ABSTRACT
Turner syndrome (TS) is caused by a complete or partial absence of an X or Y chromosome, including chromosomal mosaicism, affecting 1 in 2500 female live births. Sister chromatid exchange (SCE) is used as a sensitive indicator of spontaneous chromosome instability. Cells from mosaic patients constitute useful material for SCE evaluations as they grow under the influence of the same genetic background and endogenous and exogenous factors. We evaluated the proliferation dynamics and SCE frequencies of 45,X and 46,XN cells of 17 mosaic TS patients. In two participants, the 45,X cells exhibited a proliferative disadvantage in relation to 46,XN cells after 72 h of cultivation. The analysis of the mean proliferation index (PI) showed a trend for a significant difference between the 45,X and 46,X+der(X)/der(Y) cell lineages; however, there were no intra-individual differences. On the other hand, mean SCE frequencies showed that 46,X+der(X) had the highest mean value and 46,XX the lowest, with 45,X occupying an intermediate position among the lineages found in at least three participants; moreover, there were intra-individual differences in five patients. Although 46,X+der(X)/der(Y) cell lineages, found in more than 70% of participants, were the most unstable, they had a slightly higher mean PI than the 45,X cell lineages in younger (≤17 years) mosaic TS participants. This suggests that cells with a karyotype distinct from 45,X may increase with time in mosaic TS children and adolescents.
ABSTRACT
Chronic Granulomatous Disease (CGD) is an inborn error of immunity characterized by impaired phagocyte function, recurrent fungal and bacterial infections and granuloma formation in multiple organs. Pediatric myelodysplastic Syndrome (MDS) is a rare hematological stem cell disease that leads to an ineffective hematopoiesis with variable risk of evolution to acute leukemias. Both disorders are rare and have distinct pathophysiologic mechanisms, with no known association. A 7-month-old boy presenting with recurrent infections and anemia at age 2 months underwent immunological, hematological and genetic investigation that culminated in the diagnosis of both CGD and MDS. Next generation sequencing was performed and identified a silent variant predicted as of Uncertain Significance, located in the splicing site at the end of exon 5 in CYBB. CYBB variants account for at least two thirds of CGD cases, but no previous descriptions of this variant were found in ClinVar or The Human Gene Mutation Database (HGMD) databases. We were able to demonstrate an exon 5 skipping on the proband's cDNA, which strongly suggests the disruption of the NADPH oxidase complex, abrogating the formation of reactive oxygen species from neutrophils. Moreover, erythroid cell lineage could be also affected by NADPH oxidase complex damages. Further investigation is needed to evaluate the potential effect of CYBB gene alterations in hematopoiesis, as well as in MDS and CGD association.
Subject(s)
Granulomatous Disease, Chronic/genetics , Hematopoiesis/genetics , Myelodysplastic Syndromes/genetics , NADPH Oxidase 2/genetics , Exons/genetics , Granulomatous Disease, Chronic/diagnosis , Granulomatous Disease, Chronic/pathology , Humans , Infant , Male , Mutation/genetics , Myelodysplastic Syndromes/diagnosis , Myelodysplastic Syndromes/pathology , NADPH Oxidases/genetics , Neutrophils/metabolism , Neutrophils/pathology , Pediatrics , Phagocytes/metabolism , RNA Splicing/genetics , Reactive Oxygen Species/metabolismABSTRACT
[This corrects the article DOI: 10.1371/journal.pone.0156692.].
ABSTRACT
L-asparaginase is an enzyme used as a chemotherapeutic agent, mainly for treating acute lymphoblastic leukemia. In this study, the gene of L-asparaginase from Zymomonas mobilis was cloned in pET vectors, fused to a histidine tag, and had its codons optimized. The L-asparaginase was expressed extracellularly and intracellularly (cytoplasmically) in Escherichia coli in far larger quantities than obtained from the microorganism of origin, and sufficient for initial cytotoxicity tests on leukemic cells. The in silico analysis of the protein from Z. mobilis indicated the presence of a signal peptide in the sequence, as well as high identity to other sequences of L-asparaginases with antileukemic activity. The protein was expressed in a bioreactor with a complex culture medium, yielding 0.13 IU/mL extracellular L-asparaginase and 3.6 IU/mL intracellular L-asparaginase after 4 h of induction with IPTG. The cytotoxicity results suggest that recombinant L-asparaginase from Z. mobilis expressed extracellularly in E.coli has a cytotoxic and cytostatic effect on leukemic cells.
Subject(s)
Antineoplastic Agents/therapeutic use , Asparaginase/therapeutic use , Escherichia coli/metabolism , Leukemia/drug therapy , Recombinant Proteins/therapeutic use , Zymomonas/enzymology , Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Asparaginase/genetics , Asparaginase/pharmacology , Base Sequence , Bioreactors , Cell Cycle/drug effects , Cell Line, Tumor , Cell Nucleus Shape/drug effects , Cell Proliferation/drug effects , Cell Survival/drug effects , Child , Child, Preschool , Cloning, Molecular , Computer Simulation , Female , Humans , Infant , Leukemia/pathology , Male , Phylogeny , Recombinant Proteins/genetics , Recombinant Proteins/pharmacologyABSTRACT
A técnica de AgNOR é um método histoquímico que evidencia as Regiões organizadoras de nucléolo (NORs), sendo considerado um bom marcador de proliferação celular e consequentemente pode ser usado para avaliar malignidade. O Câncer de colo uterino pode ser detectado através do exame preventivo do câncer, através do Método de Papanicolaou, possibilitando o tratamento precose. Os casos sugestivos bem como os inconclusivos devem ser confirmados pela histologia e/ou colposcopia, o que pode acarretar evasão das pacientes dos Serviços de Saúde. Os testes moleculares, quando apresentam resultados positivos para vírus oncogênicos, não determinam necessariamente o desenvolvimento de uma displasia, sendo necessário outros fatores. O objetivo deste trabalho foi avaliar a técnica de AgNOR aplicada em esfregaços de material cervical uterino e analisar a sua associação com o diagnóstico citológico e histológico em lesões do epitélio cervical uterino. Foram estudadas 85 pacientes na faixa etária de 18 a 65 anos de idade. Amostras citológicas de material cervical foram submetidas à coloração de AgNOR e Papanicolaou e abiópsia à coloração de HE. As laminas coradas com AgNOR e o diagnóstico citológico e histológico com correlação positiva para número dos pontos de AgNOR (r de Spearman: 0,8468 e 0,8441 respectivamente). Portanto conclui-se que a Técnica de AgNOR é eficiente, e que pode ser empregada em material cervical citológico nos casos inclusivos.
Subject(s)
Female , Humans , Cell Biology , Histology , Neoplasms , ColposcopyABSTRACT
Objetivo: Padronizar em nosso meio a identificaçao da mutaçao de ponto no gene de transtiretina responsável pela polineuropatia amilóide familiar. Métodos: Amplificaçao do exon 2 do gene da transtiretina, seguida pela digestao com endonuclease Nsi I, foi realizada em amostra de DNA obtida de uma paciente afetada. Resultados: Aproximadamente metade do produto de 215 pares de base é clivada em fragmentos de 120 pb e 95 pb, enquanto o DNA correspondente ao alelo normal é resistente à digestao enzimática. Conclusao: A técnica de PCR é um método confiável e reprodutivo para identificaçao de portadores do gene mutante associado à forma brasileira de polineuropatia amilóide familiar
Subject(s)
Humans , Female , Adult , Amyloidosis , Polymerase Chain ReactionABSTRACT
O artigo refere o resultado da análise cromossômica de 27 portadores de retinoblastoma uni ou bilateral, de ambos os sexos, esporádicos ou familiais, virgens de tratamento ou após 6 meses do término de quimioterapia sistêmica, triados no Ambulatório de Oftalmologia do Hospital A.C. Camargo da Fundaçäo Antônio Prudente/SP. Foi dada preferência àqueles que apresentavam, no quadro clínico geral, malformaçöes congênitas, alteraçöes fenotípicas ou retardo mental. Encontrou-se cromossomopatia inespecífica em 2 pacientes submetidos à poliquimioterapia e cromosssomopatia conhecidas em outros 3 (47,XX+G; 46,XY/47,XY+G; 46,XY/46, XY13q-). Os autores discutem os resultados, a associaçäo de cromossomopatia e retinoblastoma e a indicaçäo do exame do cariótipo nesses pacientes
Subject(s)
Child , Female , Male , Chromosomes, Human, Pair 13/pathology , Eye Neoplasms/physiopathology , Karyotyping/instrumentation , Retinoblastoma/analysis , BrazilABSTRACT
No presente trabalho descrevemos uma modificaçäo da técnica de baneamento R, que utiliza incorporaçäo de 5-BrdU, tratamento pelo Hoechst 33258 e coloraçäo pelo Giemsa (RBG). Esta técnica produz boa qualidade de bandamento além de alta reprodutibilidade
Subject(s)
Chromosome Banding/methodsABSTRACT
Analisa-se a evoluçäo de 48 pacientes portadores de retinoblastoma, com um total de 58 olhos afetados, atendidos no Hospital A.C. Camargo - Fundaçäo Antonio Prudente, Säo Paulo, no período de agosto de 1979 a agosto de 1983. Ao diagnóstico, 43,8% dos pacientes apresentavam tumores intra-oculares e 56,2%, extra oculares. A sobrevida foi respectivamente de 90,1% e 40,7%. Estes dados mostram a evidente piora do prognóstico nos casos avançados. A partir do estudo das faixas de incidência, das causas do diagnóstico tardio e da evoluçäo da doença, propöe-se a intensificaçäo da divulgaçäo desta moléstia entre pediatras, oftalmologistas, populaçäo leiga, principalmente profissionais que lidam com crianças com baixa idade, a fim de facilitar o diagnóstico precoce. Propöe-se ainda a introduçäo de novos esquemas terapêuticos para os casos avançados, com a utilizaçäo de drogas que atravessem a barreira hematencefálica