RESUMEN
High levels of D-amino acid oxidase (DAO) are associated with neurological and psychiatric disorders, while L-amino acid oxidase (LAO) exhibits antimicrobial and antitumor properties. The enzymatic conversion of the non-fluorescent kynurenine (KYN) into the endogenous weak fluorescent kynurenic acid (KYNA) by the action of DAO has previously been reported. However, the fluorescence of KYNA can be improved by changing the substituents on the aromatic rings. In this study, we prepared different 6-phenyl-substituted KYNA derivatives and investigated their fluorescence properties. Among them, 2-MePh-KYNA showed the maximum fluorescence quantum yield of 0.881 at 340 nm excitation and 418 nm emission wavelengths. The effects of solvent properties (dielectric constant, pKa, viscosity, and proticity) on the fluorescence intensity (FLI) of the KYNA derivatives were explored. The FLI of 2-MePh-KYNA was significantly large in protic solvents. Subsequently, 2-MePh-D-KYN and 2-MePh-L-KYN were prepared with high enantiopurity (>99.25%) for the enzymatic conversion. 2-MePh-D-KYN exhibited high sensitivity (â¼19 times that of a commercial DAO substrate and â¼60 times that of the previously reported MeS-D-KYN) and high selectivity, as it was not cross-reactive towards LAO, while 2-MePh-L-KYN was also converted into 2-MePh-KYNA by LAO. Furthermore, the 2-MePh-D-KYN probe successfully detected DAO in eel liver, kidney, and heparin-anticoagulated plasma in the in vitro study.
Asunto(s)
D-Aminoácido Oxidasa , Ácido Quinurénico , L-Aminoácido Oxidasa , Ácido Quinurénico/química , Colorantes Fluorescentes , Anguilas , Animales , L-Aminoácido Oxidasa/análisis , D-Aminoácido Oxidasa/análisis , Bioensayo , Fluorescencia , Cinética , Hígado/enzimología , Riñón/enzimologíaRESUMEN
Snakebite envenomation is a serious neglected tropical disease, and its management is often complicated by the diversity of snake venoms. In Asia, pit vipers of the Ovophis species complex are medically important venomous snakes whose venom properties have not been investigated in depth. This study characterized the venom proteomes of Ovophis convictus (West Malaysia), Ovophis tonkinensis (northern Vietnam, southern China), and Ovophis okinavensis (Okinawa, Japan) by applying liquid chromatography-tandem mass spectrometry, which detected a high abundance of snake venom serine proteases (SVSP, constituting 40-60% of total venom proteins), followed by phospholipases A2, snake venom metalloproteinases of mainly P-III class, L-amino acid oxidases, and toxins from other protein families which were less abundant. The venoms exhibited different procoagulant activities in human plasma, with potency decreasing from O. tonkinensis > O. okinavensis > O. convictus. The procoagulant nature of venom confirms that consumptive coagulopathy underlies the pathophysiology of Ovophis pit viper envenomation. The hetero-specific antivenoms Gloydius brevicaudus monovalent antivenom (GbMAV) and Trimeresurus albolabris monovalent antivenom (TaMAV) were immunoreactive toward the venoms, and cross-neutralized their procoagulant activities, albeit at variably limited efficacy. In the absence of species-specific antivenom, these hetero-specific antivenoms may be useful in treating coagulotoxic envenomation caused by the different snakes in their respective regions.
Asunto(s)
Crotalinae , Proteoma , Proteínas de Reptiles , Venenos de Víboras , Animales , Antivenenos/inmunología , Coagulantes/análisis , Coagulantes/inmunología , Coagulantes/toxicidad , Humanos , L-Aminoácido Oxidasa/análisis , L-Aminoácido Oxidasa/inmunología , L-Aminoácido Oxidasa/toxicidad , Metaloproteasas/análisis , Metaloproteasas/inmunología , Metaloproteasas/toxicidad , Fosfolipasas A2/análisis , Fosfolipasas A2/inmunología , Fosfolipasas A2/toxicidad , Plasma/efectos de los fármacos , Proteoma/análisis , Proteoma/inmunología , Proteoma/toxicidad , Proteómica , Proteínas de Reptiles/análisis , Proteínas de Reptiles/inmunología , Proteínas de Reptiles/toxicidad , Serina Proteasas/análisis , Serina Proteasas/inmunología , Serina Proteasas/toxicidad , Venenos de Víboras/química , Venenos de Víboras/inmunología , Venenos de Víboras/toxicidadRESUMEN
Trimeresurus nebularis is a montane pit viper that causes bites and envenomation to various communities in the central highland region of Malaysia, in particular Cameron's Highlands. To unravel the venom composition of this species, the venom proteins were digested by trypsin and subjected to nano-liquid chromatography-tandem mass spectrometry (LC-MS/MS) for proteomic profiling. Snake venom metalloproteinases (SVMP) dominated the venom proteome by 48.42% of total venom proteins, with a characteristic distribution of P-III: P-II classes in a ratio of 2:1, while P-I class was undetected. Snaclecs constituted the second most venomous protein family (19.43%), followed by snake venom serine proteases (SVSP, 14.27%), phospholipases A2 (5.40%), disintegrins (5.26%) and minor proteins including cysteine-rich secretory proteins, L-amino acid oxidases, phosphodiesterases, 5'-nucleotidases. The venomic profile correlates with local (painful progressive edema) and systemic (hemorrhage, coagulopathy, thrombocytopenia) manifestation of T. nebularis envenoming. As specific antivenom is unavailable for T. nebularis, the hetero-specific Thai Green Pit viper Monovalent Antivenom (GPVAV) was examined for immunological cross-reactivity. GPVAV exhibited good immunoreactivity to T. nebularis venom and the antivenom effectively cross-neutralized the hemotoxic and lethal effects of T. nebularis (lethality neutralizing potency = 1.6 mg venom per mL antivenom). The findings supported GPVAV use in treating T. nebularis envenoming.
Asunto(s)
Venenos de Crotálidos/química , Venenos de Crotálidos/toxicidad , Trimeresurus , Animales , Antivenenos/farmacología , Desintegrinas/análisis , Femenino , Hidrolasas/análisis , L-Aminoácido Oxidasa/análisis , Malasia , Masculino , Ratones Endogámicos ICR , Proteoma , Proteínas de Reptiles/análisisRESUMEN
BACKGROUND: Naja annulifera is a medically important venomous snake occurring in some of the countries in Sub-Saharan Africa. Accidental bites result in severe coagulation disturbances, systemic inflammation and heart damage, as reported in dogs, and death, by respiratory arrest, in humans. Despite the medical importance of N. annulifera, little is known about its venom composition and the pathogenesis of envenomation. In this paper, the toxic, inflammatory and immunogenic properties of N. annulifera venom were analyzed. METHODOLOGY/PRINCIPAL FINDINGS: Venom proteomic analysis identified 79 different proteins, including Three Finger Toxins, Cysteine Rich Secretory Proteins, Metalloproteinases, Phospholipases A2 (PLA2), Hyaluronidase, L-amino-acid oxidase, Cobra Venom Factor and Serine Proteinase. The presence of PLA2, hyaluronidase, fibrinogenolytic and anticoagulant activities was detected using functional assays. The venom was cytotoxic to human keratinocytes. In an experimental murine model of envenomation, it was found that the venom induced local changes, such as swelling, which was controlled by anti-inflammatory drugs. Moreover, the venom caused death, which was preceded by systemic inflammation and pulmonary hemorrhage. The venom was shown to be immunogenic, inducing a strong humoral immune response, with the production of antibodies able to recognize venom components with high molecular weight and to neutralize its lethal activity. CONCLUSIONS/SIGNIFICANCE: The results obtained in this study demonstrate that N. annulifera venom contains toxins able to induce local and systemic inflammation, which can contribute to lung damage and death. Moreover, the venom is immunogenic, an important feature that must be considered during the production of a therapeutic anti-N. annulifera antivenom.
Asunto(s)
Venenos Elapídicos/análisis , Venenos Elapídicos/toxicidad , Animales , Antivenenos/farmacología , Femenino , Hialuronoglucosaminidasa/análisis , L-Aminoácido Oxidasa/análisis , Masculino , Metaloproteasas/análisis , Ratones , Ratones Endogámicos BALB C , Naja , Fosfolipasas A2/análisis , Proteómica , Serina Proteasas/análisisRESUMEN
In order to determine Bothriopsis bilineata smaragdina venom (BbsV) composition, proteomic approaches were performed. Venom components were analyzed by RP-HPLC, SDS- PAGE and nano LC on line with LTQ Orbitrap XL. Results showed a total of 189 identified proteins, grouped into 11 different subgroups, which include snake venom metalloproteinases (SVMPs, 54.67%), snake C-type lectins (Snaclecs, 15.78%), snake venom serine proteinases (SVSPs, 14.69%), cystein-rich secretory proteins (CRISP, 2.61%), phospholipases A2 (PLA2, 1.14%), phosphodiesterase (PDE, 1.17%), venom endothelial growth factor (VEGF, 1.06%) 5'nucleotidases (0.33%), L-amino acid oxidases (LAAOs, 0.28%) and other proteins. In vitro enzymatic activities (SVMP, SVSP, LAAO, Hyal and PLA2) of BbsV were also analyzed. BbsV showed high SVSP activity but low PLA2 activity, when compared to other Bothrops venoms. In vivo, BbsV induced hemorrhage and edema in mice and showed intraperitoneal median lethal dose (LD50) of 92.74 (± 0.15) µg/20â¯g of mice. Furthermore, BbsV reduced cell viability when incubated with VERO cells. Peruvian and Brazilian bothropic antivenoms recognize BbsV proteins, as detected by ELISA and Western Blotting. Both antivenoms were able to neutralize in vivo edema and hemorrhage. SIGNIFICANCE: In Peru, snakebite is a public health problem, especially in the rain forest, as a result of progressive colonization of this geographical area. This country is the second in Latin America, after Brazil, to exhibit the largest variety of venomous snakes. B. atrox and B. b. smaragdina snakes are sympatric species in Peruvian Amazon region and are responsible for approximately 95% of the envenomings reported in this region. B. b. smaragdina may cause a smaller share (3 to 38%) of those accidents, due to its arboreal habits, that make human encounters with these snakes less likely to happen. Despite B. b. smaragdina recognized medical importance, its venom composition and biological activities have been poorly studied. Furthermore, BbsV is not a component of the antigenic pool used to produce the corresponding Peruvian bothropic antivenom (P-BAV). Our results not only provide new insights on BbsV composition and biological activity, but also demonstrate that both P-BAV and B-BAV polyvalent antivenoms have a considerable recognition of proteins from BbsV and, more importantly, neutralized hemorrhage and edema, the main local effects of bothropic envenomation.
Asunto(s)
Antivenenos/análisis , Bothrops , Venenos de Crotálidos/inmunología , Venenos de Crotálidos/metabolismo , Venenos de Crotálidos/farmacología , Animales , Antivenenos/metabolismo , Chlorocebus aethiops , Venenos de Crotálidos/análisis , Femenino , Hemorragia/inducido químicamente , Hemorragia/patología , L-Aminoácido Oxidasa/análisis , L-Aminoácido Oxidasa/metabolismo , Dosificación Letal Mediana , Metaloproteasas/análisis , Metaloproteasas/metabolismo , Ratones , Perú , Fosfolipasas A2/análisis , Fosfolipasas A2/metabolismo , Proteoma/análisis , Proteoma/metabolismo , Proteómica , Serina Proteasas/análisis , Serina Proteasas/metabolismo , Células VeroRESUMEN
UNLABELLED: The venom of Malayan pit viper (Calloselasma rhodostoma) is highly toxic but also valuable in drug discovery. However, a comprehensive proteome of the venom that details its toxin composition and abundance is lacking. This study aimed to unravel the venom complexity through a multi-step venomic approach. At least 96 distinct proteins (29 basic, 67 acidic) in 11 families were identified from the venom. The venom consists of mainly snake venom metalloproteinases (SVMP, 41.17% of total venom proteins), within which the P-I (kistomin, 20.4%) and P-II (rhodostoxin, 19.8%) classes predominate. This is followed by C-type lectins (snaclec, 26.3%), snake venom serine protease (SVSP, 14.9%), L-amino acid oxidase (7.0%), phospholipase A2 (4.4%), cysteine-rich secretory protein (2.5%), and five minor toxins (nerve growth factor, neurotrophin, phospholipase B, 5' nucleotidase and phosphodiesterase, totaling 2.6%) not reported in the proteome hitherto. Importantly, all principal hemotoxins unveiled correlate with the syndrome: SVSP ancrod causes venom-induced consumptive coagulopathy, aggravated by thrombocytopenia caused by snaclec rhodocytin, a platelet aggregation inducer, while P-II rhodostoxin mediates hemorrhage, exacerbated by P-I kistomin and snaclec rhodocetin that inhibit platelet plug formation. These toxins exist in multiple isoforms and/or complex subunits, deserving further characterization for the development of an effective, polyspecific regional antivenom. BIOLOGICAL SIGNIFICANCE: Advents in proteomics and bioinformatics have vigorously propelled the scientific discoveries of toxins from various lineages of venomous snakes. The Malayan pit viper, Calloselasma rhodostoma, is a medically important species in Southeast Asia as its bite can cause envenomation, while the venom is also a source of bioactive compounds for drug discovery. Detailed profiling of the venom, however, is inadequate possibly due to the complex nature of the venom and technical limitation in separating the constituents into details. Integrating a multi-step fractionation method, this study successfully revealed a comprehensive and quantitative profile of the composition of the venom of this medically important venomous snake. The relative abundance of the various venom proteins is determined in a global profile, providing useful information for understanding the pathogenic roles of the different toxins in C. rhodostoma envenomation. Notably, the principal hemotoxins were identified in great details, including the variety of toxin subunits and isoforms. The findings indicate that these toxins are the principal targets for effective antivenom neutralization, and should be addressed in the production of a pan-regional polyspecific antivenom. In addition, minor toxin components not reported previously in the venom were also detected in this study, enriching the current toxin database for the venomous snakes.
Asunto(s)
Crotalinae , Proteoma/análisis , Venenos de Víboras/química , Animales , Hemolíticos/análisis , L-Aminoácido Oxidasa/análisis , Lectinas Tipo C/análisis , Metaloproteasas/análisis , Fosfolipasas A2/análisis , Serina Proteasas/análisis , Venenos de Víboras/enzimologíaRESUMEN
L-amino acid oxidase (LAAO) is widely distributed in many different organisms and found to play important biological roles, thus attracting a great deal of attention for characterization of its activity. Diverse detection methods with their own properties have been established. This review advanced different LAAO activity assays based on substrate consumption, cofactor amount, and product accumulation. The description of benefits and drawbacks of each method is expected to help researchers find appropriate detection method of LAAO activity for their own purpose.
Asunto(s)
L-Aminoácido Oxidasa/análisis , Animales , Coenzimas/análisis , Coenzimas/química , L-Aminoácido Oxidasa/química , Especificidad por Sustrato/fisiologíaRESUMEN
The L-amino acid oxidases (LAAOs) constitute a major component of snake venoms and have been widely studied due to their widespread presence and various effects, such as apoptosis induction, cytotoxicity, induction and/or inhibition of platelet aggregation, hemorrhage, hemolysis, edema, as well as antimicrobial, antiparasitic and anti-HIV activities. The isolated and characterized snake venom LAAOs have become important research targets due to their potential biotechnological applications in pursuit for new drugs of interest in the scientific and medical fields. The current study discusses the antitumor effects of snake venom LAAOs described in the literature to date, highlighting the mechanisms of apoptosis induction proposed for this class of proteins.
Asunto(s)
Animales , L-Aminoácido Oxidasa/análisis , Oxidorreductasas/análisis , Venenos/administración & dosificación , Serpientes/clasificaciónRESUMEN
The L-amino acid oxidases (LAAOs) constitute a major component of snake venoms and have been widely studied due to their widespread presence and various effects, such as apoptosis induction, cytotoxicity, induction and/or inhibition of platelet aggregation, hemorrhage, hemolysis, edema, as well as antimicrobial, antiparasitic and anti-HIV activities. The isolated and characterized snake venom LAAOs have become important research targets due to their potential biotechnological applications in pursuit for new drugs of interest in the scientific and medical fields. The current study discusses the antitumor effects of snake venom LAAOs described in the literature to date, highlighting the mechanisms of apoptosis induction proposed for this class of proteins.
Asunto(s)
Animales , L-Aminoácido Oxidasa/análisis , Oxidorreductasas/análisis , Venenos/administración & dosificación , Serpientes/clasificaciónRESUMEN
Venoms of the redtail coral snake Micrurus mipartitus from Colombia and Costa Rica were analyzed by "venomics", a proteomic strategy to determine their composition. Proteins were separated by RP-HPLC, followed by SDS-PAGE, in-gel tryptic digestion, identification by MALDI or ESI tandem mass spectrometry, and assignment to known protein families by similarity. These analyses were complemented with a characterization of venom activities in vitro and in vivo. Proteins belonging to seven families were found in Colombian M. mipartitus venom, including abundant three-finger toxins (3FTx; ~60% of total proteins) and phospholipases A(2) (PLA(2); ~30%), with the remaining ~10% distributed among l-amino acid oxidase, P-III metalloproteinase, Kunitz-type inhibitor, serine proteinase, and C-type lectin-like families. The venoms of two M. mipartitus specimens from Costa Rica, also referred to as M. multifasciatus in some taxonomic classifications, were also analyzed. Both samples were highly similar to each other, and partially resembled the chromatographic and identity profiles of M. mipartitus from Colombia, although presenting a markedly higher proportion of 3FTxs (~83.0%) in relation to PLA(2)s (~8.2%), and a small amount of acetylcholinesterase, not detected in the venom from Colombia. An equine antivenom against the Central American coral snake, M. nigrocinctus, did not recognize venom components of M. mipartitus from Colombia or Costa Rica by enzyme-immunoassay. Four major components of Colombian M. mipartitus venom were isolated and partially characterized. Venomics of Micrurus species may provide a valuable platform for the rational design of immunizing cocktails to obtain polyspecific antivenoms for this highly diverse group of American elapids.
Asunto(s)
Venenos Elapídicos/química , Venenos Elapídicos/farmacología , Secuencia de Aminoácidos , Animales , Antivenenos/análisis , Cromatografía Líquida de Alta Presión , Venenos Elapídicos/metabolismo , Elapidae , L-Aminoácido Oxidasa/análisis , Ratones , Fosfolipasas A2/análisis , Proteoma/análisisRESUMEN
A simple and practicable method for in-gel detection of bacterial produced L-amino acid oxidases (L-AAOs) after non-denaturing SDS-PAGE based on the visualisation of occurring hydrogen peroxide production is described. Advantages of this screening method for L-AAOs are the determination of their numbers and approximate molecular weights additionally in one approach.
Asunto(s)
Bacterias/enzimología , Electroforesis en Gel de Poliacrilamida/métodos , Peróxido de Hidrógeno/metabolismo , L-Aminoácido Oxidasa/análisis , Aminoácidos/metabolismo , L-Aminoácido Oxidasa/aislamiento & purificación , Oxidación-ReducciónRESUMEN
A new strategy for the enantioseparation of D,L-amino acids employing the principle of ligand exchange capillary electrophoresis with Zn(II)-L-valine complex as a chiral selecting system in the presence of beta-cyclodextrin has been designed. Successful enantioseparation of label free and labeled amino acids have been achieved with a buffer of 100.0mM boric acid, 5.0mM ammonium acetate, 4.0mM beta-cyclodextrin, 4.0mM ZnSO(4) and 8.0mM L-valine at pH 8.1. This new method was shown to be applicable to the quantitative analysis of label free D- and L-aromatic amino acids. Furthermore, the expanding enzymatic use of L-amino acid oxidase to incubate with different L-amino acids has allowed understanding of the substrate's specificity. An on-column incubation assay has been developed to study the L-amino acid oxidase's catalytic efficiency. It was demonstrated that the enzyme kinetic constant could be determined by using this new method.
Asunto(s)
Bioquímica/métodos , Técnicas de Química Analítica , Electroforesis Capilar/métodos , L-Aminoácido Oxidasa/análisis , Valina/química , Zinc/química , Animales , Catálisis , Electroforesis Capilar/instrumentación , Concentración de Iones de Hidrógeno , Cetoácidos/química , Cinética , Ligandos , Venenos de Serpiente/enzimología , Serpientes , Estereoisomerismo , Especificidad por SustratoRESUMEN
Vipera palaestinae (Werner, 1938) es una de las serpientes venenosas presentes en Oriente Próximo, incluido el sur del Líbano donde las Fuerzas Armadas españolas tienen desplegadas tropas que forman parte de la Fuerza Interina de las Naciones Unidas en el Líbano (UNIFIL) en la Operación Libre Hidalgo (L/H). En el presente trabajo se revisan los componentes del veneno de esta serpiente, así como la clínica y el tratamiento de los efectos tóxicos causados por su mordedura venenosa (AU)
Vipera palaestinae (Werner, 1938) is one of the dangerous venomous snakes present in the Middle East, including the south of Lebanon where the Spanish Armed Forces have deployed troops as part of the United Nations Interim Force in Lebanon (UNIFIL) in Operation Libre Hidalgo (L/H). This article reviews the different components present in the venom, as well as the clinical signs and treatment of the toxicological effects caused by its venomous bite (AU)
Asunto(s)
Humanos , Viperidae , Mordeduras de Serpientes/tratamiento farmacológico , Venenos de Serpiente/envenenamiento , Personal Militar , L-Aminoácido Oxidasa/análisis , Fosfolipasas A/análisis , Metaloproteínas/análisis , Hialuronoglucosaminidasa/análisis , Antídotos/administración & dosificaciónRESUMEN
We report the comparative proteomic characterization of the venoms of two related neotropical arboreal pitvipers from Costa Rica of the genus Bothriechis, B. lateralis (side-striped palm pit viper) and B. schlegelii (eyelash pit viper). The crude venoms were fractionated by reverse-phase HPLC, followed by analysis of each chromatographic fraction by SDS-PAGE, N-terminal sequencing, MALDI-TOF mass fingerprinting, and collision-induced dissociation tandem mass spectrometry of tryptic peptides. The venom proteomes of B. lateralis and B. schlegelii comprise similar number of distinct proteins belonging, respectively, to 8 and 7 protein families. The two Bothriechis venoms contain bradykinin-potentiating peptides (BPPs), and proteins from the phospholipase A 2 (PLA 2), serine proteinase, l-amino acid oxidase (LAO), cysteine-rich secretory protein (CRISP), and Zn (2+)-dependent metalloproteinase (SVMP) families, albeit each species exhibit different relative abundances. Each venom also contains unique components, for example, snake venom vascular endothelial growth factor (svVEGF) and C-type lectin-like molecules in B. lateralis, and Kazal-type serine proteinase inhibitor-like proteins in B. schlegelii. Using a similarity coefficient, we estimate that the similarity of the venom proteins between the two Bothriechis taxa may be <10%, indicating a high divergence in their venom compositions, in spite of the fact that both species have evolved to adapt to arboreal habits. The major toxin families of B. lateralis and B. schlegelii are SVMP (55% of the total venom proteins) and PLA 2 (44%), respectively. Their different venom toxin compositions provide clues for rationalizing the distinct signs of envenomation caused by B. schlegelii and B. lateralis. An antivenomic study of the immunoreactivity of the Instituto Clodomiro Picado (ICP) polyvalent antivenom toward Bothriechis venoms revealed that l-amino acid oxidase and SVMPs represent the major antigenic protein species in both venoms. Our results provide a ground for rationalizing the reported protection of the ICP polyvalent antivenom against the hemorrhagic, coagulant, defibrinating, caseinolytic and fibrin(ogen)olytic activities of Bothriechis ( schlegelii, lateralis) venoms. However, these analyses also evidenced the limited recognition capability of the polyvalent antivenom toward a number of Bothriechis venom components, predominantly BPPs, svVEGF, Kazal-type inhibitors, some PLA 2 proteins, some serine proteinases, and CRISP molecules.
Asunto(s)
Antivenenos/análisis , Venenos de Crotálidos/metabolismo , Proteoma/análisis , Viperidae/metabolismo , Animales , Antivenenos/inmunología , Cromatografía Líquida de Alta Presión , Venenos de Crotálidos/química , Venenos de Crotálidos/inmunología , Electroforesis en Gel de Poliacrilamida , L-Aminoácido Oxidasa/análisis , L-Aminoácido Oxidasa/inmunología , Lectinas Tipo C/análisis , Metaloproteasas/análisis , Metaloproteasas/inmunología , Oligopéptidos/análisis , Fosfolipasas A2 Secretoras/análisis , Proteoma/inmunología , Serina Endopeptidasas/análisis , Inhibidores de Serina Proteinasa/análisis , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Espectrometría de Masas en Tándem , Factor A de Crecimiento Endotelial Vascular/análisisRESUMEN
To better understand the effects of antiandrogens on the prostate, we investigated the changes in the proteome of rat ventral prostate (VP) following treatment with a well characterized 5alpha-reductase inhibitor, finasteride. Sprague-Dawley rats were treated daily by gavage with finasteride at 0, 1, 5, 25, and 125 mg/kg/day. Changes in plasma hormone levels as well as the weight and histology of sex accessory tissues were determined after 28 days of treatment and showed a dose-related decrease of VP weights together with a marked atrophy of the tissue visible at the macroscopic and microscopic levels. In addition, significant reductions in seminal vesicle and epididymis weights were noted. VP proteins were analyzed by two-dimensional gel electrophoresis: 37 proteins, mainly involved in protein synthesis, processing, and cellular trafficking and in metabolism, detoxification, and oxidative stress, were identified as modulated by finasteride. The prominent feature of this study is the demonstration of finasteride dose-dependent up-regulation of a protein similar to l-amino-acid oxidase 1 (Lao1). An up-regulation of this protein was also observed with the antiandrogen flutamide. Lao1 expression occurred as early as 48 h after antiandrogen administration and persisted throughout the treatment duration. Immunohistochemistry showed that this protein was only detectable in epithelial cells and secretory vesicles. Altogether these data point to a potential use of Lao1 to reveal antiandrogen-induced prostate injury.