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1.
Anticancer Res ; 39(8): 4329-4332, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31366525

RESUMO

BACKGROUND/AIM: Acute myeloid leukemia is well characterized by chromosomal aberrations that correspond to various subtypes of acute leukemias. The t(8;21)(q22;q22) is a frequent chromosomal abnormality strongly associated with acute myeloblastic leukemia with maturation (AML-M2), but is rarely associated with other subtypes. Translocation involving a third chromosome could produce new genetic rearrangements that lead to leukemogenesis. PATIENTS AND METHODS: Conventional cytogenetic analysis and fluorescence in situ hybridization (FISH) were performed to identify the karyotype. Reverse transcriptase polymerase chain reaction (RT-PCR) was used to detect the AML1/ETO transcript. RESULTS/CONCLUSION: We herein report a novel rearrangement with a three-way translocation involving chromosomes 8, 21 and another unknown chromosome, in an 83-year-old female patient diagnosed as AML-M4, with an ALM1/ETO negative transcript. This is an uncommon case of AML-M4 with three-way translocation in a new variant of t(8;21) acute myeloid leukaemia. The detailed mechanism of different phenotype expression is unclear. Further study is needed to identify the leukemogenetic transformation resulting from t(8;21) translocation.


Assuntos
Análise Citogenética , Cariótipo , Leucemia Mielomonocítica Aguda/genética , Translocação Genética/genética , Idoso de 80 Anos ou mais , Cromossomos Humanos Par 21/genética , Cromossomos Humanos Par 8/genética , Feminino , Humanos , Hibridização in Situ Fluorescente/métodos , Cariotipagem/métodos , Leucemia Mielomonocítica Aguda/diagnóstico , Leucemia Mielomonocítica Aguda/patologia
2.
Cytogenet Genome Res ; 158(2): 63-73, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31261151

RESUMO

Terminal deletion of chromosome 4 (4q deletion syndrome) is a rare genetic condition that is characterized by a broad clinical spectrum and phenotypic variability. Diagnosis of the distinct condition can be identified by conventional chromosome analysis and small deletions by novel molecular cytogenetic methods such as microarray comparative genome hybridization (aCGH). Prenatal diagnosis is challenging; to date 10 cases have been described. We report a prenatally diagnosed case of de novo 4q deletion syndrome confirmed by conventional karyotyping and FISH due to an elevated combined risk for Down syndrome and prenatal ultrasound findings. aCGH validated the diagnosis and offered exact characterization of the disorder. Cytogenetic and microarray results described a 4q32.1qter terminal deletion of the fetus. Prenatal ultrasound detected multiple nonstructural findings (micrognathia, choroid plexus cysts, echogenic fetal bowel, short femur, and cardiac axis deviation). Pregnancy was terminated at 20 weeks. In addition to the index patient, we reviewed the 10 prenatally published cases of 4q deletion syndrome in the literature and compared these with our results. We summarize the patients' characteristics and prenatal clinical findings. Alterations of maternal serum biochemical factors, an elevated combined risk for trisomies, and distinct ultrasonographic findings can often be observed in cases of prenatal 4q deletion syndrome and may facilitate the otherwise difficult prenatal diagnosis.


Assuntos
Transtornos Cromossômicos/diagnóstico , Hibridização in Situ Fluorescente/métodos , Cariotipagem/métodos , Diagnóstico Pré-Natal/métodos , Aborto Induzido , Adulto , Deleção Cromossômica , Transtornos Cromossômicos/genética , Cromossomos Humanos Par 4/genética , Hibridização Genômica Comparativa , Feminino , Humanos , Idade Materna , Fenótipo , Gravidez , Fatores de Risco
3.
Methods Mol Biol ; 1975: 407-426, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31062320

RESUMO

Due to their unique cellular features, pluripotent stem cells (PSCs) acquire chromosomal aberrations at a rather high frequency during their growth in culture. Analysis of chromosomal integrity should be routinely performed and usually is done at the DNA level of the cells. RNA sequencing (RNA-Seq) has recently become the basic tool for transcriptional studies. Therefore, methods that utilize this already available data to inspect the genomic integrity are very valuable. In this chapter, we provide a practical guide to implement methods of detection of chromosomal aberrations, which are based on RNA-Seq data. The expression-based karyotyping (e-Karyotyping) method is based on global gene expression analysis, while the expressed-SNP-karyotyping (eSNP-Karyotyping) method is based on changes in the ratio between alleles.


Assuntos
Aneuploidia , Biologia Computacional/métodos , Regulação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Cariotipagem/métodos , Células-Tronco Pluripotentes/citologia , Polimorfismo de Nucleotídeo Único , Humanos
4.
Fertil Steril ; 111(5): 842-850, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31029238

RESUMO

Male infertility is a heterogenous disease process requiring the proper functioning and interaction of thousands of genes. Given the number of genes involved, it is thought that genetic causes contribute to most cases of infertility. Identifying these causes, however, is challenging. Infertility is associated with negative health outcomes, such as cancer, highlighting the need to further understand the genetic underpinnings of this condition. This paper describes the genetic and genomic tests currently available to identify the etiology of male infertility and then will discuss emerging technologies that may facilitate diagnosis and treatment of in the future.


Assuntos
Testes Genéticos/métodos , Infertilidade Masculina/genética , Transtornos do Cromossomo Sexual no Desenvolvimento Sexual/genética , Deleção Cromossômica , Cromossomos Humanos Y/genética , Testes Genéticos/tendências , Humanos , Infertilidade Masculina/diagnóstico , Cariotipagem/métodos , Cariotipagem/tendências , Masculino , Análise Serial de Proteínas/métodos , Análise Serial de Proteínas/tendências , Aberrações dos Cromossomos Sexuais , Transtornos do Cromossomo Sexual no Desenvolvimento Sexual/diagnóstico
6.
Cytogenet Genome Res ; 157(1-2): 46-52, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30904910

RESUMO

Chromosome homologies in reptiles have been investigated extensively by gene mapping and chromosome painting. Relative chromosome size can be estimated roughly from conventional karyotypes, but chromosome GC content cannot be evaluated by any of these approaches. However, GC content can be obtained by whole-genome sequencing, although complete data are available only for a limited number of reptilian species. Chromosomes can be characterized by size and GC content in bivariate flow karyotypes, in which the distribution of peaks represents the differences. We have analysed flow karyotypes from 9 representative squamate species and show chromosome profiles for each species based on the relationship between size and GC content. Our results reveal that the GC content of macrochromosomes is invariable in the 9 species. A higher GC content was found in microchromosomes, similar to profiles previously determined in crocodile, turtle, and chicken. The findings suggest that karyotype evolution in reptiles is characterized by unique features of chromosome GC content.


Assuntos
Composição de Bases/genética , Cromossomos/genética , Cariotipagem/métodos , Répteis/genética , Animais , Evolução Molecular , Tamanho do Genoma , Filogenia , Répteis/classificação , Especificidade da Espécie , Sequenciamento Completo do Genoma/métodos
7.
Transl Psychiatry ; 9(1): 60, 2019 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-30718465

RESUMO

We searched for genetic causes of major psychiatric disorders (bipolar disorder, schizoaffective disorder, and schizophrenia) in a large, densely affected pedigree from Northern Sweden that originated with three pairs of founders born around 1650. We applied a systematic genomic approach to the pedigree via karyotyping (N = 9), genome-wide SNP arrays (N = 418), whole-exome sequencing (N = 26), and whole-genome sequencing (N = 10). Comprehensive analysis did not identify plausible variants of strong effect. Rather, pedigree cases had significantly higher genetic risk scores compared to pedigree and community controls.


Assuntos
Transtorno Bipolar/genética , Genômica/métodos , Transtornos Psicóticos/genética , Esquizofrenia/genética , Adulto , Idoso , Transtorno Bipolar/epidemiologia , Estudos de Casos e Controles , Feminino , Humanos , Cariotipagem/métodos , Masculino , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Linhagem , Polimorfismo de Nucleotídeo Único , Transtornos Psicóticos/epidemiologia , Esquizofrenia/epidemiologia , Suécia/epidemiologia , Sequenciamento Completo do Exoma/métodos , Sequenciamento Completo do Genoma/métodos
8.
Cytogenet Genome Res ; 157(1-2): 98-106, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30754040

RESUMO

The suborder Serpentes is divided into 2 infraorders, Scolecophidia and Alethinophidia, which diverged at an early stage of snake diversification. In this study, we examined karyotypes of 4 scolecophidian species (Letheobia simonii, Xerotyphlops vermicularis, Indotyphlops braminus, and Myriopholis macrorhyncha) and performed FISH with 18S-28S rDNA as well as microchromosomal and Z chromosome-linked genes of Elaphe quadrivirgata (Alethinophidia) to investigate the karyotype evolution in the scolecophidian lineage. Diploid chromosome numbers of X. vermicularis and L. simonii were 30 (16 macrochromosomes and 14 microchromosomes) and 32 (16 macrochromosomes and 16 microchromosomes), respectively. The karyotype of a female M. macrorhyncha consisted of 15 macrochromosomes and 19 microchromosomes, including a heterochromatic microchromosome, indicating the presence of a heteromorphic chromosome pair. E. quadrivirgata Z-linked genes mapped to chromosome 4 of M. macrorhyncha, not to the heteromorphic pair. Therefore, M. macrorhyncha may have differentiated ZW sex chromosomes which are not homologous to those of E. quadrivirgata. One of the E. quadrivirgata microchromosomal genes mapped to the terminal region of chromosome 4q in X. vermicularis, suggesting that fusions between microchromosomes and macrochromosomes occurred in this species. rDNA was localized in different macrochromosomal pairs in the 2 diploid scolecophidian snakes examined here, whereas the gene location in a microchromosomal pair was conserved in 5 alethinophidian species examined. These results might imply the occurrence of chromosome fusions in the scolecophidian lineages. In I. braminus, a unique parthenogenetic snake with a triploid karyotype (21 macrochromosomes and 21 microchromosomes), morphological heteromorphisms were identified in chromosomes 1 and 7. Such heteromorphisms in 2 chromosomes were also observed in individuals from distant locations in the broad distribution range of this species, suggesting that the heteromorphisms were fixed in the genome at an early stage of its speciation.


Assuntos
Cromossomos/genética , Cariotipagem/métodos , Cromossomos Sexuais/genética , Serpentes/genética , Animais , Mapeamento Cromossômico , Evolução Molecular , Feminino , Hibridização in Situ Fluorescente/métodos , Cariótipo , Masculino , Serpentes/classificação , Especificidade da Espécie
9.
Mol Biol Rep ; 46(2): 1931-1940, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30710232

RESUMO

The aim of the study was to characterize the type of aneuploidy present in the hybrid Urochloa ruziziensis × Urochloa decumbens and to confirm the origin of the additional chromosomes through comparative analysis of the hybrid and parental karyotypes. C and CMA banding techniques were used for chromosome differentiation. The parental genotypes showed 36 chromosomes. The hybrid presented plants with 36 + 2 chromosomes and plants with 36 + 1 chromosomes. Urochloa ruziziensis (4x) presented four chromosomes with CMA and C bands co-located in the terminal position. In U. decumbens, four chromosomes presented terminal CMA bands, eight chromosomes were distinguished by C banding with pericentromeric and terminal bands, one chromosome with terminal band at both ends and one chromosome presented one C terminal band. For the hybrid, CMA bands were found on five chromosomes and C bands on seven chromosomes, all in terminal position. Aneuploidy was identified in pairs 3' and 4' in the hybrid plants with 36 + 2 chromosomes, characterizing it as double trisomy. The karyotype of hybrid plants with 36 + 1 chromosomes indicated elimination of the additional chromosome identified in pair 4' and maintenance of trisomy on pair 3'. The comparative analysis of karyotypes indicates that the additional chromosomes that characterize the trisomy were inherited from U. ruziziensis (artificial tetraploid).


Assuntos
Quimera/genética , Poaceae/genética , Aneuploidia , Cromossomos de Plantas/genética , Transferência Genética Horizontal/genética , Genótipo , Cariotipagem/métodos , Plantas/genética , Poaceae/metabolismo , Poliploidia , Trissomia/genética
10.
Medicine (Baltimore) ; 98(7): e14599, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30762813

RESUMO

BACKGROUND: Prenatal screening for aneuploidies has seen great changes over the last 2 decades. But there is still no non-invasive diagnostic test. Therefore, prenatal invasive procedures are still being routinely performed due to maternal anxiety. The association of cardiac anomalies and abnormal findings with aneuploidies has been known for a long time. This prospective study was done to evaluate abnormal fetal cardiac examination (FCE) findings on patients undergoing diagnostic invasive procedures due to maternal anxiety and to assess the predictive value of abnormal cardiac findings on abnormal karyotype. MATERIALS AND METHODS: Patients who underwent prenatal diagnostic invasive tests due to maternal anxiety indication between March 2013 and September 2016 were included in this study. FCE was performed in the study group immediately prior to invasive tests. Findings of fetal cardiac examination are classified as normal, major-minor cardiac anomalies and soft markers. Fetal karyotypes were compared among groups depending on cardiac findings. RESULTS: One hundred eighty-two invasive procedures were performed because of maternal anxiety during this period. There were 29 abnormal findings detected on FCE. A total of 7 abnormal karyotypes were detected. FCE was abnormal in 5 of the abnormal karyotypes (71.4%). The presence of a major cardiac anomaly was most predictive for abnormal karyotype (LR+: 96,67, LR-: 0,34). No association was detected between the presence of minor cardiac anomalies and abnormal karyotype. Normal FCE appeared to be a good predictive factor for normal karyotype (LR-: 0.20). CONCLUSIONS: This is the first study evaluating the power of early fetal cardiac examination findings on fetal aneuploidies. This study suggested that the application of fetal cardiac examination findings to genetic counseling for screening aneuploidies may be efficient on patients' preference about invasive tests. Due to the small number of abnormal findings and karyotypes detected (not the large study group), further studies on large study groups are needed to confirm these results.


Assuntos
Cardiopatias Congênitas/diagnóstico , Mães/psicologia , Preferência do Paciente , Diagnóstico Pré-Natal/métodos , Diagnóstico Pré-Natal/psicologia , Adolescente , Adulto , Aneuploidia , Feminino , Humanos , Cariotipagem/métodos , Pessoa de Meia-Idade , Gravidez , Estudos Prospectivos , Adulto Jovem
11.
Braz J Med Biol Res ; 52(2): e8194, 2019 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-30785480

RESUMO

Cytogenetics is essential in myeloid neoplasms (MN) and pre-analytical variables are important for karyotyping. We assessed the relationship between pre-analytical variables (time from collection to sample processing, material type, sample cellularity, and diagnosis) and failures of karyotyping. Bone marrow (BM, n=352) and peripheral blood (PB, n=69) samples were analyzed from acute myeloid leukemia (n=113), myelodysplastic syndromes (n=73), myelodysplastic syndromes/myeloproliferative neoplasms (n=17), myeloproliferative neoplasms (n=137), and other with conclusive diagnosis (n=6), and reactive disorders/no conclusive diagnosis (n=75). The rate of unsuccessful karyotyping was 18.5% and was associated with the use of PB and a low number of nucleated cells (≤7×103/µL) in the sample. High and low cellularity in BM and high and low cellularity in PB samples showed no metaphases in 3.9, 39.7, 41.9, and 84.6% of cases, respectively. Collecting a good BM sample is the key for the success of karyotyping in MN and avoids the use of expensive molecular techniques.


Assuntos
Células da Medula Óssea/patologia , Cariotipagem/métodos , Leucemia Mieloide/genética , Síndromes Mielodisplásicas/genética , Transtornos Mieloproliferativos/genética , Manejo de Espécimes/métodos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Leucemia Mieloide/diagnóstico , Masculino , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/diagnóstico , Transtornos Mieloproliferativos/diagnóstico , Manejo de Espécimes/normas , Adulto Jovem
12.
Taiwan J Obstet Gynecol ; 58(1): 15-28, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30638470

RESUMO

The aim of the current review is to report a-CGH abnormalities identified in fetuses with prenatally diagnosed fetal malformations in whom a normal karyotype was diagnosed with conventional cytogenetic analysis. A systematic electronic search of databases (PubMed/Medline, EMBASE/SCOPUS) has been conducted from inception to May, 2017. Bibliographic analysis has been performed according to PRISMA statement for review. The following keywords were used: 'array-CGH' and 'fetal malformations" and "prenatal diagnosis"; alternatively, "microarray", "oligonucleotide array", "molecular biology", "antenatal diagnostics", "fetal diagnostics", "congenital malformations" and "ultrasound" were used to capture both "a-CGH" and "prenatal". One-hundred and twelve fetuses with prenatally diagnosed fetal malformations with normal karyotyping and a-CGH abnormalities detected are described. Single or multiple microarray abnormalities diagnosed have been classified in relation to different organ/system affected. The most frequent a-CGH abnormalities were detected in cases of congenital heart diseases (CDHs), multiple malformations and central nervous system (CNS) malformations. Maternal or paternal carrier-state was seen in 19.64% (22/112), of cases while the number of reported de novo mutations accounted for 46.42% (52/112) of all CNVs microarray abnormalities. Array-comparative genomic hydridization (a-CGH) may become an integral and complemantary genetic testing when fetal malformations are detected prenatally in fetuses with normal cytogenetic karyotype. In addition, a-CGH enables the identification of CNVs and VOUS and improves the calculation of recurrent risk and the genetic counseling.


Assuntos
Anormalidades Múltiplas/genética , Aberrações Cromossômicas/embriologia , Hibridização Genômica Comparativa , Cardiopatias Congênitas/genética , Análise de Sequência com Séries de Oligonucleotídeos , Diagnóstico Pré-Natal/métodos , Anormalidades Múltiplas/diagnóstico , Sistema Nervoso Central/anormalidades , Sistema Nervoso Central/embriologia , Feminino , Cardiopatias Congênitas/diagnóstico , Humanos , Cariotipagem/métodos , Idade Materna , Gravidez , Fatores de Risco
13.
Asian Pac J Cancer Prev ; 20(1): 235-241, 2019 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-30678438

RESUMO

Objective: Multiple myeloma (MM) is a clinically and genetically heterogeneous plasma cell neoplasm. The prognosis of MM patients is dependent on several factors including the patient's age, the stage of disease and genetic alterations. This study aimed to determine the frequency of common chromosomal abnormalities and their significance in MM patients referred to a tertiary healthcare center in India. Methods: Fluorescence in situ hybridization on interphase nuclei from bone marrow cells using seven MM-specific probes for recurrent aberrations was performed in a total of 215 newly diagnosed patients. Results: Chromosomal abnormalities were detected in 161 (74.9%) MM patients in this study. The most frequent aberration was trisomy(ies) involving only gain of chromosomes in 48 (22.3%) cases. A translocation involving the IGH gene alone or accompanied by trisomy(ies) or by monosomy 13/13q deletion or by both was registered in 80 (37.2%) patients. Atypical patterns such as a deletion of the IGH variable segment (IGHv) on the derivative chromosome 14 or on the native (normal) chromosome 14, biallelic deletion of IGHv, deletion of the IGH constant segment on the rearranged chromosome14 and extra fusions were noticed in 21 (9.8%) patients with an IGH rearrangement. Monosomy 13/deletion 13q was identified singly or as part of a complex karyotype in 74 patients (34.4%). Clonal heterogeneity and additional abnormalities including TP53 deletion and monosomies of chromosomes 4, 9, 14 and 16 were recorded in 18.6% and 16.3% of patients respectively. Patients with abnormalities exhibited plasmacytosis, reduced hemoglobin value and high level of ß2-microglobulin. Conclusions: A lower median age and a low frequency of IGH translocations particularly t(11;14) and chromosome 13 abnormalities suggest ethnic diversity. Further investigations on genetic alterations including IGH deletions will contribute to improved insights into the biology of myeloma disease, risk stratification and patient management.


Assuntos
Cromossomos/genética , Mieloma Múltiplo/genética , Idoso , Aberrações Cromossômicas , Deleção Cromossômica , Transtornos Cromossômicos/genética , Cromossomos Humanos Par 13/genética , Feminino , Humanos , Índia , Cariotipagem/métodos , Masculino , Pessoa de Meia-Idade , Prognóstico , Atenção Terciária à Saúde/métodos , Translocação Genética/genética
14.
PLoS One ; 14(1): e0210079, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30608972

RESUMO

Previous studies, including our own, have reported that spermatozoa isolated from the testis have remarkably higher occurrence of aneuploidy once isolated from azoospermic men. This notion, however, did not translate into a lower pregnancy rate nor a greater proportion of miscarriages. Indeed, ICSI offspring generated from surgically retrieved gametes did not suffer from increased karyotypic aneuploidy than children generated from ejaculated specimens. In recent years, aneuploidy assessments on a larger number of cells and utilizing more chromosome probes have reported a progressive decrease in chromosomal aberrations in spermatozoa directly retrieved from the seminiferous tubules. In light of the availability of more accurate molecular genetic techniques, we have decided to challenge the notion that sampling epididymal and testicular tissues yields spermatozoa with higher incidence of aneuploidy than those retrieved in the ejaculate. In a retrospective manner, we have carried out an analysis by FISH with 9 chromosome probes on at least 1000 cells from the ejaculates of 87 consenting men and the specimens of 6 azoospermic men, while spermatozoa of fertile donors were used as control. Aneuploidy by FISH yielded 0.9% for the donor control but rose in the study group to 3.6% in the ejaculated, 1.2% for the epididymal, and 1.1% for testicular spermatozoa. There were no differences in autosomal or gonosomal disomies, nor nullisomies. In this group, once the specimens of these men were used for ICSI, ejaculated spermatozoa yielded a 22% clinical pregnancy rate that resulted in 62.5% pregnancy loss. The surgically retrieved specimens yielded a 50% clinical pregnancy rate that progressed to term. To confirm our findings, in a prospective analysis, DNA sequencing was carried out on the ejaculates and surgical samples of 22 men with various spermatogenic characteristics. In this comparison, the findings were similar with actually a higher incidence of aneuploidy in the ejaculated spermatozoa (n = 16) compared to those surgically retrieved (n = 6) (P<0.0001). For this group, the clinical pregnancy rate for the ejaculated specimens was 47.2% with 29.4% pregnancy loss, while the surgically retrieved yielded a 50% clinical pregnancy rate, all progressing to term. A subsequent prospective combined assessment on ejaculated and surgically retrieved spermatozoa by FISH and NGS was performed on non-azoospermic men with high DNA fragmentation in their ejaculate. The assessment by FISH evidenced 2.8% chromosomal defects in the ejaculated and 1.2% in testicular biopsies while by NGS became 8.4% and 1.3% (P = 0.02), respectively. Interestingly, we evidenced a pregnancy rate of 0% with ejaculated while 100% with the testicular spermatozoa in this latter group. This indicates that improved techniques for assessing sperm aneuploidy on a wider number of cells disproves earlier reports and corroborates the safe utilization of testicular spermatozoa with a positive impact on chances of pregnancy.


Assuntos
Aneuploidia , Azoospermia/terapia , Análise do Sêmen/métodos , Recuperação Espermática , Aborto Espontâneo/epidemiologia , Adulto , Feminino , Humanos , Hibridização in Situ Fluorescente/métodos , Cariotipagem/métodos , Masculino , Pessoa de Meia-Idade , Gravidez , Taxa de Gravidez , Estudos Prospectivos , Estudos Retrospectivos , Injeções de Esperma Intracitoplásmicas/métodos , Espermatogênese/fisiologia , Espermatozoides/fisiologia , Resultado do Tratamento , Sequenciamento Completo do Exoma/métodos
15.
Methods Mol Biol ; 1881: 27-34, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30350195

RESUMO

Chromosome analysis of chronic lymphocytic leukemia (CLL) is an important clinical tool for evaluating prognosis and disease progression. Visualizing chromosomes microscopically using traditional cytogenetic techniques requires dividing cells to be arrested during metaphase. The major challenge for performing this analysis on CLL samples is stimulating the cells to divide in culture. Stimulation of CLL cells with CpG oligodeoxynucleotides has improved our ability to perform chromosome analysis for this leukemia. This protocol should help the reader successfully culture CLL samples for clinical chromosome analysis.


Assuntos
Leucemia Linfocítica Crônica de Células B/genética , Oligodesoxirribonucleotídeos/farmacologia , Cultura Primária de Células/métodos , Linfócitos B/efeitos dos fármacos , Células da Medula Óssea/efeitos dos fármacos , Humanos , Cariotipagem/instrumentação , Cariotipagem/métodos , Leucemia Linfocítica Crônica de Células B/sangue , Leucemia Linfocítica Crônica de Células B/patologia , Metáfase/efeitos dos fármacos , Mitógenos/farmacologia , Cultura Primária de Células/instrumentação , Manejo de Espécimes/instrumentação , Manejo de Espécimes/métodos , Células Tumorais Cultivadas
16.
Environ Mol Mutagen ; 60(4): 331-347, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30592088

RESUMO

To develop an improved in vitro mammalian cell gene mutation assay, it is imperative to address the known deficiencies associated with existing assays. Primary hepatocytes isolated from the MutaMouse are ideal for an in vitro gene mutation assay due to their metabolic competence, their "normal" karyotype (i.e., neither transformed nor immortalized), and the presence of the MutaMouse transgene for rapid and reliable mutation scoring. The cells were extensively characterized to confirm their utility. Freshly isolated cells were found to have a hepatocyte-like morphology, predominantly consisting of binucleated cells. These cells maintain hepatocyte-specific markers for up to 3 days in culture. Analyses revealed a normal murine hepatocyte karyotype with a modal ploidy number of 4n. Fluorescence in situ hybridization analysis confirmed the presence of the lambda shuttle vector on chromosome 3. The doubling time was determined to be 22.5 ± 3.3 h. Gene expression and enzymatic activity of key Phase I and Phase II metabolic enzymes were maintained for at least 8 and 24 h in culture, respectively. Exposure to ß-naphthoflavone led to approximately 900- and 9-fold increases in Cyp1a1 and Cyp1a2 gene expression, respectively, and approximately twofold induction in cytochrome P450 (CYP) 1A1/1A2 activity. Exposure to phenobarbital resulted in an approximately twofold increase in CYP 2B6 enzyme activity. Following this characterization, it is evident that MutaMouse primary hepatocytes have considerable promise for in vitro mutagenicity assessment. The performance of these cells in an in vitro gene mutation assay is assessed in Part II. Environ. Mol. Mutagen. 60:331-347, 2019. © 2018 The Authors. Environmental and Molecular Mutagenesis published by Wiley Periodicals, Inc. on behalf of Environmental Mutagen Society.


Assuntos
Separação Celular/métodos , Hepatócitos/efeitos dos fármacos , Testes de Mutagenicidade/métodos , Mutagênicos/toxicidade , Animais , Linhagem Celular , Células Cultivadas , Expressão Gênica/efeitos dos fármacos , Hepatócitos/citologia , Hepatócitos/metabolismo , Humanos , Hibridização in Situ Fluorescente/métodos , Cariótipo , Cariotipagem/métodos , Camundongos , Camundongos Endogâmicos BALB C , Mutagênese/efeitos dos fármacos
17.
Medicine (Baltimore) ; 97(50): e13617, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30558042

RESUMO

BACKGROUND: Congenital heart disease (CHD) is one of the most common birth defects; however, the mechanisms underlying its development are poorly understood. Recently, heritable genetic factors, including copy number variations (CNVs) and single nucleotide polymorphisms (SNPs), have been implicated in its etiology. The aim of this study was to investigate the utility of a SNP array for the prenatal diagnosis of CHD and the improvement of prenatal genetic counseling and to compare this approach to traditional chromosome analysis. METHODS: One hundred and fortysix cases of CHD detected by prenatal echocardiography were analyzed. Of these, 110 were isolated CHD and 36 were of CHD with extracardiac defects. SNP analysis was performed using the Affymetrix CytoScan HD platform, which was followed by karyotype analysis. All annotated CNVs were validated by fluorescence in situ hybridization. RESULTS: Karyotype analysis identified chromosomal abnormalities in 19 of 146 cases. In addition to the 15 chromosomal abnormalities that were consistent with the results of karyotype analysis, the SNP array identified abnormal CNVs in an additional 15.2% (22/145) cases; of these, 15 were pathogenic CNVs, three were variations of uncertain clinical significance, and four were benign CNVs. The rates at which the SNP array detected pathogenic CNVs differed significantly between cases of isolated CHD and CHD with extracardiac defects (13.6% vs. 72.2%, P = .001). The results of the SNP array also affected the rate of pregnancy termination. CONCLUSION: Combining SNP array with cytogenetic analyses is particularly effective for identifying chromosomal abnormalities in CNVs in fetuses with CHD, which also affects obstetrical outcomes.


Assuntos
Análise Citogenética/métodos , Cardiopatias Congênitas , Cariotipagem/métodos , Polimorfismo de Nucleotídeo Único/genética , Diagnóstico Pré-Natal/métodos , China/epidemiologia , Aberrações Cromossômicas , Variações do Número de Cópias de DNA/genética , Ecocardiografia/métodos , Feminino , Doenças Fetais/diagnóstico , Doenças Fetais/epidemiologia , Doenças Fetais/genética , Cardiopatias Congênitas/diagnóstico , Cardiopatias Congênitas/epidemiologia , Cardiopatias Congênitas/etiologia , Humanos , Gravidez , Resultado da Gravidez/epidemiologia , Cuidado Pré-Natal/métodos , Reprodutibilidade dos Testes
18.
Zhonghua Fu Chan Ke Za Zhi ; 53(7): 464-470, 2018 Jul 25.
Artigo em Chinês | MEDLINE | ID: mdl-30078256

RESUMO

Objective: To investigate the value of single nucleotide polymorphism array (SNP-array) for fetuses with abnormal ultrasound findings. Method: A total of 904 fetuses with abnormal ultrasound findings were enrolled in this study from May 2015 to November 2017, and 434 (48.0%) cases received conventional karyotyping analysis at the same time. According to different abnormal ultrasound category, 904 cases were divided into 5 groups: 280 cases (31.0%) in single system structural anomalies, 31 cases (3.4%) in multiple system structural anomalies, 331 cases (36.6%) in single ultrasound soft marker abnormalities without structural anomalies, 107 cases (11.8%) in multiple soft marker abnormalities and 155 cases (17.2%) in structural abnormalities combined with soft markers abnormalities. Abnormal detection rates by SNP-array among 5 groups of abnormal ultrasound category were calculated. Result: (1) Total SNP-array results: 171 (19.0%) cases out of 904 cases analyzed by SNP-array, presented chromosomal abnormalities. Pathogenic copy number variants were detected in 27 cases (3.0%) and variants of unknown significance were detected in 81 cases (7.8%) . In addition, 7 cases (26.0%) were found with new mutation by parental validation. (2) SNP-array of 5 groups: among the 5 groups of abnormal ultrasound category, chromosomal abnormalities were identified by SNP-array in 19.3% (54/280) with single system structural abnormalities, 25.8% (8/31) with multiple system structural abnormalities, 13.9% (46/331) with single nonstructural anomalies, 19.6% (21/107) with multiple nonstructural anomalies and 27.1% (42/155) with structural abnormalities combined with nonstructural anomalies. The differences were significant (P=0.010) . No chromosome abnormalities was identified in single soft marker abnormalities, such as choroid plexus cysts, echogenic foci in the heart, single umbilical artery and pyelectasis. (3) Chromosomal abnormalities: the abnormal detection rate of aneuploidy chromosomal abnormalities by SNP-array increased with the maternal age, decreased with the gestational weeks (all P<0.05) . However, the pathogenic copy number variants and variants of unknown significance rates did not change with maternal age and gestational weeks (all P>0.05) . (4) SNP-array and karyotyping: 434 cases were analyzed by conventional karyotyping and SNP-array respectively, 10.3% (43/419) of which presented chromosomal abnormalities by conventional karyotyping and 18.7% (81/434) of which presented chromosomal abnormalities by SNP-array. Conclusions: SNP-array could be a useful genetic analysis method in prenatal diagnosis for fetuses with abnormal ultrasound findings. For different abnormal ultrasound category, SNP-array has different detection rate. Compared with conventional karyotyping analysis, SNP-array can improve the detection rates for chromosomal abnormalities and find the chromosome abnormalities which can't be detected by conventional karyotyping analysis. In clinical prenatal genetic counseling, SNP-array should be selected rationally in combination with the various abnormal ultrasound category.


Assuntos
Anormalidades Múltiplas/genética , Transtornos Cromossômicos/genética , Anormalidades Congênitas/genética , Variações do Número de Cópias de DNA , Cariotipagem/métodos , Reação em Cadeia da Polimerase/métodos , Polimorfismo de Nucleotídeo Único , Diagnóstico Pré-Natal/métodos , Ultrassonografia Pré-Natal/métodos , Anormalidades Múltiplas/diagnóstico por imagem , Aneuploidia , Aberrações Cromossômicas , Anormalidades Congênitas/diagnóstico por imagem , Feminino , Feto , Aconselhamento Genético , Testes Genéticos , Humanos , Idade Materna , Gravidez
19.
Ann Clin Lab Sci ; 48(3): 264-272, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29970427

RESUMO

Conventional cytogenetic and routine I-FISH (interphase fluorescence in-situ hybridization) studies periodically present discrepant results on the same sample calling into question their validity. Generally it is expected that these tests confirm each other, otherwise there is concern that they may represent laboratory error. We present data showing that these discrepant results are rarely due to laboratory error, and that M-FISH (metaphase fluorescence in-situ hybridization) can usually reconcile them by identifying the nature of these differences. This report includes 32 bone marrow (BM) samples from patients with hematologic neoplasms that showed incongruent cytogenetic/I-FISH results. M-FISH was selectively applied for further clarification of these discrepancies when deemed necessary. This study evaluated BM samples in our laboratory (Integrated Oncology, Phoenix, AZ) that represented 5 major categories of hematologic disorders (MDS/AML, MPN, NHL, CLL, & PCN). Five general categories of these cases were identified: 1) laboratory error (clerical), 2) limited resolution of testing methods, 3) cellular response to culture/preparative conditions, 4) cytogenetic bi-clonality and 5) failed hybridizations due to cover-slipping. Our results suggest that the majority of discrepant results are related to the intrinsic nature of the malignant cells (and how they respond to their growth environment) evaluated by these two testing methods.


Assuntos
Medula Óssea/patologia , Neoplasias Hematológicas/genética , Hibridização in Situ Fluorescente/métodos , Interfase , Cariotipagem/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Medula Óssea/metabolismo , Estudos de Casos e Controles , Feminino , Seguimentos , Humanos , Masculino , Metáfase , Pessoa de Meia-Idade , Prognóstico
20.
Genes Genomics ; 40(5): 465-473, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29892954

RESUMO

Gender Dysphoria is characterized by a marked incongruence between the cerebral sex and biological sex. To investigate the possible influence of karyotype on the etiology of Gender Dysphoria we carried out the cytogenetic analysis of karyotypes in 444 male-to-females (MtFs) and 273 female-to-males (FtMs) that attended the Gender Identity Units of Barcelona and Málaga (Spain) between 2000 and 2016. The karyotypes from 23 subjects (18 MtFs and 5 FtMs) were also analysed by Affymetrix CytoScan™ high-density (HD) arrays. Our data showed a higher incidence of cytogenetic alterations in Gender Dysphoria (2.65%) than in the general population (0.53%) (p < 0.0001). When G-banding was performed, 11 MtFs (2.48%) and 8 FtMs (2.93%) showed a cytogenetic alteration. Specifically, Klinefelter syndrome frequency was significantly higher (1.13%) (p < 0.0001), however Turner syndrome was not represented in our sample (p < 0.61). At molecular level, HD microarray analysis revealed a 17q21.31 microduplication which encompasses the gene KANSL1 (MIM612452) in 5 out of 18 MtFs and 2 out of 5 FtMs that corresponds to a copy-number variation region in chromosome 17q21.31. In conclusion, we confirm a significantly high frequency of aneuploidy, specifically Klinefelter syndrome and we identified in 7 out of 23 GD individuals the same microduplication of 572 Kb which encompasses the KANSL1 gene.


Assuntos
Disforia de Gênero/etiologia , Disforia de Gênero/genética , Cariotipagem/métodos , Adulto , Bandeamento Cromossômico/métodos , Cromossomos Humanos Par 17/genética , Feminino , Identidade de Gênero , Duplicação Gênica/genética , Humanos , Cariótipo , Síndrome de Klinefelter , Masculino , Pessoa de Meia-Idade , Proteínas Nucleares/genética , Transexualismo/genética , Síndrome de Turner
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