RESUMO
Self-adaptive methods are recognized as important tools in signal process and analysis. A signal can be decomposed into a serious of new components with these mentioned methods, thus the amount of information is also increased. In order to use these components effectively, a feature set is used to describe them. With the development of pattern recognition, the analysis of self-adaptive components is becoming more intelligent and depend on feature sets. Thus, a new feature is proposed to express the signal based on the hidden property between extreme values. In this investigation, the components are first simplified through a symbolization method. The entropy analysis is incorporated into the establishment of the characteristics to describe those self-adaptive decomposition components according to the relationship between extreme values. Subsequently, Extreme Interval Entropy is proposed and used to realize the pattern recognition, with two typical self-adaptive methods, based on both Empirical Mode Decomposition (EMD) and Empirical Wavelet Transform (EWT). Later, extreme interval entropy is applied in two fault diagnosis experiments. One experiment is the fault diagnosis for rolling bearings with both different faults and damage degrees, the other experiment is about rolling bearing in a printing press. The effectiveness of the proposed method is evaluated in both experiments with K-means cluster. The accuracy rate of the fault diagnosis in rolling bearing is in the range of 75% through 100% using EMD, 95% through 100% using EWT. In the printing press experiment, the proposed method can reach 100% using EWT to distinguish the normal bearing (but cannot distinguish normal samples at different speeds), with fault bearing in 4 r/s and in 8 r/s. The fault samples are identified only according to a single proposed feature with EMD and EWT. Therefore, the extreme interval entropy is proved to be a reliable and effective tool for fault diagnosis and other similar applications.
RESUMO
The purpose of this study was to investigate the role of retinoic acid (RA) and/or dexamethasone and growth hormone releasing hormone (GHRH) in the induction of somatotroph cell differentiation. Immunohistochemistry, radioimmunoassay, 3-(4,5-dimethylthiazol -1,2-y1)-2,5-diphenyltetrazolium bromide assay, and immune electron microscopy were employed to determine the effect of incubation with these constituents on the differentiation into somatotrophs of cells isolated from the rat embryonic pituitary gland. RA administration increased the proportion of growth hormone (GH) positive somatotroph cells and GH secretion in embryonic pituitary cells (P<0.01). After 4 days of incubation with RA, additional administration of dexamethasone further increased the proportion of somatotroph cells and GH secretion (P<0.01), and increased the number of secretory granules in the somatotroph cells. Addition of GHRH alone had no such effect (P>0.05). However, addition of GHRH to treatment with RA plus dexamethasone significantly increased both the proportion of somatotroph cells and the secretion of GH compared to treatment with RA or dexamethasone alone or RA plus dexamethasone (P<0.01). RA promoted the early differentiation of somatotroph cells, dexamethasone promoted the differentiation and maturation of somatotroph cells and in addition, RA, dexamethasone and GHRH together exerted synergistic effects that markedly promoted somatotroph cell differentiation, maturation and GH secretion.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Dexametasona/farmacologia , Somatotrofos/citologia , Somatotrofos/efeitos dos fármacos , Tretinoína/farmacologia , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Feminino , Hormônio do Crescimento/metabolismo , Hormônio Liberador de Hormônio do Crescimento/farmacologia , Masculino , Modelos Animais , Hipófise/citologia , Hipófise/embriologia , Gravidez , Ratos , Ratos Sprague-Dawley , Somatotrofos/metabolismo , Fatores de TempoRESUMO
Butyrylcholinesterase (BChE, EC 3.1.1.8) is important in human cocaine metabolism despite its limited ability to hydrolyze this drug. Efforts to improve the catalytic efficiency of this enzyme have led to a quadruple mutant cocaine hydrolase, "CocH", that in animal models of addiction appears promising for treatment of overdose and relapse. We incorporated the CocH mutations into a BChE-albumin fusion protein, "Albu-CocH", and evaluated the pharmacokinetics of the enzyme after i.v. injection in rats. As assessed from the time course of cocaine hydrolyzing activity in plasma, Albu-CocH redistributed into extracellular fluid (16% of estimated total body water) with a t(1/2) of 0.66h and it underwent elimination with a t(1/2) of 8h. These results indicate that the enzyme has ample stability for short-term applications and may be suitable for longer-term treatment as well. Present data also confirm the markedly enhanced power of Albu-CocH for cocaine hydrolysis and they support the view that Albu-CocH might prove valuable in treating phenomena associated with cocaine abuse.
Assuntos
Albuminas/metabolismo , Butirilcolinesterase/metabolismo , Transtornos Relacionados ao Uso de Cocaína/tratamento farmacológico , Cocaína/toxicidade , Albuminas/farmacocinética , Animais , Biocatálise , Butirilcolinesterase/farmacocinética , Feminino , Masculino , Ratos , Ratos WistarRESUMO
A coordinated functional genomics program was implemented to identify secreted polypeptides with therapeutic applications in the treatment of diabetes. Secreted factors were predicted from a diverse expressed-sequence tags (EST) database, representing >1,000 cDNA libraries, using a combination of bioinformatic algorithms. Subsequently, approximately 8,000 human proteins were screened in high-throughput cell-based assays designed to monitor key physiological transitions known to be centrally involved in the physiology of type 2 diabetes. Bone morphogenetic protein-9 (BMP-9) gave a positive response in two independent assays: reducing phosphoenolpyruvate carboxykinase (PEPCK) expression in hepatocytes and activating Akt kinase in differentiated myotubes. Purified recombinant BMP-9 potently inhibited hepatic glucose production and activated expression of key enzymes of lipid metabolism. In freely fed diabetic mice, a single subcutaneous injection of BMP-9 reduced glycemia to near-normal levels, with maximal reduction observed 30 hours after treatment. BMP-9 represents the first hepatic factor shown to regulate blood glucose concentration. Using a combination of bioinformatic and high-throughput functional analyses, we have identified a factor that may be exploited for the treatment of diabetes.
Assuntos
Proteínas Morfogenéticas Ósseas/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Diabetes Mellitus/genética , Diabetes Mellitus/metabolismo , Perfilação da Expressão Gênica/métodos , Animais , Proteínas Morfogenéticas Ósseas/química , Proteínas Morfogenéticas Ósseas/uso terapêutico , Células Cultivadas , Diabetes Mellitus/tratamento farmacológico , Desenho de Fármacos , Glucose/metabolismo , Fator 2 de Diferenciação de Crescimento , Fatores de Diferenciação de Crescimento , Humanos , Rim/química , Rim/embriologia , Rim/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas/química , Proteínas/genética , Proteínas/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Valores de Referência , Alinhamento de Sequência/métodos , Análise de Sequência de Proteína/métodos , Integração de SistemasRESUMO
Successive rational mutations of human butyrylcholinesterase (BChE) followed by fusion to human serum albumin have yielded an efficient hydrolase that offers realistic options for therapy of cocaine overdose and abuse. This albumin-BChE prevented seizures in rats given a normally lethal cocaine injection (100 mg/kg, i.p.), lowered brain cocaine levels even when administered after the drug, and provided rescue after convulsions commenced. Moreover, it selectively blocked cocaine-induced reinstatement of drug seeking in rats that had previously self-administered cocaine. The enzyme treatment was well tolerated and may be worth exploring for clinical application in humans.
Assuntos
Comportamento Aditivo/enzimologia , Comportamento Aditivo/prevenção & controle , Butirilcolinesterase/farmacologia , Cocaína/antagonistas & inibidores , Cocaína/toxicidade , Hidrolases/farmacologia , Animais , Butirilcolinesterase/química , Butirilcolinesterase/genética , Células CHO , Cocaína/administração & dosagem , Cricetinae , Cricetulus , Relação Dose-Resposta a Droga , Feminino , Humanos , Hidrolases/síntese química , Hidrolases/genética , Masculino , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Engenharia de Proteínas/métodos , Ratos , Ratos Sprague-Dawley , Ratos Wistar , AutoadministraçãoRESUMO
Genetic studies in mice and humans have shown that the transforming growth factor-beta (TGF-beta) type-I receptor activin receptor-like kinase 1 (ALK1) and its co-receptor endoglin play an important role in vascular development and angiogenesis. Here, we demonstrate that ALK1 is a signalling receptor for bone morphogenetic protein-9 (BMP-9) in endothelial cells (ECs). BMP-9 bound with high affinity to ALK1 and endoglin, and weakly to the type-I receptor ALK2 and to the BMP type-II receptor (BMPR-II) and activin type-II receptor (ActR-II) in transfected COS cells. Binding of BMP-9 to ALK2 was greatly facilitated when BMPR-II or ActR-II were co-expressed. Whereas BMP-9 predominantly bound to ALK1 and BMPR-II in ECs, it bound to ALK2 and BMPR-II in myoblasts. In addition, we observed binding of BMP-9 to ALK1 and endoglin in glioblastoma cells. BMP-9 activated Smad1 and/or Smad5, and induced ID1 protein and endoglin mRNA expression in ECs. Furthermore, BMP-9 was found to inhibit basic fibroblast growth factor (bFGF)-stimulated proliferation and migration of bovine aortic ECs (BAECs) and to block vascular endothelial growth factor (VEGF)-induced angiogenesis. Taken together, these results suggest that BMP-9 is a physiological ALK1 ligand that plays an important role in the regulation of angiogenesis.
Assuntos
Receptores de Activinas Tipo II/fisiologia , Proteínas Morfogenéticas Ósseas/fisiologia , Proliferação de Células/efeitos dos fármacos , Células Endoteliais/fisiologia , Fator 2 de Crescimento de Fibroblastos/farmacologia , Neovascularização Fisiológica/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/farmacologia , Receptores de Ativinas Tipo I/fisiologia , Animais , Células COS , Bovinos , Linhagem Celular Tumoral , Movimento Celular , Células Cultivadas , Chlorocebus aethiops , Células Endoteliais/efeitos dos fármacos , Fator 2 de Diferenciação de Crescimento , Fatores de Diferenciação de Crescimento , Humanos , Hipoxantina Fosforribosiltransferase/metabolismo , Camundongos , Ligação Proteica , Receptores de Superfície Celular/metabolismo , Transdução de SinaisRESUMO
CC chemokine ligand 14, CCL14, is a human CC chemokine that is of recent interest because of its natural ability, upon proteolytic processing of the first eight NH2-terminal residues, to bind to and signal through the human immunodeficiency virus type-1 (HIV-1) co-receptor, CC chemokine receptor 5 (CCR5). We report X-ray crystallographic structures of both full-length CCL14 and signaling-active, truncated CCL14 [9-74] determined at 2.23 and 1.8 A, respectively. Although CCL14 and CCL14 [9-74] differ in their ability to bind CCR5 for biological signaling, we find that the NH2-terminal eight amino acids (residues 1 through 8) are completely disordered in CCL14 and both show the identical mode of the dimeric assembly characteristic of the CC type chemokine structures. However, analytical ultracentrifugation studies reveal that the CCL14 is stable as a dimer at a concentration as low as 100 nM, whereas CCL14 [9-74] is fully monomeric at the same concentration. By the same method, the equilibrium between monomers of CCL14 [9-74] and higher order oligomers is estimated to be of EC1,4 = 4.98 microM for monomer-tetramer conversion. The relative instability of CCL14 [9-74] oligomers as compared to CCL14 is also reflected in the Kd's that are estimated by the surface plasmon resonance method to be approximately 9.84 and 667 nM for CCL14 and CCL14 [9-74], respectively. This approximately 60-fold difference in stability at a physiologically relevant concentration can potentially account for their different signaling ability. Functional data from the activity assays by intracellular calcium flux and inhibition of CCR5-mediated HIV-1 entry show that only CCL14 [9-74] is fully active at these near-physiological concentrations where CCL14 [9-74] is monomeric and CCL14 is dimeric. These results together suggest that the ability of CCL14 [9-74] to monomerize can play a role for cellular activation.
Assuntos
Sinalização do Cálcio/efeitos dos fármacos , Quimiocinas CC/química , Quimiocinas CC/farmacologia , Fragmentos de Peptídeos/farmacologia , Processamento de Proteína Pós-Traducional , Receptores CCR5/agonistas , Sítios de Ligação , Quimiocinas CC/metabolismo , Cristalografia por Raios X , Dimerização , Endopeptidases/metabolismo , HIV-1/efeitos dos fármacos , HIV-1/metabolismo , Humanos , Concentração Inibidora 50 , Fragmentos de Peptídeos/metabolismo , Receptores CCR5/metabolismo , Receptores de HIV/efeitos dos fármacos , Receptores de HIV/metabolismo , Relação Estrutura-Atividade , Ressonância de Plasmônio de Superfície , Linfócitos T/efeitos dos fármacos , Linfócitos T/virologia , Ultracentrifugação , Internalização do Vírus/efeitos dos fármacosRESUMO
Escherichia coli SeqA binds clusters of transiently hemimethylated GATC sequences and sequesters the origin of replication, oriC, from methylation and premature reinitiation. Besides oriC, SeqA binds and organizes newly synthesized DNA at replication forks. Binding to multiple GATC sites is crucial for the formation of stable SeqA-DNA complexes. Here we report the crystal structure of the oligomerization domain of SeqA (SeqA-N). The structural unit of SeqA-N is a dimer, which oligomerizes to form a filament. Mutations that disrupt filament formation lead to asynchronous DNA replication, but the resulting SeqA dimer can still bind two GATC sites separated from 5 to 34 base pairs. Truncation of the linker between the oligomerization and DNA-binding domains restricts SeqA to bind two GATC sites separated by one or two full turns. We propose a model of a SeqA filament interacting with multiple GATC sites that accounts for both origin sequestration and chromosome organization.
Assuntos
Proteínas da Membrana Bacteriana Externa/química , Replicação do DNA , Proteínas de Ligação a DNA/química , Proteínas de Escherichia coli/química , Estrutura Quaternária de Proteína , Proteínas da Membrana Bacteriana Externa/genética , Cristalografia por Raios X , DNA/química , DNA/metabolismo , Proteínas de Ligação a DNA/genética , Dimerização , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Modelos Moleculares , Dados de Sequência Molecular , MutaçãoRESUMO
Bone morphogenetic proteins (BMPs), a subset of the transforming growth factor (TGF)-beta superfamily, regulate a diverse array of cellular functions during development and in the adult. BMP-9 (also known as growth and differentiation factor (GDF)-2) potently induces osteogenesis and chondrogenesis, has been implicated in the differentiation of cholinergic neurons, and may help regulate glucose metabolism. We have determined the structure of BMP-9 to 2.3 A and examined the differences between our model and existing crystal structures of other BMPs, both in isolation and in complex with their receptors. TGF-beta ligands are translated as precursors, with pro-regions that generally dissociate after cleavage from the ligand, but in some cases (including GDF-8 and TGF-beta1, -2, and -3), the pro-region remains associated after secretion from the cell and inhibits binding of the ligand to its receptor. Although the proregion of BMP-9 remains tightly associated after secretion, we find, in several cell-based assays, that the activities of BMP-9 and BMP-9.pro-region complex were equivalent. Activin receptor-like kinase 1 (ALK-1), an orphan receptor in the TGF-beta family, was also identified as a potential receptor for BMP-9 based on surface plasmon resonance studies (BIAcore) and the ability of soluble ALK-1 to block the activity of BMP-9.pro-region complex in cell-based assays.
Assuntos
Proteínas Morfogenéticas Ósseas/química , Cristalografia por Raios X/métodos , Células 3T3-L1 , Receptores de Ativinas Tipo I/metabolismo , Receptores de Activinas Tipo II , Fosfatase Alcalina/metabolismo , Sequência de Aminoácidos , Animais , Proteína Morfogenética Óssea 2 , Proteína Morfogenética Óssea 6 , Proteína Morfogenética Óssea 7 , Proteínas Morfogenéticas Ósseas/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células , Condrogênese , Cromatografia , Eletroforese em Gel de Poliacrilamida , Genes Reporter , Glucose/metabolismo , Fator 2 de Diferenciação de Crescimento , Ligantes , Camundongos , Modelos Moleculares , Dados de Sequência Molecular , Miostatina , Neurônios/metabolismo , Osteogênese , Ligação Proteica , Ratos , Homologia de Sequência de Aminoácidos , Transdução de Sinais , Ressonância de Plasmônio de Superfície , Fator de Crescimento Transformador beta/metabolismoRESUMO
The SeqA protein binds clusters of fully methylated or hemimethylated GATC sequences at oriC and negatively modulates the initiation of DNA replication. We find that SeqA can be proteolytically cleaved into an N-terminal multimerization and a C-terminal DNA-binding domain and have determined the crystal structure of the C-terminal domain in complex with a hemimethylated GATC site. SeqA makes direct hydrogen bonds and van der Waals contacts with the hemimethylated A-T base pair in addition to interactions with the surrounding bases and DNA backbone. The tetrameric protein-DNA complex found in the crystal suggests that SeqA binds multiple GATC sites on separate DNA duplexes, altering the overall DNA topology and sequestering oriC from replication initiation.