RESUMO
Resistance to thyroid hormone alpha is an emerging syndrome, with up to now a limited number of published cases. Some features are common to most of the patients, but there is still some work to provide a comprehensive description of the full spectrum of the syndrome. A survey of the strategy to screen for and characterize the mutations in TR α gene is given.
Assuntos
Mutação , Receptores alfa dos Hormônios Tireóideos/genética , Síndrome da Resistência aos Hormônios Tireóideos/genética , Animais , Animais Geneticamente Modificados , Biomarcadores , Criança , Biologia Computacional/métodos , Análise Mutacional de DNA , Modelos Animais de Doenças , Feminino , Estudos de Associação Genética/métodos , Predisposição Genética para Doença , Testes Genéticos , Humanos , Receptores alfa dos Hormônios Tireóideos/metabolismo , Síndrome da Resistência aos Hormônios Tireóideos/diagnóstico , Síndrome da Resistência aos Hormônios Tireóideos/metabolismo , Hormônios Tireóideos/sangue , Hormônios Tireóideos/metabolismoRESUMO
BACKGROUND: Determination of BRAF mutation status is mandatory in the management of patients with inoperable stage IIIC or stage IV melanoma. Currently, molecular biology (MB) has been validated for detecting the presence of BRAF mutations. OBJECTIVE: To compare the sensitivity, specificity and cost of immunohistochemistry (IHC) (clone VE1) versus BM methods (qPCR and Sanger sequencing). PATIENTS AND METHODS: All the samples for which BRAF mutation status was requested between March 2013 and February 2015 at the cellular and molecular analysis laboratory of the Angers Hospital were included retrospectively and consecutively. The IHC (clone VE1) and BM analyses were performed with the same formalin-fixed paraffin embedded tumour samples. The cost of these two methods was determined on the basis of the cost for the French Health Insurance. RESULTS: Two hundred and seven samples were subjected to a determination of BRAF mutational status in IHC and BM. Only one sample was discordant between these two methods (positive in IHC, negative in BM). The sensitivity and specificity of the IHC was 100% and 99.25% respectively. The ratio of the cost of IHC/BM testing was 1:2.1. CONCLUSION: IHC (clone VE1) is a specific, sensitive and economic method for determining BRAFV600E mutation status. Nevertheless, this method must be validated in order to be integrated into a decisional algorithm, alongside BM methods, to determine whether targeted BRAF-inhibitor therapy is indicated.
Assuntos
Biomarcadores Tumorais/genética , Imuno-Histoquímica , Melanoma , Biologia Molecular , Mutação , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias Cutâneas , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Células Clonais , Feminino , França , Humanos , Imuno-Histoquímica/economia , Imuno-Histoquímica/métodos , Masculino , Melanoma/diagnóstico , Melanoma/economia , Melanoma/genética , Melanoma/patologia , Pessoa de Meia-Idade , Biologia Molecular/economia , Estadiamento de Neoplasias , Valor Preditivo dos Testes , Estudos Retrospectivos , Sensibilidade e Especificidade , Neoplasias Cutâneas/diagnóstico , Neoplasias Cutâneas/economia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologiaRESUMO
INTRODUCTION: Fondaparinux (FDX) was demonstrated to be cardioprotective in a rat model of myocardial ischemia reperfusion. In this model, FDX reduced infarct size after 2h of reperfusion, involving the activation of the survivor activating factor enhancement (SAFE) pathway as early as 30min post-reperfusion. Our aim was to study if this cardioprotection could be explained by anti-inflammatory mechanisms and a protective effect on vessels. METHODS: Wistar male rats were subjected to 40minutes (min) of myocardial ischemia, followed by 30min or 2h of reperfusion. Rats were randomized into four groups: control 30min (n=7), FDX 30min (n=7), control 2h (n=7), and FDX 2h (n=7). The FDX groups received 10mg/kg injection of FDX 10min prior to initiating reperfusion. We studied: 1) mRNA expression of endothelial markers, such as thrombomodulin (TM), endothelial protein C receptor (EPCR), and tissue factor (TF) and 2) proteic expression of ICAM-1, NF-κB, IκB, and JNK. Leukocyte infiltration was assessed by histochemistry. We also evaluated TM and EPCR mRNA expression in a model of isolated rat mesenteric arteries incubated with FDX. RESULTS: FDX upregulated the expression of TM and EPCR mRNA in the models of myocardial infarction and isolated mesenteric arteries. No difference was observed between the treated and control groups regarding the expression of pro-inflammatory signaling proteins, adhesion molecules, and leukocyte infiltration after 2h of reperfusion. CONCLUSION: The cardioprotective effect of FDX at early-stage reperfusion could be related to vascular protection, yet not to an anti-inflammatory effect.
Assuntos
Anticoagulantes/uso terapêutico , Cardiotônicos/uso terapêutico , Infarto do Miocárdio/tratamento farmacológico , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Polissacarídeos/uso terapêutico , Receptores de Endotelina/genética , Trombomodulina/genética , Animais , Modelos Animais de Doenças , Fondaparinux , Masculino , Artérias Mesentéricas/efeitos dos fármacos , Artérias Mesentéricas/metabolismo , Artérias Mesentéricas/patologia , Infarto do Miocárdio/genética , Infarto do Miocárdio/patologia , Traumatismo por Reperfusão Miocárdica/genética , Traumatismo por Reperfusão Miocárdica/patologia , Miocárdio/metabolismo , Miocárdio/patologia , RNA Mensageiro/genética , Ratos , Ratos Wistar , Regulação para Cima/efeitos dos fármacosRESUMO
Germline anomalies of the INK4a-ARF and Cdk4 genes were sought in a series of 89 patients suspected of having a genetic predisposition to melanoma. Patients were selected based on the following criteria: (a) familial melanoma (23 cases), (b) multiple primary melanoma (MPM; 18 cases), (c) melanoma and additional unrelated cancers (13 cases), (d) age at diagnosis less than 25 years (21 cases), and (e) nonphoto-induced melanoma (NPIM; 14 cases). Mutations of INK4a-ARF and Cdk4 were characterised by automated sequencing, and germline deletions of INK4a-ARF were also examined by real-time quantitative PCR. Seven germline changes of INK4a-ARF, five of which were novel, were found in seven patients (8%). Four were very likely to be pathogenic mutations and were found in three high-risk melanoma families and in a patient who had a pancreatic carcinoma in addition to melanoma. Three variants of uncertain significance were detected in one MPM patient, one patient <25 years, and one NPIM patient. No germline deletion of INK4a-ARF was found in 71 patients, and no Cdk4 mutation was observed in the 89 patients. This study confirms that INK4a-ARF mutations are infrequent outside stringent familial criteria, and that germline INK4a-ARF deletions are rarely involved in genetic predisposition to melanoma.
Assuntos
Inibidor p16 de Quinase Dependente de Ciclina/genética , Predisposição Genética para Doença , Mutação em Linhagem Germinativa , Melanoma/genética , Neoplasias Cutâneas/genética , Proteína Supressora de Tumor p14ARF/genética , Adolescente , Adulto , Idade de Início , Idoso , Análise Mutacional de DNA , DNA de Neoplasias , Feminino , Humanos , Masculino , Melanoma/patologia , Pessoa de Meia-Idade , Linhagem , Reação em Cadeia da Polimerase , Fatores de Risco , Neoplasias Cutâneas/patologiaAssuntos
Transplante de Medula Óssea , DNA/metabolismo , Eletroforese Capilar/métodos , Repetições Minissatélites , Reação em Cadeia da Polimerase , Quimeras de Transplante/genética , Adolescente , Alelos , Medula Óssea/patologia , Criança , Pré-Escolar , DNA/sangue , Primers do DNA/química , Eletroforese Capilar/economia , Estudos de Avaliação como Assunto , Feminino , Fluorescência , Humanos , Lactente , Leucemia/genética , Leucemia/terapia , Masculino , Paris , Reação em Cadeia da Polimerase/economia , Polimorfismo Genético/genética , Estudos Prospectivos , Sensibilidade e Especificidade , Fatores de TempoRESUMO
Human DAP3 (death-associated protein-3) has been identified as an essential positive mediator of programmed cell death. Structure-function studies have shown previously the N-terminal extremity of the protein to be required in apoptosis induction. Analysis of human DAP3 gene structure predicted 13 exons and subsequent targeting prediction by two software packages (MITOPROT and TargetP) gave a high probability for mitochondrial targeting. The predicted N-terminal targeting structure was also found in the mouse, Drosophila, and C. elegans orthologues with a strong sequence homology between mouse and human. Secondary structure analyses identified alpha-helical structures typical of mitochondrial target peptides. To confirm experimentally this targeting we constructed a fusion protein with N-terminal human DAP3 upstream of enhanced green fluorescent protein (EGFP). Confocal analysis of transfected human fibroblasts clearly demonstrated EGFP localization exclusive to mitochondria. The positioning of this key apoptotic factor at the heart of the mitochondrial pathway provides exciting insight into its role in programmed cell death.