cytogenetic results in 668 infertile men with oligozoospermia and azoospermia. High resolution Giemsa banding chromosome analysis and/or fluorescence in situ hybridization were done in 668 infertile males referred for routine cytogenetic analysis between January 2004 and March 2009. The overall incidence of chromosomal abnormalities was about 8.2%. Of the 55 patients with abnormal cytogenetic findings sex chromosome aneuploidies were observed in 29 (53%), including Klinefelter syndrome in 27 (49%). Structural chromosome abnormalities involving autosomes (29%) and sex chromosomes (18%) were detected in 26 infertile men. Abnormal cytogenetic findings were observed in 35 of 264 patients (13.3%) with azoospermia and 19 of 365 (5.2%) with oligozoospermia. Structural chromosomal defects and low level sex chromosome mosaicism are common in oligozoospermia cases. Extensive cytogenetic assessment and fluorescence in situ hybridization may improve the detection rate in males with oligozoospermia. These findings highlight the need for efficient genetic testing in infertile men so that couples may make informed decisions on assisted reproductive technologies to achieve parenthood.

A clinical and laboratory study on acute myeloid leukemia with t(6;9)(p23;q34)

To explore the clinical and laboratory features of 6 cases of acute myeloid leukemia (AML) with t(6;9)(p23;q34). Chromosome preparation of bone marrow cells was performed with regular method. R-banding by heating using Giemsa banding technique (RHG) was used for karyotype analysis. The immunoprofile was studied by flow cytometry (FCM) using a panel of monoclonal antibodies. Chromosome painting was performed by using whole chromosome paint probes for chromosomes 6 and 9 in all the 6 cases. The expression of fusion gene DEK/CAN and FLT3-ITD mutation were analyzed by reverse transcription-PCR(RT-PCR). The t(6;9)(p23;q34) was found in all the 6 cases including 4 cases of M2 and 2 cases of M4. Blast cells were positive for CD13 and CD33 in 6 patients, for HLA-DR in 4 patients, for CD34 and CD117 in 3 cases, for CD38 or CD15 each in 1 case, respectively. A reciprocal translocation between chromosome 6 and 9 was confirmed by chromosome painting technique in the 6 cases. The DEK/CAN fusion gene was found in all the 6 cases, FLT3-ITD mutation was detected in three of them. Follow-up showed that 3 patients died with a survival time of 3 months, 5 months and 6 months, respectively. The other three obtained complete remission and are still alive. The t(6;9)(p23;q34) is a rare recurrent abnormity. AML with t(6;9)(p23;q34) has unique clinical and laboratory features and its prognosis is poor in most cases.

Automating Giemsa banding of chromosomes: protocol for and evaluation of the use of a programmable, high-throughput automatic stainer

The use of prometaphase chromosomes prepared for high-resolution imaging is essential for accurate cytogenetic investigations. The process of Giemsa chromosome banding (G-banding), however, is often time consuming and difficult to standardize owing to differing ambient conditions and inter-operator variability. Consequently, many laboratories currently are introducing automatic metaphase finder and analysis systems to achieve the goals of higher throughput of samples and more consistent chromosome quality. In this context, we investigated the use of automation to improve the G-banding process. We investigated the use of the Shandon Thermo Varistain Gemini automatic stainer to replicate the manual process of G-banding. We compared the current manual method and the automated procedure and found that automation provided equivalent quality, and increased consistency while decreasing the time required and reducing the cost per preparation.