RESUMEN
BACKGROUND: In patients with coronary artery disease and reduced ejection fraction, amiodarone reduces mortality by decreasing sudden death. Because the latter may be triggered by coronary artery thrombosis as much as ventricular arrhythmias, amiodarone might interfere with tissue factor (TF) expression and thrombus formation. METHODS AND RESULTS: Clinically relevant plasma concentrations of amiodarone reduced TF activity and impaired carotid artery thrombus formation in a mouse photochemical injury model in vivo. PTT, aPTT, and tail bleeding time were not affected; platelet number was slightly decreased. In human endothelial and vascular smooth muscle cells, amiodarone inhibited tumor necrosis factor (TNF)-alpha and thrombin-induced TF expression as well as surface activity. Amiodarone lacking iodine and the main metabolite of amiodarone, N-monodesethylamiodarone, inhibited TF expression. Amiodarone did not affect mitogen-activated protein kinase activation, TF mRNA expression, and TF protein degradation. Metabolic labeling confirmed that amiodarone inhibited TF protein translation. CONCLUSIONS: Amiodarone impairs thrombus formation in vivo; in line with this, it inhibits TF protein expression and surface activity in human vascular cells. These pleiotropic actions occur within the range of amiodarone concentrations measured in patients, and thus may account at least in part for its beneficial effects in patients with coronary artery disease.
Asunto(s)
Amiodarona/farmacología , Trombosis de las Arterias Carótidas/metabolismo , Trombosis de las Arterias Carótidas/prevención & control , Tromboplastina/biosíntesis , Amiodarona/análogos & derivados , Animales , Antiarrítmicos/farmacología , Traumatismos de las Arterias Carótidas/tratamiento farmacológico , Traumatismos de las Arterias Carótidas/etiología , Traumatismos de las Arterias Carótidas/genética , Traumatismos de las Arterias Carótidas/metabolismo , Trombosis de las Arterias Carótidas/genética , Células Cultivadas , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Humanos , Ratones , Ratones Endogámicos C57BL , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Biosíntesis de Proteínas/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Tromboplastina/genéticaRESUMEN
The nuclear protein LF-B1 (also referred to as HNF-1) is a transcription activator required for the expression of several liver-specific genes. LF-B1 has been purified to homogeneity from rat liver nuclear extracts. The sequence of the protein has been partially determined and, subsequently, overlapping cDNA clones containing the entire open reading frame of LF-B1 were isolated. The full-length cDNA encodes a 628 amino acid protein and directs the synthesis in vitro of a protein capable of binding DNA with the same specificity as LF-B1. The cDNA was recombined into a vaccinia virus vector and active LF-B1 was obtained from infected HeLa cells. Addition of the vaccinia recombinant protein to rat spleen extracts results in activation of transcription of an LF-B1-dependent promoter. The DNA binding domain of LF-B1 is located in the amino-terminal part of the protein and displays distant structural similarity to the homeobox domain. The distribution of LF-B1 mRNA is restricted to liver, which correlates with the tissue-specific expression of its target genes.
Asunto(s)
Proteínas de Unión al ADN/aislamiento & purificación , Genes Homeobox , Hígado/análisis , Proteínas Nucleares/aislamiento & purificación , Factores de Transcripción/aislamiento & purificación , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Cromatografía de Afinidad , Clonación Molecular , ADN/aislamiento & purificación , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Regulación de la Expresión Génica , Factor Nuclear 1 del Hepatocito , Factor Nuclear 1-alfa del Hepatocito , Factor Nuclear 1-beta del Hepatocito , Masculino , Datos de Secuencia Molecular , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Especificidad de Órganos , Ratas , Ratas Endogámicas , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
A full-length cDNA of the rat liver Nuclear Factor 1 (NF1L21) has been cloned and expressed in S. cerevisiae to analyse the architecture of its activation domain. NF1L21 displays a specific DNA-binding activity, as well as the ability to activate transcription from an artificial NF 1-responsive promoter in yeast. Interaction of two or more NF1L21 molecules with multiple sites on the same promoter activated transcription in a synergistic fashion. Functional analysis of the activation domain of NF1L21 reveals a tripartite structure. Two distinct positive elements are required for NF1L21 -mediated transcription activation. A proline-rich element sandwiched between these two positive domains attenuates their transactivation potential. A shorter NF1L variant (NFlL4) in which the distal positive element is replaced by a different sequence was also isolated. NF1L4 displays the same DNA-binding activity and dimerisation properties as NF1L21, but is unable to activate transcription in yeast.