RESUMO
Complex chromosome rearrangements (CCRs) are structural rearrangements involving at least three chromosomes and three or more chromosome breakpoints. Generally, balanced CCR carriers have a normal phenotype but they are at a higher reproductive risk. Azoospermia was discovered in the male partner of a couple with primary infertility. Conventional cytogenetics identified a CCR refined by fluorescent in situ hybridisation. The CCR involved three chromosomes, four breakpoints and an insertion. A literature search identified 43 phenotypically normal males referred for reproductive problems presenting a CCR. More males were ascertained because of spermatogenesis failure or disturbances than because of repeated abortions and/or birth of a malformed child. Male carriers of CCR produce a high frequency of chromosomally abnormal spermatozoa due to the aberrant segregation of the rearranged chromosomes. The number of chromosomes and breakpoints involved in the rearrangement, the position of breakpoints, the relative size of the resultant chromosomes and the presence or absence of recombination inside the paired-rearranged segments are presumed to affect the fertility of the carrier. Testicular biopsy should not be performed in males with azoospermia. Intracytoplasmic sperm injection should not be proposed as a procedure for treating the infertility of CCR male carriers as a successful result is unlikely.
Assuntos
Azoospermia/genética , Pontos de Quebra do Cromossomo , Rearranjo Gênico/genética , Infertilidade Masculina/genética , Adulto , Azoospermia/complicações , Azoospermia/diagnóstico , Cromossomos Humanos Par 1/genética , Cromossomos Humanos Par 15/genética , Cromossomos Humanos Par 5/genética , Humanos , Infertilidade Masculina/diagnóstico , Infertilidade Masculina/etiologia , Masculino , Mutagênese Insercional/genética , FenótipoRESUMO
Among various causes responsible for infertility, it has been admitted for a long time that male infertility can be due to impaired spermatogenesis and/or balanced structural chromosomal abnormalities. Sperm DNA fragmentation is also considered as another cause of infertility. Most of the studies on male infertility have concerned either aneuploidy in the sperm of carriers of constitutional chromosomal abnormalities or sperm DNA fragmentation. This review is aimed at analyzing these 2 parameters in the same patients. Furthermore, we present work on the study of these 2 parameters in the same gametes of 4 carriers of a balanced chromosomal abnormality. Meiotic segregation was analyzed by fluorescent in situ hybridization and DNA fragmentation was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay. It was shown that aneuploidy and DNA fragmentation were increased in the sperm of carriers of a balanced chromosomal abnormality. For all 4 carriers of a balanced structural abnormality, there was a 2-5 times higher proportion of spermatozoa with unbalanced chromosomal content and fragmented DNA than among those with normal/balanced content. Moreover, we found a non-random distribution with more gametes with DNA fragmentation when these arose from a particular segregation mode. The mechanism which would tend to explain our results is abortive apoptosis. In conclusion, both meiotic segregation and DNA fragmentation studies should be integrated in the genetic exploration of male carriers of a chromosomal structural abnormality.
Assuntos
Aneuploidia , Aberrações Cromossômicas , Fragmentação do DNA , Espermatozoides , Ejaculação , Feminino , Humanos , Masculino , Interações Espermatozoide-Óvulo , Espermatozoides/metabolismoRESUMO
In the infertile male population, there is a 2-20-time higher probability of having a structural chromosomal abnormality than in general population. Generally, these men have a normal phenotype but they can have sperm abnormalities. As they can produce a variable proportion of unbalanced gametes, it is important to evaluate the percentage of unbalanced chromosomal spermatozoa to assess the risk of injecting a chromosomally unbalanced gamete during ICSI procedure. We report here the meiotic segregation analysis of chromosomes in spermatozoa of 12 men with a balanced reciprocal translocation and 4 men with a Robertsonian translocation using a fluorescent in situ hybridisation analysis. The frequencies of normal or balanced spermatozoa ranged from 34.4% to 49.1% in balanced reciprocal translocation carriers. For Robertsonian translocation, the frequencies of normal or balanced spermatozoa ranged from 78.4% to 91.2%. These analyses allow us to define the orientation of genetic counselling according to the results of meiotic segregation obtained. As a last resort, it could then be discussed of the possibility of having recourse to donor spermatozoa or adoption.
Assuntos
Segregação de Cromossomos , Hibridização in Situ Fluorescente , Infertilidade Masculina/diagnóstico , Meiose/genética , Espermatozoides/patologia , Translocação Genética , Humanos , Infertilidade Masculina/genética , Cariotipagem , MasculinoAssuntos
Deleção Cromossômica , Cromossomos Humanos Par 20/genética , Hibridização Genômica Comparativa/métodos , Isocromossomos/genética , Mielofibrose Primária/genética , Idoso , Quebra Cromossômica , Feminino , Dosagem de Genes , Humanos , Hibridização in Situ Fluorescente , Janus Quinase 2/genéticaRESUMO
Semen analysis of a 31-year-old infertile man showed a severe oligoteratozoospermia. Karyotyping of peripheral blood lymphocytes showed a 47,XY,+18[13]/46,XY[16] mosaicism. Cultured skin fibroblasts, right and left jugal smears showed 3, 50 and 65% trisomic cells respectively. The aim of the study was to evaluate the aneuploidy rates of chromosomes X, Y, 13, 18 and 21 and the diploidy rate in his spermatozoa by fluorescence in situ hybridization. The rate of disomy 18 was significantly increased in the spermatozoa of the patient (0.68%) compared to the control group (0.06%). A statistically significant difference in the rates of disomy for chromosome 13 (0.46% vs. 0.14%) and the gonosomes (0.78% vs. 0.24%) and diploidy (0.93% vs. 0.34%) was also found between the patient and the control group. However, no significant difference was observed for chromosome 21 (0.34% vs. 0.15%). Our results show evidence of a generalized perturbation of the meiotic mechanism that could lead to an increased risk for a mosaic trisomy 18 infertile male of producing offspring with aneuploidy that is not only on account of the father's mosaicism, but also more particularly because of severe oligoteratozoospermia.
Assuntos
Aneuploidia , Cromossomos Humanos Par 18/genética , Oligospermia/genética , Cromossomos Humanos Par 13/genética , Cromossomos Humanos Par 21/genética , Cromossomos Humanos X/genética , Cromossomos Humanos Y/genética , Humanos , Hibridização in Situ Fluorescente , Masculino , Meiose , Mosaicismo , Análise do Sêmen , Espermatozoides , TrissomiaRESUMO
Isochromosome of the long arm of chromosome 20 with loss of interstitial material [ider(20q)] is a variant of deletion of chromosome 20q and a rare abnormality in myelodysplastic syndrome (MDS). We studied seven cases with an ider(20q) in MDS. Fluorescence in situ hybridization (FISH) studies showed all proximal breakpoints to be consistently located in 20q11.21 band whereas distal breakpoints were variable. Amplification of HCK, TNFRSF6B and DIDO1 genes included in retained regions associated with loss of tumour suppressor genes in deleted regions could explain cell tumour progression and possibly the less favourable prognosis of ider(20q) compared with del(20q).
Assuntos
Cromossomos Humanos Par 20 , Isocromossomos , Síndromes Mielodisplásicas/genética , Idoso de 80 Anos ou mais , Quebra Cromossômica , Proteínas de Ligação a DNA/genética , Feminino , Amplificação de Genes , Deleção de Genes , Humanos , Hibridização in Situ Fluorescente/métodos , Masculino , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-hck/genética , Membro 6b de Receptores do Fator de Necrose Tumoral/genéticaRESUMO
Supernumerary marker chromosomes (SMCs) are defined as extrastructurally abnormal chromosomes which origin and composition cannot be determined by conventional cytogenetics. SMCs are an heterogeneous group of abnormalities concerning all chromosomes with variable structure and size and are associated with phenotypic heterogeneity. The characterisation of SMCs is of utmost importance for genetic counselling. Different molecular techniques are used to identify chromosomal material present in markers such as 24-colour FISH (MFISH, SKY), centromere specific multicolour FISH (cenMFISH) and derivatives (acroMFISH, subcenMFISH), comparative genomic hybridisation (CGH), arrayCGH, and targeted FISH techniques (banding techniques, whole chromosome painting...). Based on the morphology of SMC with conventional cytogenetic and clinical data, we tried to set up different molecular strategies with all available techniques.
Assuntos
Transtornos Cromossômicos/diagnóstico , Análise Citogenética/métodos , Marcadores Genéticos , Técnicas de Diagnóstico Molecular/métodos , Aneuploidia , Centrômero/genética , Bandeamento Cromossômico/métodos , Transtornos Cromossômicos/genética , Coloração Cromossômica/métodos , Duplicação Gênica , Humanos , Hibridização in Situ Fluorescente/métodos , Cariotipagem/métodos , Hibridização de Ácido Nucleico/métodos , Sondas de OligonucleotídeosRESUMO
Balanced reciprocal translocations are the most common structural abnormalities; most involve two autosomes while a few involve a gonosome (X or Y chromosome) and an autosome. These rearrangements are usually associated with infertility and/or a higher risk of chromosomal imbalances among offspring. This 26 years old man was first seen because of a 3-year history of primary infertility. He had been found to have a translocation, t(X;18)(q11;p11.1), inherited from his mother when he was 9 years old. Semen analysis showed a very severe oligoasthenoteratozoospermia (OAT). A total of 447 spermatozoa were analysed using three-colour fluorescent in situ hybridization (FISH). The alternate segregation pattern, leading to a normal or balanced chromosomal content, was found in 54.36% of the spermatozoa studied. The frequencies of Adjacent I, Adjacent II, 3:1 segregation and diploidy (or 4:0 segregation) were 8.28, 5.14, 22.37 and 2.01%, respectively. Balanced reciprocal translocations between an autosome and the X chromosome lead to important disruptions in human spermatogenesis. Almost all the males with an X-autosome translocation have azoospermia. The man reported here had very severe OAT and is the first in whom the meiotic segregation pattern was analysed. This case further emphasizes the interest in performing FISH studies in infertile males with a chromosomal translocation to provide them with a personalized imbalance risk.
Assuntos
Segregação de Cromossomos , Cromossomos Humanos X , Heterozigoto , Infertilidade Masculina/genética , Espermatozoides/fisiologia , Translocação Genética , Adulto , Astenozoospermia/genética , Humanos , Cariotipagem , Masculino , Meiose/genética , Mães , Oligospermia/genética , Espermatozoides/anormalidadesRESUMO
Rearrangements involving the IGH gene have been identified in about 50% of non-Hodgkin B-cell lymphomas (NHLs) and correlated to clinically relevant subgroups. However, the detection rate largely varied with the technique used. We analyzed the incidence of IGH rearrangements using several fluorescence in situ hybridization (FISH) techniques on metaphases obtained from 96 patients with nodal NHL. An IGH rearrangement was identified in 71 cases (74%). A t(14;18)(q32;q21) was found in 37 of the 42 follicular lymphomas (88.1%) studied and a t(11;14)(q13;q32) in 12 of the 14 mantle cell lymphomas (85.7%). IGH rearrangements were identified in 21 of the 40 diffuse large B-cell lymphomas (52.5%), including seven t(14;18)(q32;q21) and four t(3;14)(q27;q32). Conventional cytogenetics was uninformative in several cases. However, the complemented analysis using 24-color FISH, chromosomal whole paints, telomeric probes and locus specific identifiers enabled us to characterize complex and/or masked IGH translocations in follicular lymphomas and mantle cell lymphomas and to identify all the chromosomal partners involved in IGH rearrangements in diffuse large B-cell lymphomas. This study shows the interest of using metaphase FISH in addition to conventional cytogenetics. Following banding techniques, FISH with the IGH dual color probe can be the first approach in NHL, after which chromosome painting and 24-color FISH can be used to identify the chromosomal partners involved in IGH rearrangements. The identification of these genes is of utmost importance for a better understanding of the molecular mechanisms involved in the genesis of lymphoma.
Assuntos
Rearranjo Gênico , Cadeias Pesadas de Imunoglobulinas/genética , Linfoma de Células B/genética , Translocação Genética , Humanos , Hibridização in Situ Fluorescente , Cariotipagem , Linfoma de Células B/patologia , MetáfaseAssuntos
Cromossomos Humanos Par 1 , Cromossomos Humanos Par 9 , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Leucemia de Células B/genética , Proteínas de Fusão Oncogênica/química , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Proteínas Proto-Oncogênicas c-abl/genética , Translocação Genética , Criança , Clonagem Molecular , Humanos , Imunofenotipagem , Cariotipagem , Masculino , Proteínas de Fusão Oncogênica/metabolismo , FosforilaçãoRESUMO
Marker chromosomes are defined as 'structurally abnormal chromosomes in which no part can be identified' (ISCN 1995). Supernumerary marker chromosomes (SMC) are 'additional markers' whose origin and composition cannot be determined by conventional cytogenetics. Molecular cytogenetic methods are necessary to identify these additional chromosomal markers. In one third, the SMCs are clinically well-defined in the literature, the remaining two thirds present a major problem for genetic counselling in prenatal diagnosis. At present, different molecular cytogenetic methods are used to determine the origin of SMCs. In this work, we studied 13 SMCs detected by RHG-banding, completed by C-banding and/or NOR-staining. 24-color FISH was used as the primary technique when the chromosomal origin was unknown. Targeted FISH procedures with specific probes (whole chromosome painting, centromeric probe, locus-specific identifier, BAC, etc.) were then performed to confirm and/or specify the chromosomal material present in the SMC. Seven SMCs were found to be associated with phenotypic abnormalities. Five derived from autosomes and two from gonosomes; these are: der(12)t(4;12), dic(15), i(18p), r(19), der(22)t(11;22), r(X), and der(Y). Two markers, r(8) and idic(15), were identified during investigations of infertile couples. Three cases seemed to be phenotypically normal. Four were discovered prenatally: r(2) and r(19) referred for elevated maternal serum markers, der(13/21) referred for advanced maternal age. The fourth SMC, der(14/22), was found during familial investigation following the identification of the same marker in an infertile son. The precise characterisation of the SMCs is of utmost importance for genetic counselling, especially in prenatal diagnosis.
Assuntos
Mapeamento Cromossômico/métodos , Cromossomos Humanos/ultraestrutura , Citogenética/métodos , Animais , Aberrações Cromossômicas , Bandeamento Cromossômico , Transtornos Cromossômicos , Feminino , Marcadores Genéticos , Humanos , Hibridização in Situ Fluorescente , Fenótipo , Gravidez , Diagnóstico Pré-NatalRESUMO
A 35-year-old male was found to have a 45,XY,-14,der(18)t(14;18)(q11;p11.3) karyotype during the investigations for a couple with infertility for 8 years. Two sperm samples were obtained and analysed in triple fluorescence in situ hybridization (FISH) with the D18Z1 and LSI IGH/BCL2 probes. The frequency of gametes exhibiting a normal or balanced chromosomal equipment was 87.26 and 90.97% in samples 1 and 2, respectively. No statistically significant difference was found between the results of meiotic segregation of both samples. These proportions are close to those observed among Robertsonian translocation carriers. They can probably be explained by the formation of trivalent in cis configuration during meiosis I between the derivative chromosome and the normal chromosomes 14 and 18, as in Robertsonian translocation carriers. These results suggest that the configuration adopted at pachytene strongly determines the segregation mode that will be preferentially followed during anaphase I.
Assuntos
Deleção Cromossômica , Segregação de Cromossomos/genética , Cromossomos Humanos Par 14/genética , Cromossomos Humanos Par 18/genética , Infertilidade Masculina/genética , Meiose/genética , Translocação Genética , Adulto , Humanos , Hibridização in Situ Fluorescente , MasculinoRESUMO
We report the prenatal diagnosis of a mosaic 45,X/46,X,r(X)/46,XX foetus after amniocentesis for maternal second-trimester serum screening. Biparental contribution for the X chromosomes suggest the postzygotic formation of the X ring. The ring is tiny but contains the X-inactive-specific transcript gene (XIST). However, we could not determine whether XIST was correctly expressed or not. The foetal ultrasound examination at 21, 25, 31 weeks' gestation showed no physical abnormalities, prompting the parents to continue the pregnancy. Physical examination at one year-old revealed normal growth and psychomotor development. Only three cases exhibiting an identical 45,X/46,X,r(X)/46,XX mosaicism, but detected postnatally, are reported in the literature. This is the first reported case ofa mosaic 45,X/46,X,r(X)/46,XX identified in the prenatal period.
Assuntos
Cromossomos Humanos X/genética , Doenças Fetais/diagnóstico por imagem , Mosaicismo , Diagnóstico Pré-Natal , Cromossomos em Anel , Adulto , Feminino , Humanos , Fenótipo , Gravidez , Ultrassonografia , Inativação do Cromossomo X/genéticaRESUMO
Unlike the small proximal 15q deletions causing Prader-Willi and/or Angelman syndrome, distal deletions of the terminal long arm of chromosome 15 have rarely been described. To the best of our knowledge, only four patients with a pure terminal 15q deletion have been documented in the literature. We report here on an unexpected abnormal hybridization pattern for the 15q specific subtelomeric control probe (clone 154P1) of the commercial SNRPN probe in a girl referred for suspicion of Angelman syndrome. Investigation by fluorescent in situ hybridization (FISH) using bacterial artificial chromosome (BAC) clones defined a partial monosomy 15q26.2 --> 15qter for a minimal critical region of approximately 5.7 Mb, which is the most distal de novo 15qter deletion reported to date. All the de novo 15qter deletion cases, including ours, presented with pre- and post-natal growth retardation related to the loss of one copy of the IGF1R gene. Based on the comparaison with the previous published cases and owing to the clinical phenotype of our patient, we define a new subtelomeric 15qter syndrome which would be characterized by intrauterine growth retardation and global post-natal growth failure, variable mental retardation, facial anomalies including relative micrognathia and triangular facies and minor malformations of the extremities including proximally placed thumbs, cubitus valgus, and brachydactyly with tappering of the digits.
Assuntos
Deleção Cromossômica , Cromossomos Humanos Par 15/genética , Anormalidades Múltiplas/genética , Anormalidades Múltiplas/patologia , Pré-Escolar , Feminino , Dedos/anormalidades , Transtornos do Crescimento/patologia , Humanos , Hibridização in Situ Fluorescente , Cariotipagem , Pescoço/anormalidades , Telômero/genéticaRESUMO
The safe and efficient delivery of nucleic acids into haematopoietic stem cells (HSCs) has a wide range of therapeutic applications. Although viruses are being used in most clinical trials owing to their high transfection efficacy, recent results highlight many concerns about their use. Synthetic transfection reagents, in contrast, have the advantage of being safe and easy to manage while their low transfection efficiency remains a hurdle that needs to be addressed before they can be widely used. Using information on transfection mechanisms, a new family of monocationic lipids called lipophosphoramides was synthesized. Their efficiency to transfer genes into haematopoietic cell lines (K562, Jurkat and Daudi) and CD34+ cells was assessed. In this study, we report that one of these new compounds, KLN-5, leads to more efficient transfection activity than one of our previously most efficient reagents (EG-308) and the commercially available monocationic lipids (DC-CHOL and DOTAP/DOPE) (P<0.05). In addition, only a slight toxicity related to the chemical structure of the new compounds is observed. Moreover, we show that KLN-5 can successfully carry the transgene into haematopoietic progenitor cells (CD34+). These results demonstrate that synthetic transfection reagents represent a viable alternative to viruses and could have potential practical utility in a number of applications.
Assuntos
Células Sanguíneas/metabolismo , Colesterol/análogos & derivados , Células-Tronco Hematopoéticas/metabolismo , Lipídeos , Compostos Organofosforados/química , Transfecção/métodos , Amidas , Antígenos CD34 , Linhagem Celular Tumoral , Ácidos Graxos Monoinsaturados , Humanos , Fosfatidiletanolaminas , Fosforamidas , Ácidos Fosfóricos , Plasmídeos , Compostos de Amônio Quaternário , Relação Estrutura-Atividade , Transfecção/normas , beta-Galactosidase/genéticaRESUMO
The objective of this study was to determine the aneuploidy level in spermatozoa in two men with globozoospermia. Sperm nuclei were analysed by fluorescence in-situ hybridization (FISH) in two infertile males with globozoospermia. Dual FISH for chromosomes 7 and 9, 13 and 21, and triple FISH for chromosomes X, Y, and 18 was performed. The main outcome measured was meiotic segregation differences between both globozoospermic men and controls. A statistically significant difference in disomies 13 and 21 was found between patients 1 and 2. The diploidy rate of spermatozoa of patient 1 (0.876%) was significantly increased compared with that of patient 2 (0.304%) and control men (0.293%). In conclusion there seems to be a slightly increased frequency of aneuploidy in round-headed spermatozoa. However, it is unlikely that these aneuploid spermatozoa would be used in assisted reproduction techniques.
Assuntos
Aneuploidia , Cromossomos Humanos/genética , Oligospermia/genética , Espermatozoides/patologia , Adulto , Segregação de Cromossomos , Diploide , Humanos , Masculino , Meiose , Técnicas de Reprodução AssistidaRESUMO
The meiotic segregation pattern of 83 men carrying a balanced reciprocal translocation between two autosomes has already been published. Nevertheless, the question of intraindividual variations has not been addressed yet. A 32-year-old patient was found to be a carrier of a t(9;22)(q21;q11.2) during the investigations for a couple with infertility for 3 years. Two sperm samples were obtained at more than 3 months interval. Both sperm samples were analyzed in triple FISH with the D9Z1 and LSI BCR/ABL ES translocation probes. The frequency of gametes exhibiting a chromosomal imbalance was 45.32% and 42.1% in samples 1 and 2, respectively, with the unbalanced spermatozoa resulting from adjacent 1, adjacent 2, and 3:1 segregation in decreasing frequencies. No statistically significant difference was found between both segregation profiles. Four studies have analyzed the meiotic segregation pattern of translocations within families; they found similar profiles of meiotic segregation in each family, but not between families. This suggests, along with our results, that meiotic segregation is not a random process. More studies on intraindividual variations are necessary to allow a better understanding of the meiotic behaviour of chromosomal rearrangements and the practical interest of studies of this kind.