RESUMEN
OBJECTIVE: Trisomy of chromosome 13, 18, 21 and sex chromosome aneuploidies are the most common chromosomal abnormalities encountered in prenatal screening and are responsible for polymaformative syndrome associated with severe mental retardation. This high degree of morbidity justifies the prenatal diagnosis of these aneuploidies. Fetal nuchal translucency measurement and maternal serum biochemical marker assessment are the method of choice used for antenatal screening of aneuploidies. This prenatal screening leads to numerous maternal samplings followed by karyotyping which is cost-effective, time consuming, while results are generally returned between 2 and 3 weeks. Our study describes the research of common aneuploidies by molecular biology. We have used on one hand the MLPA kit (MRC Holland) based on amplification of specific DNA probes that hybridize with chromosomes 13, 18, 21, X, Y. On the other hand we have developed multiplex fluorescent PCR, amplifying microsatellite DNA sequences. PATIENTS AND METHODS: We have evaluated the efficiency of these two techniques to detect chromosomal abnormalities by screening 400 amniotic fluids or chorionic villi samples obtained from pregnant women presenting a high risk of chromosomal aneuploidy. RESULTS: We have found four trisomies 21, one trisomy 13, one monosomy 13, one trisomy 18, two triploidies, one trisomy X and one Klinefelter syndrome. DISCUSSION AND CONCLUSION: In our study we have detected by molecular biology, in less than 48 h, 100% of common chromosomal aneuploidies without false positive or false negative results which could lead molecular biology as a method of choice for the rapid detection of common aneuploidies in addition to fetal karyotyping.
Asunto(s)
Aneuploidia , Síndrome de Down/diagnóstico , Diagnóstico Prenatal/métodos , Ultrasonografía Prenatal/métodos , Muestra de la Vellosidad Coriónica , Aberraciones Cromosómicas , Femenino , Humanos , Cariotipificación , Edad Materna , Repeticiones de Microsatélite , Medida de Translucencia Nucal , Valor Predictivo de las Pruebas , Embarazo , Diagnóstico Prenatal/normas , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Factores de Tiempo , Ultrasonografía Prenatal/normasAsunto(s)
Regiones no Traducidas 3'/genética , Resistencia a la Proteína C Activada/diagnóstico , Electroforesis en Gel de Poliacrilamida , Factor V/genética , Hipoprotrombinemias/diagnóstico , Mutación Puntual , Reacción en Cadena de la Polimerasa/métodos , Protrombina/genética , Trombofilia/genética , Resistencia a la Proteína C Activada/genética , Sustitución de Aminoácidos , Predisposición Genética a la Enfermedad , Humanos , Hipoprotrombinemias/genética , Mutagénesis Sitio-Dirigida , Mutación Missense , Trombofilia/diagnóstico , Trombosis de la Vena/etiologíaAsunto(s)
Alelos , Electroforesis/métodos , Mutación , Protrombina/genética , Humanos , Datos de Secuencia MolecularRESUMEN
The Bcr-Abl fusion protein plays a crucial role in the initiation and maintenance of chronic myelogenous leukemia (CML). However, additional events are necessary for the transition from the chronic phase to the terminal phase of the disease. To identify genes involved in the disease progression, we constructed a subtractive library from enriched K562 cell mRNA. We obtained 1084 cDNA clones. After a specific hybridization of these clones with a cDNA probe from either chronic phase or K562 cells, 43 clones which present a differential hybridization level have been selected. Among them, several clones corresponded to ribosomal protein genes showing an increased transcription level during the blast crisis. We observed variations in the expression of a cellular adhesion molecule, a laminin-binding protein. An increased transcription level of the MAZ gene has been shown in the terminal phase of the disease. This gene encodes a protein that regulates the transcription of myc.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Transcripción Genética , Crisis Blástica , Proteínas Portadoras/biosíntesis , Moléculas de Adhesión Celular/biosíntesis , Cartilla de ADN , Proteínas de Unión al ADN , Progresión de la Enfermedad , Biblioteca de Genes , Humanos , Laminina/metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva/sangre , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Recuento de Leucocitos , Reacción en Cadena de la Polimerasa , Proteínas Proto-Oncogénicas c-myc/biosíntesis , ARN Mensajero/biosíntesis , Proteínas Ribosómicas/biosíntesis , Factores de Transcripción/biosíntesis , Células Tumorales CultivadasRESUMEN
Mouse resistance to several intracellular pathogens including Mycobacteria, Leishmania, and Salmonella is under the control of the Chromosome (Chr) 1 Natural Resistance Associated Macrophage Protein I gene (Nramp1). This gene could have an economic and health importance for domestic animals and humans as well. Therefore, equivalents of the NRAMP1 gene have been cloned by several research groups in various animal species. To study in sheep the influence of the NRAMP1 gene on the susceptibility to intracellular pathogens induced diseases, we have cloned the sheep NRAMP1 cDNA by screening a splenic cDNA library. The genomic organization of the sheep NRAMP1 gene was then determined by sequencing the exon/intron boundaries. The transcription start points (tsp) from the NRAMP1 mRNA have been located with primer extension experiments. RT-PCR reactions have been used to determine the profile of mRNA expression of this gene.
Asunto(s)
Proteínas Portadoras/genética , Proteínas de Transporte de Catión , ADN Complementario/genética , Genes/genética , Proteínas de la Membrana/genética , Ovinos/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Bovinos , Pollos , Clonación Molecular , ADN Complementario/química , ADN Complementario/aislamiento & purificación , Exones , Expresión Génica , Humanos , Intrones , Ratones , Datos de Secuencia Molecular , ARN Mensajero/genética , ARN Mensajero/metabolismo , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Distribución Tisular , Transcripción GenéticaRESUMEN
A 33-year-old man with an atypical course of hypereosinophilic syndrome including malignant hypercalcemia, osteolytic lesions and evolution into severe myelofibrosis was treated by allogeneic bone marrow transplantation after conditioning with cytoxan and total body irradiation. As the transplant was sex-mismatched, chimerism was studied by means of cytogenetic analysis and Y chromosomal DNA amplification by PCR assay. Long-term complete remission has been assessed by normalization of blood cell counts, magnetic resonance imaging and karyotypic analysis. A relapse was observed 40 months after transplantation. The patient remains alive 44 months post-BMT. This case report is compared with those reported in the literature.
Asunto(s)
Trasplante de Médula Ósea , Síndrome Hipereosinofílico/terapia , Mielofibrosis Primaria/terapia , Adulto , Humanos , Síndrome Hipereosinofílico/fisiopatología , Masculino , Trasplante HomólogoRESUMEN
In one case out of four, allogeneic BMT concerns a male recipient and a female donor. The monitoring of sex-matched BMT can be carried out by PCR amplification on Y-specific chromosome sequences (YCS), whatever the hematological disease. Twelve patients with sex-mismatched non-T-depleted BMT were first studied through a qualitative PCR, which gave semi-quantitative results. When the qualitative PCR revealed YCS, a competitive amplification was performed in order to estimate the YCS amount in the patient blood sample. For the purpose of the study, we classified the patients in two categories according to the results obtained 9 months after BMT. For 10 patients, we did not detect any YCS amplification after this time. These patients were in complete cytogenetic and clinical remission. For the remaining two patients, we always found male DNA in their blood samples. These patients were in cytogenetic remission but relapsed and died 21 and 25 months after BMT. Our results suggest that the persistence of male cells in peripheral blood, even at the low rate of 1% or 0.1%, 1 year after sex-mismatched BMT, is a bad prognosis.
Asunto(s)
Trasplante de Médula Ósea/patología , Leucemia/patología , Recurrencia Local de Neoplasia/epidemiología , Reacción en Cadena de la Polimerasa , Cromosoma Y , Secuencia de Bases , Trasplante de Médula Ósea/estadística & datos numéricos , Supervivencia Celular , Quimera , ADN/sangre , Sondas de ADN , Femenino , Estudios de Seguimiento , Marcadores Genéticos , Supervivencia de Injerto , Enfermedad Injerto contra Huésped , Humanos , Leucemia/mortalidad , Leucemia/terapia , Masculino , Neoplasia Residual , Pronóstico , Inducción de Remisión , Trasplante Homólogo , Insuficiencia del Tratamiento , Cromosoma Y/genéticaRESUMEN
A 51-year-old man with previously treated CLL received an allogeneic sex mismatched BMT after total body irradiation and high dose chemotherapy. Residual disease was studied at phenotypic and molecular levels including Y chromosome DNA amplification by PCR assay. The patient was clinically disease-free 20 months after BMT with disappearance of the leukemic clone assessed by the most sensitive methods of detection. Long-term follow-up is necessary to ascertain the relevance of Y DNA amplification in predicting outcome in this patient.
Asunto(s)
Trasplante de Médula Ósea , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/terapia , Antineoplásicos/uso terapéutico , Secuencia de Bases , Southern Blotting , ADN de Neoplasias/genética , Amplificación de Genes , Humanos , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Fenotipo , Reacción en Cadena de la Polimerasa , Inducción de Remisión , Irradiación Corporal Total , Cromosoma YRESUMEN
A sheep cDNA library was screened with a human C mu probe, and the complete nucleotide sequence of a 1923 nt cDNA was determined. It contains sequences corresponding to all the exons (VH, DH, JH, CH1, CH2, CH3 and CH4) characteristic of the immunoglobulin mu heavy chain regions. The deduced amino acid sequence shows a percentage of identical residues in the range 65-45% when compared with the mu chains of various species. The VH region of this clone is clearly related to a group of genes that includes mouse VH36-60 and VHQ52, human VH2, VH4 and VH6 gene families and Xenopus VHII gene families. The constant region shows an unusual repartition of cystein and proline residues at the beginning of the CH2 domain, that may result in a molecule with enhanced stability and reduced flexibility.