RESUMEN
This work was developed to evaluate the impact of the addition of proteases on the performance characteristics, egg quality, relative weight of digestive organs, and intestinal morphometry of laying hens. 390 Hy Line W36® hens were allocated into five treatments and six replicates with 13 animals. The treatments were: 1) Control (standard formulation), 2) Negative control A - NCA (nutritional reduction according to protease A matrix), 3) Negative control B - NCB (nutritional reduction according to protease B matrix), 4) NCA+protease A (0.250 g/kg of feed) and 5) NCB+protease B (0.125 g/kg of feed). Hens fed the NCA, NCB, and NCA+protease A diets showed reductions in feed intake and egg mass. The addition of protease B provided better results for egg production in both percentage and per dozen as compared to the group fed with the NCA+protease A diets. The hens subjected to diets NCA and NCB showed eggs with a reduced eggshell and thickness percentage. However, supplementation with proteases A and B improved these parameters to values similar to the controls. There was no significant effect of the treatments on the relative weight of the liver, proventricle, gizzard, pancreas, and small intestine. However, the addition of protease A resulted in a decreased value for the relative weight of the large intestine. The jejunum and ileum crypt depths were, respectively, smaller in hens fed the control diet in relation to the NCB diet and the NCA and NCB diets. As it can be concluded, Protease B supplementation provided the best performance results.(AU)
Asunto(s)
Animales , Péptido Hidrolasas/fisiología , Pollos/fisiología , Ingestión de Alimentos/fisiologíaRESUMEN
Background: Serratia marcescens is an enteric bacterium with increasing incidence in clinical settings, attributed mainly to the opportune expression of diverse virulence determinants plus a wide intrinsic and acquired antibiotic resistance. Methods: The aim of this study was to compare the virulence factor profiles of 185 Serratia marcescens isolates from different clinical origins. In vitro proteolytic and hemolytic activities, biofilm formation, and motility were assessed in each strain. Additionally, the pathogenicity of four hypervirulent strains was analyzed in vivo in Galleria mellonella. Results: We found that bacterial isolates from wound/abscess and respiratory tract specimens exhibited the highest protease activity along with a strong biofilm production, while uropathogenic isolates showed the highest hemolytic activity. Swarming and swimming motilities were similar among all the strains. However, respiratory tract isolates showed the most efficient motility. Two hyperhemolytic and two hyperproteolytic strains were detected; the latter were more efficient killing Galleria mellonella with a 50%-60% larval mortality 48 hours after challenge. Conclusion: A correlation was found between biofilm formation and proteolytic and hemolytic activities in biopsy specimens and bloodstream isolates, respectively. Overall, it becomes critical to evaluate and compare the clinical strains virulence diversity in order to understand the underlying mechanisms that allow the establishment and persistence of opportunistic bacterial infections in the host.
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Antibacterianos/farmacología , Farmacorresistencia Microbiana/efectos de los fármacos , Serratia marcescens/patogenicidad , Biopelículas/crecimiento & desarrollo , Infección Hospitalaria , Hemólisis/fisiología , Humanos , México/epidemiología , Péptido Hidrolasas/fisiología , Serratia marcescens/aislamiento & purificación , Virulencia , Factores de VirulenciaRESUMEN
Latex, a milky fluid found in several plants, is widely used for many purposes, and its proteins have been investigated by researchers. Many studies have shown that latex produced by some plant species is a natural source of biologically active compounds, and many of the hydrolytic enzymes are related to health benefits. Research on the characterization and industrial and pharmaceutical utility of latex has progressed in recent years. Latex proteins are associated with plants' defense mechanisms, against attacks by fungi. In this respect, there are several biotechnological applications of antifungal proteins. Some findings reveal that antifungal proteins inhibit fungi by interrupting the synthesis of fungal cell walls or rupturing the membrane. Moreover, both phytopathogenic and clinical fungal strains are susceptible to latex proteins. The present review describes some important features of proteins isolated from plant latex which presented in vitro antifungal activities: protein classification, function, molecular weight, isoelectric point, as well as the fungal species that are inhibited by them. We also discuss their mechanisms of action.
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Antifúngicos/farmacología , Quitinasas/farmacología , Látex/química , Péptido Hidrolasas/farmacología , Peroxidasas/farmacología , Lectinas de Plantas/farmacología , Proteínas de Plantas/farmacología , Antifúngicos/clasificación , Antifúngicos/aislamiento & purificación , Botrytis/efectos de los fármacos , Botrytis/crecimiento & desarrollo , Candida albicans/efectos de los fármacos , Candida albicans/crecimiento & desarrollo , Quitinasas/clasificación , Quitinasas/aislamiento & purificación , Quitinasas/fisiología , Fusarium/efectos de los fármacos , Fusarium/crecimiento & desarrollo , Punto Isoeléctrico , Pruebas de Sensibilidad Microbiana , Peso Molecular , Péptido Hidrolasas/clasificación , Péptido Hidrolasas/aislamiento & purificación , Péptido Hidrolasas/fisiología , Peroxidasas/clasificación , Peroxidasas/aislamiento & purificación , Peroxidasas/fisiología , Enfermedades de las Plantas/microbiología , Extractos Vegetales/química , Lectinas de Plantas/clasificación , Lectinas de Plantas/aislamiento & purificación , Lectinas de Plantas/fisiología , Proteínas de Plantas/clasificación , Proteínas de Plantas/aislamiento & purificación , Proteínas de Plantas/fisiología , Plantas/químicaRESUMEN
Demineralization in dentinal caries and erosion exposes dentine organic matrix. This exposed matrix, containing type I collagen and non-collagenous proteins, is then degraded by host collagenolytic enzymes, matrix metalloproteinases (MMPs) and cysteine cathepsins. The knowledge of the identities and function of these enzymes in dentine has accumulated only within the last 15 years, but has already formed a field of research called 'dentine degradomics'. This research has demonstrated the role of endogenous collagenolytic enzymes in caries and erosion development. In demineralized dentine, the enzymes degrade triple-helical collagen molecules, leading to the gradual loss of collagen matrix. Even before that, they can cleave off the terminal non-helical ends of collagen molecules called telopeptides, leading to the structural changes at the intramolecular gap areas, which may affect or even prevent intrafibrillar remineralization, which is considered essential in restoring the dentine's mechanical properties. They may also cause the loss of non-collagenous proteins that could serve as nucleation sites for remineralization. Here we review the findings demonstrating that inhibition of salivary or dentine endogenous MMPs and cysteine cathepsins may provide preventive means against the progression of caries or erosion. Furthermore, we also suggest the future directions for the new experimental preventive research to gain more knowledge of the enzymes and their function during and after dentine demineralization, and the pathways to find the clinically acceptable means to prevent the functional activity of these enzymes.
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Caries Dental/enzimología , Dentina/enzimología , Péptido Hidrolasas/fisiología , Erosión de los Dientes/enzimología , Catepsinas/antagonistas & inhibidores , Catepsinas/metabolismo , Colágeno Tipo I/metabolismo , Colagenasas/metabolismo , Caries Dental/prevención & control , Humanos , Inhibidores de la Metaloproteinasa de la Matriz/uso terapéutico , Metaloproteinasas de la Matriz/metabolismo , Fosfoproteínas/metabolismo , Erosión de los Dientes/prevención & controlRESUMEN
BACKGROUND: Candida species, in conditions of microbiota imbalance or decreased immune defenses, may be one of the main human fungal pathogens. Virulence factors constitute the mechanisms used by the fungus to avoid host defenses. AIMS: This study aimed to investigate the in vitro production of virulence factors, such as hemolytic activity, and deoxyribonuclease (DNase), proteinase, and phospholipase activities in Candida spp. METHODS: Fifty clinical isolates were analyzed for virulence factors: Candida albicans (15), Candida tropicalis (15), Candida parapsilosis (10), Candida glabrata (5), and Candida krusei (5). Hemolytic activity was determined in Sabouraud dextrose agar plates containing 3% glucose and 7% sheep red cells. Culture media containing, respectively, agar-base DNA, egg yolk, and bovine albumin were used to determine DNase, phospholipase and proteinase activities, respectively. RESULTS: Forty-eight (96%) of 50 isolates showed hemolytic activity, with 10 (20%) positive for DNase, 19 (38%) for proteinase, and 16 (32%) for phospholipase. Statistically significant differences were observed between species for phospholipase (p<0.0001) and proteinase (p<0.05) production. CONCLUSIONS: It is concluded that all species had hemolytic activity. DNase activity was detected in all species except in C. glabrata; proteinase activity was detected in C. albicans, C. tropicalis, and C. parapsilosis; and phospholipase activity was observed in C. albicans and C. tropicalis.
Asunto(s)
Candida/enzimología , Proteínas Fúngicas/fisiología , Animales , Candida/clasificación , Candida/aislamiento & purificación , Candida/patogenicidad , Candidiasis/microbiología , Medios de Cultivo , Desoxirribonucleasas/aislamiento & purificación , Desoxirribonucleasas/fisiología , Eritrocitos , Proteínas Fúngicas/aislamiento & purificación , Hemólisis , Humanos , Péptido Hidrolasas/aislamiento & purificación , Péptido Hidrolasas/fisiología , Fosfolipasas/aislamiento & purificación , Fosfolipasas/fisiología , Ovinos , Especificidad de la Especie , VirulenciaRESUMEN
Many cases of infection caused by the oral transmission of Trypanosoma cruzi have been reported during the last decade. These have been due to the contamination of food by faeces from sylvatic triatomines or by leakage from reservoirs in areas where domiciliated vectors have been controlled or where there has been no prior background of domiciliation. The United Nations Food and Agriculture Organization (FAO) and the World Health Organization (WHO) have used epidemiological, clinical and socioeconomic criteria for ranking parasites transmitted by the contamination of food in different areas of the world; T. cruzi was placed tenth in importance amongst a group of 24 parasites in such ranking. Environmental changes such as deforestation and global warming have affected ecotopes and the behaviour of T. cruzi vectors and reservoirs so that these have become displaced to new areas, thereby leading to such new transmission scenario caused by the contamination of food, which requires evaluation in Colombia. The current review deals with the oral transmission of Chagas' disease, emphasising studies aimed at identifying the pertinent risk factors, the triatomine species involved, the physiopathology of oral infection, the parasite's genotypes implicated in this type of transmission in Colombia and other Latin American regions, as well as the need for ongoing epidemiological surveillance and control policies.
Asunto(s)
Enfermedad de Chagas/transmisión , Heces/parasitología , Parasitología de Alimentos , Frutas/parasitología , Insectos Vectores/parasitología , Carne/parasitología , Rhodnius/parasitología , Trypanosoma cruzi/aislamiento & purificación , Verduras/parasitología , Animales , Animales Salvajes/parasitología , Armadillos/parasitología , Bebidas/parasitología , Donantes de Sangre , Enfermedad de Chagas/congénito , Enfermedad de Chagas/epidemiología , Enfermedad de Chagas/parasitología , Colombia , Reservorios de Enfermedades/parasitología , Femenino , Mucosa Gástrica/parasitología , Genotipo , Vivienda , Humanos , Mucosa Bucal/parasitología , Parasitemia/parasitología , Parasitemia/transmisión , Péptido Hidrolasas/fisiología , Embarazo , Complicaciones Infecciosas del Embarazo/parasitología , Proteínas Protozoarias/química , Proteínas Protozoarias/fisiología , Factores de Riesgo , Reacción a la Transfusión , Trypanosoma cruzi/genética , Trypanosoma cruzi/patogenicidad , Trypanosoma cruzi/fisiología , Glicoproteínas Variantes de Superficie de Trypanosoma/química , Glicoproteínas Variantes de Superficie de Trypanosoma/fisiologíaRESUMEN
Durante la última década se han reportado numerosos casos de infección por Trypanosoma cruzi por vía oral, debidos a la contaminación de alimentos con heces de triatominos silvestres o con secreciones de reservorios en áreas donde los vectores domiciliados han sido controlados o no hay antecedentes de domiciliación. Con base en criterios epidemiológicos, clínicos y socioeconómicos, la Organización de las Naciones Unidas para la Agricultura y la Alimentación (FAO) y la Organización Mundial de la Salud (OMS) establecieron una clasificación de los parásitos transmitidos por contaminación de alimentos en diferentes regiones del mundo, en la cual T. cruzi ocupó el décimo lugar de importancia en un grupo de 24 parásitos. Los cambios ambientales, como la deforestación y el calentamiento global, han afectado los ecotopos y el comportamiento de los vectores y de los reservorios de T. cruzi , de manera que estos se han desplazado a nuevas zonas, generando una nueva forma de transmisión por contaminación de alimentos que requiere su evaluación en el país. La presente revisión aborda la transmisión oral de la enfermedad de Chagas con énfasis en los estudios orientados a identificar los factores de riesgo, las especies de triatominos involucrados, la fisiopatología de la infección oral y los genotipos del parásito que están implicados en esta forma de transmisión en Colombia y en otras regiones de América Latina, así como la necesidad de adoptar políticas para su control y vigilancia epidemiológica.
Many cases of infection caused by the oral transmission of Trypanosoma cruzi have been reported during the last decade. These have been due to the contamination of food by faeces from sylvatic triatomines or by leakage from reservoirs in areas where domiciliated vectors have been controlled or where there has been no prior background of domiciliation. The United Nations Food and Agriculture Organization (FAO) and the World Health Organization (WHO) have used epidemiological, clinical and socioeconomic criteria for ranking parasites transmitted by the contamination of food in different areas of the world; T. cruzi was placed tenth in importance amongst a group of 24 parasites in such ranking. Environmental changes such as deforestation and global warming have affected ecotopes and the behaviour of T. cruzi vectors and reservoirs so that these have become displaced to new areas, thereby leading to such new transmission scenario caused by the contamination of food, which requires evaluation in Colombia. The current review deals with the oral transmission of Chagas´ disease, emphasising studies aimed at identifying the pertinent risk factors, the triatomine species involved, the physiopathology of oral infection, the parasite´s genotypes implicated in this type of transmission in Colombia and other Latin American regions, as well as the need for ongoing epidemiological surveillance and control policies.
Asunto(s)
Animales , Femenino , Humanos , Embarazo , Enfermedad de Chagas/transmisión , Parasitología de Alimentos , Heces/parasitología , Frutas/parasitología , Insectos Vectores/parasitología , Carne/parasitología , Rhodnius/parasitología , Trypanosoma cruzi/aislamiento & purificación , Verduras/parasitología , Animales Salvajes/parasitología , Armadillos/parasitología , Donantes de Sangre , Bebidas/parasitología , Transfusión Sanguínea/efectos adversos , Colombia , Enfermedad de Chagas/congénito , Enfermedad de Chagas/epidemiología , Enfermedad de Chagas/parasitología , Reservorios de Enfermedades/parasitología , Genotipo , Mucosa Gástrica/parasitología , Vivienda , Mucosa Bucal/parasitología , Parasitemia/parasitología , Parasitemia/transmisión , Péptido Hidrolasas/fisiología , Complicaciones Infecciosas del Embarazo/parasitología , Proteínas Protozoarias/química , Proteínas Protozoarias/fisiología , Factores de Riesgo , Trypanosoma cruzi/genética , Trypanosoma cruzi/patogenicidad , Trypanosoma cruzi/fisiología , Glicoproteínas Variantes de Superficie de Trypanosoma/química , Glicoproteínas Variantes de Superficie de Trypanosoma/fisiologíaRESUMEN
Cockroaches are among the first insects to appear in the fossil record. This work is part of ongoing research on insects at critical points in the evolutionary tree to disclose evolutionary trends in the digestive characteristics of insects. A transcriptome (454 Roche platform) of the midgut of Periplanetaamericana was searched for sequences of digestive enzymes. The selected sequences were manually curated. The complete or nearly complete sequences showing all characteristic motifs and highly expressed (reads counting) had their predicted sequences checked by cloning and Sanger sequencing. There are two chitinases (lacking mucin and chitin-binding domains), one amylase, two α- and three ß-glucosidases, one ß-galactosidase, two aminopeptidases (none of the N-group), one chymotrypsin, 5 trypsins, and none ß-glucanase. Electrophoretic and enzymological data agreed with transcriptome data in showing that there is a single ß-galactosidase, two α-glucosidases, one preferring as substrate maltase and the other aryl α-glucoside, and two ß-glucosidases. Chromatographic and enzymological data identified 4 trypsins, one chymotrypsin (also found in the transcriptome), and one non-identified proteinase. The major digestive trypsin is identifiable to a major P. americana allergen (Per a 10). The lack of ß-glucanase expression in midguts was confirmed, thus lending support to claims that those enzymes are salivary. A salivary amylase was molecularly cloned and shown to be different from the one from the midgut. Enzyme distribution showed that most digestion occurs under the action of salivary and midgut enzymes in the foregut and anterior midgut, except the posterior terminal digestion of proteins. A counter-flux of fluid may be functional in the midgut of the cockroach to explain the low excretory rate of digestive enzymes. Ultrastructural and immunocytochemical localization data showed that amylase and trypsin are released by both merocrine and apocrine secretion mainly from gastric caeca. Finally, a discussion on Polyneoptera digestive physiology is provided.
Asunto(s)
Digestión/fisiología , Periplaneta/fisiología , Aminopeptidasas/genética , Aminopeptidasas/fisiología , Animales , Secuencia de Bases , Quitinasas/genética , Quitinasas/fisiología , Quimotripsina/genética , Quimotripsina/fisiología , Tracto Gastrointestinal/anatomía & histología , Tracto Gastrointestinal/diagnóstico por imagen , Glucosidasas/genética , Glucosidasas/fisiología , Microscopía Electrónica , Datos de Secuencia Molecular , Péptido Hidrolasas/genética , Péptido Hidrolasas/fisiología , Periplaneta/anatomía & histología , Periplaneta/enzimología , Periplaneta/genética , Reacción en Cadena de la Polimerasa , Transcriptoma/genética , Tripsina/genética , Tripsina/fisiología , Ultrasonografía , beta-Galactosidasa/genética , beta-Galactosidasa/fisiología , beta-Glucosidasa/genética , beta-Glucosidasa/fisiologíaRESUMEN
Trypanosoma cruzi, the agent of the American Trypanosomiasis, Chagas disease, contains cysteine, serine, threonine, aspartyl and metallo peptidases. The most abundant among these enzymes is cruzipain, a cysteine proteinase expressed as a mixture of isoforms, some of them membrane-bound. The enzyme is an immunodominant antigen in human chronic Chagas disease and seems to be important in the host/parasite relationship. Inhibitors of cruzipain kill the parasite and cure infected mice, thus validating the enzyme as a very promising target for the development of new drugs against the disease. In addition, a 30kDa cathepsin B-like enzyme, two metacaspases and two autophagins have been described. Serine peptidases described in the parasite include oligopeptidase B, a member of the prolyl oligopeptidase family involved in Ca(2+)-signaling during mammalian cell invasion; a prolyl endopeptidase (Tc80), against which inhibitors are being developed, and a lysosomal serine carboxypeptidase. Metallopeptidases homologous to the gp63 of Leishmania spp. are present, as well as two metallocarboxypeptidases belonging to the M32 family, previously found only in prokaryotes. The proteasome has properties similar to those of other eukaryotes, and its inhibition by lactacystin blocks some differentiation steps in the life cycle of the parasite. This article is part of a Special Issue entitled: Proteolysis 50 years after the discovery of lysosome.
Asunto(s)
Apoptosis , Autofagia/fisiología , Sistema Digestivo/enzimología , Péptido Hidrolasas/fisiología , Trypanosoma cruzi/enzimología , Factores de Virulencia/fisiología , Animales , Apoptosis/genética , Apoptosis/inmunología , Apoptosis/fisiología , Autofagia/genética , Muerte Celular/genética , Muerte Celular/fisiología , Sistema Digestivo/metabolismo , Humanos , Ratones , Modelos Biológicos , Péptido Hidrolasas/genética , Péptido Hidrolasas/metabolismo , Trypanosoma cruzi/metabolismo , Trypanosoma cruzi/patogenicidad , Factores de Virulencia/genética , Factores de Virulencia/metabolismoRESUMEN
Digestive endoprotease activities of the coconut palm weevil, Homalinotus coriaceus (Coleoptera: Curculionidae), were characterized based on the ability of gut extracts to hydrolyze specific synthetic substrates, optimal pH, and hydrolysis sensitivity to protease inhibitors. Trypsin-like proteinases were major enzymes for H. coriaceus, with minor activity by chymotrypsin proteinases. More importantly, gut proteinases of H. coriaceus were inhibited by trypsin inhibitor from Inga laurina seeds. In addition, a serine proteinase inhibitor from I. laurina seeds demonstrated significant reduction of growth of H. coriaceus larvae after feeding on inhibitor incorporated artificial diets. Dietary utilization experiments show that 0.05% I. laurina trypsin inhibitor, incorporated into an artificial diet, decreases the consumption rate and fecal production of H. coriaceus larvae. Dietary utilization experiments show that 0.05% I. laurina trypsin inhibitor, incorporated into an artificial diet, decreases the consumption rate and fecal production of H. coriaceus larvae. We have constructed a three-dimensional model of the trypsin inhibitor complexed with trypsin. The model was built based on its comparative homology with soybean trypsin inhibitor. Trypsin inhibitor of I. laurina shows structural features characteristic of the Kunitz type trypsin inhibitor. In summary, these findings contribute to the development of biotechnological tools such as transgenic plants with enhanced resistance to insect pests.
Asunto(s)
Fabaceae , Proteínas de Insectos/fisiología , Péptido Hidrolasas/fisiología , Péptidos/farmacología , Proteínas de Plantas/farmacología , Gorgojos/enzimología , Secuencia de Aminoácidos , Animales , Sistema Digestivo/enzimología , Proteínas de Insectos/aislamiento & purificación , Proteínas de Insectos/farmacología , Larva/efectos de los fármacos , Larva/crecimiento & desarrollo , Modelos Moleculares , Datos de Secuencia Molecular , Inhibidores de Proteasas/farmacología , Estructura Terciaria de Proteína , Alineación de Secuencia , Homología Estructural de Proteína , Gorgojos/efectos de los fármacos , Gorgojos/crecimiento & desarrolloRESUMEN
Invasion and metastasis are the most important events in cancer progression. In these two phases, several molecules are implicated and have been long associated with several forms of cancer. Proteases play a critical role not only in tumor cell invasion, but also in the earliest stages of carcinogenesis and its associated changes: angiogenesis and metastasis. Aside from their ability to degrade the extracellular matrix, facilitate invasion and metastasis, proteases target a great variety of substrates that favor or inhibit cancer progression: b-FGF, HGF, VEGF, cell death receptors, cistatin-C, galectin, procollagen, and other proteases. Proteases are also signaling molecules that modulate other molecules by underlying pathways in addition to their degradative role. Proteases form interconnected cascades, circuits and networks that bring about the tumor's potential for malignancy. Although, proteases are regulated by diverse molecules, it is known that tumoral and stromal cells secrete several biological molecules, including cytokines and chemokines that directly or indirectly regulate the protease-expression within the tumor's microenvironment. The present review briefly summarizes some of the major aspects associated with the role of proteases in cancer progression.
Asunto(s)
Neoplasias/enzimología , Péptido Hidrolasas/fisiología , Animales , Membrana Basal/fisiología , Matriz Extracelular/fisiología , Humanos , Neovascularización PatológicaRESUMEN
La invasión y la metástasis son los eventos más importantes en la progresión del cáncer, en los cuales están implicadas muchas moléculas, entre ellas, las proteasas. Éstas desempeñan un papel importante en etapas tempranas de la carcinogénesis, en la invasión, en fenómenos asociados como la angiogénesis y en la metástasis, principalmente por su capacidad para degradar componentes de la matriz extracelular, aunque sus sustratos son de naturaleza diversa: citocinas, quimiocinas, factores de crecimiento (b- FGF, HGF, VEGF) y de muerte celular, cistatina-C, galectina, procolágena y otras proteasas, que pueden favorecer o inhibir la progresión neoplásica. Las proteasas son también moléculas de señalización que modulan a otras moléculas; forman cascadas, circuitos e incluso redes, que en conjunto determinan parte del potencial maligno. Se sabe que tanto la célula tumoral como las del estroma secretan diversos factores que regulan directa e indirectamente la expresión de proteasas en el microambiente tumoral. Esta revisión proporciona un panorama breve y actualizado sobre la participación de las proteasas en la progresión neoplásica.
Invasion and metastasis are the most important events in cancer progression. In these two phases, several molecules are implicated and have been long associated with several forms of cancer. Proteases play a critical role not only in tumor cell invasion, but also in the earliest stages of carcinogenesis and its associated changes: angiogenesis and metastasis. Aside from their ability to degrade the extracellular matrix, facilitate invasion and metastasis, proteases target a great variety of substrates that favor or inhibit cancer progression: b-FGF, HGF, VEGF, cell death receptors, cistatin-C, galectin, procollagen, and other proteases. Proteases are also signaling molecules that modulate other molecules by underlying pathways in addition to their degradative role. Proteases form interconnected cascades, circuits and networks that bring about the tumor's potential for malignancy. Although, proteases are regulated by diverse molecules, it is known that tumoral and stromal cells secrete several biological molecules, including cytokines and chemokines that directly or indirectly regulate the protease-expression within the tumor's microenvironment. The present review briefly summarizes some of the major aspects associated with the role of proteases in cancer progression.
Asunto(s)
Humanos , Animales , Neoplasias/enzimología , Péptido Hidrolasas/fisiología , Matriz Extracelular/fisiología , Membrana Basal/fisiología , Neovascularización PatológicaRESUMEN
Parasites belonging to the Leptomonas genus have been used as model organisms for studying biochemical, cellular, and genetic processes unique to members of the Trypanosomatidae family. In the present study, the cell-associated and extracellular peptidases of three Leptomonas species, Leptomonas collosoma, Leptomonas samueli, and Leptomonas wallacei, were assayed and characterized by gelatin-sodium dodecyl sulfate polyacrylamide gel electrophoresis. All parasites released metallopeptidases, whereas no cell-associated proteolytic activity could be detected in the cellular extracts from L. collosoma. Western blotting probed with a polyclonal antibody raised against gp63 from Leishmania amazonensis revealed two major reactive polypeptides of apparent molecular masses of 63 and 52 kDa, with different intensities in cellular extracts and released proteins from the studied trypanosomatids. Flow cytometry and fluorescence microscopy analyses showed that the gp63-like molecules have a surface location. This is the first report on the presence of gp63-like molecules in L. collosoma, L. samueli, and L. wallacei. The pretreatment of L. samueli and L. wallacei with anti-gp63 antibody significantly diminished their association index to Aedes albopictus cell line (C6/36), suggesting a potential involvement of the gp63-like molecules in the interaction process of these insect trypanosomatids with the vector.
Asunto(s)
Antígenos de Protozoos/fisiología , Adhesión Celular , Péptido Hidrolasas/fisiología , Proteínas Protozoarias/fisiología , Trypanosomatina/fisiología , Aedes , Animales , Antígenos de Protozoos/análisis , Western Blotting , Línea Celular , Electroforesis en Gel de Poliacrilamida , Metaloendopeptidasas/inmunología , Péptido Hidrolasas/análisis , Proteínas Protozoarias/análisis , Proteínas Protozoarias/antagonistas & inhibidores , Trypanosomatina/químicaRESUMEN
Canavalia ensiformis ureases are toxic to insects of different orders. The entomotoxicity of urease is due to a 10 kDa internal peptide released by proteinases in the insect digestive tract. We previously observed that, given orally, urease is toxic to nymphs of Dysdercus peruvianus, but does not affect adults. Here we characterized the major proteolytic activities of D. peruvianus midgut homogenates and investigated their in vitro-catalyzed release of the 10 kDa entomotoxic peptide from urease. Cysteine, aspartic and metalloproteinases are present in both homogenates. Variations in optimal pH and susceptibility to inhibitors indicated differences in the enzyme profiles in the two developmental stages. Only nymph homogenates released approximately 10 kDa fragment(s) from urease, recognized by antibodies against the entomotoxic peptide. Fluorogenic substrates containing urease partial sequences flanking the N-terminal or the C-terminal portion of the entomotoxic peptide were efficiently cleaved by homogenates from nymphs, but much more slowly by the adult homogenate. Different classes of enzymes in the homogenates cleaved both substrates suggesting that in vivo the release of the entomotoxic peptide results from the concerted action of at least two different proteinases. Our findings support the view that a differential processing of ingested urease by the insects explains at least in part the lack of toxicity in adults.
Asunto(s)
Canavalia/metabolismo , Heterópteros/enzimología , Proteínas de Insectos/fisiología , Péptido Hidrolasas/fisiología , Toxinas Biológicas/metabolismo , Ureasa/metabolismo , Animales , Caseínas/metabolismo , Cromatografía Liquida , Heterópteros/crecimiento & desarrollo , Hidrólisis , Proteínas de Insectos/metabolismo , Ninfa/enzimología , Ninfa/crecimiento & desarrollo , Péptido Hidrolasas/metabolismo , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
The cell-associated and extracellular peptidases of Herpetomonas megaseliae grown in brain-heart infusion and in modified Roitman's complex media were analyzed by measuring peptidase activity on gelatin, casein and hemoglobin in zymograms. Casein was the best proteinaceous substrate for the peptidase detection on both growth conditions. However, no proteolytic activity was detected when hemoglobin was used. Our results showed that cellular cysteine peptidase (115-100, 40 and 35 kDa) and metallopeptidase (70 and 60 kDa) activities were detected on both media in casein and gelatin zymograms. Additionally, the use of casein in the gel revealed a distinct acidic metallopeptidase of 50 kDa when the parasite was cultured in the modified Roitman's complex medium. Irrespective of the culture medium composition, H. megaseliae released metallopeptidases exclusively in the extracellular environment. The presence of gp63-like molecules on the H. megaseliae surface was shown by flow cytometry using anti-gp63 antibody raised against recombinant gp63 from Leishmania mexicana. The pre-treatment of parasites with phospholipase C reduced the number of gp63-positive cells, suggesting that these molecules were glycosylphosphatidylinositol-anchored to the surface. Additionally, the supernatant obtained from phospholipase C-treated cells and probed with anti-cross-reacting determinant confirmed that at least a 52 kDa gp63-like molecule is glycosylphosphatidylinositol-anchored. Furthermore, we assessed a possible function for the gp63-like molecules in H. megaseliae on the interaction with explanted guts of its original host, Megaselia scalaris, and with an experimental model employing Aedes aegypti. Parasites pre-treated with either anti-gp63 antibody or phospholipase C showed a significant reduction in the adhesion to M. scalaris and A. aegypti guts. Similarly, the pre-treatment of the explanted guts with purified gp63 diminished the interaction process. Collectively, these results corroborate the ubiquitous existence of gp63 homologues in insect trypanosomatids and the potential adhesion of these molecules to invertebrate host tissues.
Asunto(s)
Metaloendopeptidasas/fisiología , Péptido Hidrolasas/fisiología , Trypanosomatina/fisiología , Aedes/parasitología , Animales , Adhesión Celular/fisiología , Medios de Cultivo , Dípteros/parasitología , Citometría de Flujo/métodos , Interacciones Huésped-Parásitos , Insectos Vectores/parasitología , Intestinos/parasitología , Metaloendopeptidasas/metabolismo , Péptido Hidrolasas/metabolismo , Trypanosomatina/efectos de los fármacos , Trypanosomatina/metabolismo , Fosfolipasas de Tipo C/farmacologíaAsunto(s)
Adenoviridae/fisiología , Astroviridae/fisiología , Virus del Dengue/fisiología , Rotavirus/fisiología , Adenoviridae/genética , Proteínas E1B de Adenovirus/genética , Proteínas E4 de Adenovirus/genética , Humanos , Biología Molecular , Péptido Hidrolasas/fisiología , ARN Mensajero/biosíntesis , Proteínas Virales/fisiologíaRESUMEN
Estrous behavior induced by progesterone (P) treatment of estradiol-primed rats is followed by a period in which females do not respond behaviorally to a second administration of P [sequential inhibition (SI)]. SI is thought to involve P-dependent down-regulation of hypothalamic P receptor (PR) content. This study tested the hypothesis that the 26S proteasome participates in the regulation of SI and brain PR content in female rats. Ovariectomized, estrogen-primed (estradiol benzoate, 2 microg s.c.) adult rats were injected with P (1 mg s.c.) alone or P with the proteasome inhibitors Z-Ile-Glu (OBu(1))-Ala-Leu-H (PSI, 300 microg/100 g s.c.) or N alpha-tosyl-lysyl chloromethyl ketone (TLCK, 200 microg i.p.) administered 48 h after estradiol priming. Sexual behavior was assessed in all animals 4 h later. These two agents inhibit 26S proteasome-mediated protein degradation by different mechanisms. To explore SI, the animals received a second P injection 24 h after the first, and a second sexual behavior test was performed 4 h later. After this test, brains were excised, and proteins were extracted from the preoptic area and the hypothalamus and processed for semiquantitative immunoblotting. In the first sexual behavior test (facilitation test), all animals treated with estradiol + P exhibited intense lordosis behavior. In the second sexual behavior test (inhibition test), both lordosis and proceptivity were significantly reduced in response to the second administration of P (SI). The magnitude of SI was significantly attenuated by the administration of either PSI or TLCK concurrently with the first P injection. The first P injection reduced PR content in the hypothalamus but not in the preoptic area. In contrast, PSI and TLCK significantly increased PR content in both structures. Our results suggest that PR degradation by the 26S proteasome participates in the expression of P-induced SI in female rats.
Asunto(s)
Ciclo Estral/fisiología , Péptido Hidrolasas/fisiología , Progesterona/farmacología , Complejo de la Endopetidasa Proteasomal , Receptores de Progesterona/metabolismo , Conducta Sexual/efectos de los fármacos , Animales , Inhibidores Enzimáticos/farmacología , Estradiol/farmacología , Femenino , Hipotálamo/química , Oligopéptidos/farmacología , Ovariectomía , Postura , Área Preóptica/química , Inhibidores de Proteasas/farmacología , Ratas , Ratas Sprague-Dawley , Conducta Sexual/fisiología , Clorometilcetona Tosilisina/farmacologíaRESUMEN
AIMS: To study the influence of peptides and proteolytic enzymes in the osmotic adaptation of Lactobacillus casei. METHODS AND RESULTS: Di- and tri-peptides added individually increased the osmotolerance of Lact. casei when grown in a chemically defined medium (CDM) containing NaCl. Growth stimulation and the re-establishment in their presence of plasmid DNA supercoiling (recovery of the linking number) in hyperosmotic medium indicated that they are used as osmocompatible solutes as carnithine a known osmoprotector does. The investigation of the proteolytic system showed that in high osmolarity medium, the cell envelope-associated proteinase (PrtP), and PepX (X-prolyl-dipeptidyl aminopeptidase) increased activity and lost repression by peptides. PepI, an iminopeptidase was also derepressed. PepQ, a prolidase that specifically liberated proline from dipeptides, was almost unaffected. Derepression in the presence of peptides took place at the transcriptional level. However, the twofold activation of PrtP in CDM hyperosmotic medium was essentially through an increase of the apparent Vmax of the enzyme. CONCLUSIONS: These results strongly suggest a contribution of the proteolytic system peptide supply in the osmotic adaptation. SIGNIFICANCE AND IMPACT OF THE STUDY: Advances in understanding the role of peptides in the adaptation to high osmolarity particularly involved in dairy processes.