Neotropical Rattlesnake (Crotalus simus) Venom Pharmacokinetics in Lymph and Blood Using an Ovine Model.

The most abundant protein families in viper venoms are Snake Venom Metalloproteases (SVMPs), Snake Venom Serine Proteases (SVSPs) and Phospholipases (PLA2s). These are primarily responsible for the pathophysiology caused by the bite of pit-vipers; however, there are few studies that analyze the pharmacokinetics (PK) of whole venom (WV) and its protein families. We studied the pathophysiology, PK profile and differential absorption of representative toxins from venom of Neotropical Rattlesnake (Crotalus simus) in a large animal model (ovine). Toxins studied included crotoxin (the main lethal component), which causes moderate to severe neurotoxicity; SVSPs, which deplete fibrinogen; and SVMPs, which cause local tissue damage and local and systemic hemorrhage. We found that Whole Venom (WV) was highly bioavailable (86%) 60 h following intramuscular (IM) injection, and extrapolation suggests that bioavailability may be as high as 92%. PK profiles of individual toxins were consistent with their physicochemical properties and expected clinical effects. Lymph cannulated animals absorbed 1.9% of WV through lymph during the first 12 h. Crotoxin was minimally detectable in serum after intravenous (IV) injection; however, following IM injection it was detected in lymph but not in blood. This suggests that crotoxin is quickly released from the blood toward its tissue targets.

Harnessing the Activity of the Fungal Metalloprotease, Mpr1, To Promote Crossing of Nanocarriers through the Blood-Brain Barrier.

Cryptococcus neoformans (Cn) is the leading cause of fungal meningitis primarily in immunosuppressed patients. Cn invades the central nervous system by overcoming the highly restricted blood-brain barrier (BBB). We previously determined that a secreted fungal metalloprotease, Mpr1, that also confers crossing ability to yeast upon CnMPR1 expression in Saccharomyces cerevisiae is central to this process. This led us to question whether Mpr1 could be engineered to function as part of a nanocarrier delivery vehicle. Here, a eukaryotic expression system produced proteolytically active Mpr1 recombinant protein that was successfully conjugated to functionalized quantum dot (QD) nanoparticles and readily internalized by brain microvascular endothelial cells. An in vitro BBB model showed QD-Mpr1 crossed the BBB significantly better than mock QD, and QD-Mpr1 did not damage BBB integrity. Internalization of QD-Mpr1 occurred by membrane invaginations and endocytic pits typical of receptor-mediated endocytosis involving clathrin-coated entry points. This study substantiates the notion that fungal mechanisms of BBB entry may be harnessed for new drug delivery platform technologies.

La colagenasa y el tejido desvitalizado en el contexto de la preparación del lecho de la herida / Collagenase and devitalized tissue in the framework of the wound bed preparation

Se analiza la función de la colagenasa como un recurso imprescindible para el abordaje y control del tejido desvitalizado en el contexto de la preparación del lecho de la herida. Se realiza un análisis y discusión de los aspectos fisiopatológicos de la colagenasa mediante una revisión bibliográfica (AU)

We analize the role of collagenase, as an indispensable resource for the management and control of the devitalized tissue in the framework of the the wound bed preparation. An analysis and discussion of the physiopathological aspects of collagenase has been performed through a bibliographic review (AU)

Expresión de metaloproteasa de matriz 9 en el cáncer de próstata. Experiencia preliminar / Expression of matrix metalloproteinase-9 in prostate cancer. Preliminary experience

OBJETIVO: Estudiar la validez de la metaloproteasa 9 (MMP-9) como marcador complementario al PSA en el diagnóstico y el pronóstico del carcinoma de próstata.MÉTODO: Estudio prospectivo estructurado como cohorte de base hospitalaria. Fueron incluidos 100 pacientes consecutivos a los que se iba a practicar una biopsia prostática. La determinación sérica de MMP-9 se realizó mediante inmunoensayo, y el análisis estadístico con el programa informático stata/SE 8.2.RESULTADOS: 32 pacientes fueron diagnosticados de carcinoma prostático y el 52% de ellos con grado Gleason mayor o igual a 7. Los valores de MMP-9 sérica oscilaron entre 225,7 y 1932,3 nanogramos por mililitro, sin encontrar diferencias estadísticamente significativas entre los pacientes con histología benigna, maligna e incierta (p=0,429). Las diferencias se acercaron a la significación estadística en el subgrupo de pacientes con PSA 4-10 ng/ml (p=0,058) y en el subgrupo PSA libre/total menor de 15% se observaron diferencias significativas (p=0,037). No se encontró relación entre el grado Gleason y el nivel de MMP-9 (p=0,739). Los niveles de PSA y MMP-9 demostraron ser independientes (Coeficiente de correlación de Pearson -0,1).CONCLUSIONES: No fue posible demostrar la eficacia de la MMP-9 para predecir el resultado de la biopsia. En el grupo de pacientes con elevaciones discretas del PSA (entre 4 y 10 ng/ml) todas las variables descriptivas fueron superiores en el grupo con histología maligna, sin alcanzar la significación estadística. Sí se alcanzó la significación cuando el cociente de PSA libre entre PSA total fue menor del 15%, pero este hallazgo no tiene relevancia en la práctica clínica, pues estos pacientes ya tienen indicación clara de biopsia. Tampoco se demuestra relación con el pronóstico al no existir diferencias de expresión de MMP-9 entre diferentes grados Gleason(AU)

OBJECTIVES: To study the validity of Matrix Metalloproteinase 9 as a complementary marker to PSA for the diagnosis and prognosis of Prostate Cancer.METHODS: Prospective study structured as a hospital-based cohort of 100 consecutive patients undergoing prostate biopsy. Serum determination of MMP-9 was carried out by means of inmunoassay . Statistical analysis was performed using the Stata/SE 8.2 software.RESULTS: 32 patients were diagnosed with prostate cancer and 52% had a Gleason score equal to or greater than 7. The values of serum MMP-9 varied between 225.7 and 1932.3 ng/ml, without significant differences among patients with benign, malignant and uncertain histology (p=0.429). The differences approached statistical significance in the subgroup of patients with PSA at 4-10 ng/ml (p=0.058), and significant differences were observed in the subgroup with free PSA to total PSA coefficient of less than 15% (p=0.037). No relationship between the Gleason score and the level of MMP-9 was shown (p=0.739). The levels of PSA and MMP-9 were shown to be independent (Pearson coefficient of correlation -0.1).CONCLUSIONS: It was not possible to show the efficacy of MMP-9 in predicting the result of the biopsy. In the group of patients with slightly increased levels of PSA (between 4 and 10 ng/ml) all the descriptive variables were higher in the group with malignant histology, though they did not reach statistical significance, they did reach significance when the coefficient of free PSA over total PSA was less than 15%, but this finding is not relevant clinically, as these patients already have a clear indication for biopsy. Neither was the relationship with the prognosis shown as there are no differences of MMP-9 expression at varying Gleason scores(AU)

Metaloproteinasas, matriz extracelular y cáncer / Metalloproteinases, extracellular matrix and cancer

Las metaloproteinasas de la matriz extracelular (MMP) estánimplicadas en procesos fisiológicos y patológicos e intervienen en larotura de la matriz extracelular (ECM). Las MMP constituyen unafamilia de endopeptidasas neutras dependientes de zinc, capaces dedegradar los componentes esenciales de la matriz. Los inhibidorestisulares endógenos de las MMP (TIMP), una clase de inhibidores delas MMP, reducen la degradación proteolítica excesiva de la ECM.La degradación de la ECM es crucial para el crecimiento tumoralmaligno, invasión, metástasis y angiogénesis. Se han descrito cambiosen las MMP y sus inhibidores durante la carcinogénesis y tambiénque unas y otros regulan las vías señalizadoras mediante larotura de otros sustratos que los de la matriz, tales como citoquinas,quimioquinas y factores de crecimiento. Como ciertas MMP limitanel crecimiento tumoral, su identificación e intervención terapéuticaen combinación con la quimioterapia convencional ha de proporcionarun medio para la terapia del cáncer(AU)

The extracellular matrix metalloproteinases (MMP) are involved in physiological and pathological processes, through the cleavage ofextracellular matrix (ECM) and non-matrix substrates. MMP are afamily of zinc-dependent neutral endopeptidases capable of degradingessentially all matrix components. Endogenous tissue inhibitors ofmetalloproteinases (TIMP), one kind of MMP inhibitors, reduce theexcessive proteolytic ECM degradation. Degradation of ECM is crucialfor malignant tumor growth, invasion, metastasis and angiogenesis.A variety of reports describe the correlated changes in MMP and TIMPduring the formation of cancer, and also that MMP and TIMP may actas regulators of signaling pathways through the cleavage of nonmatrixsubstrates, including cytokines, chemokines, and growthfactors. As certain MMP limit tumor growth, identification of properMMP in combination with conventional chemotherapy is expected toprovide a feasible approach for cancer therapy(AU)

Cinética de la metaloproteasa 9: importancia del ritmo luz-oscuridad de la metaloproteasa 9 en el síndrome coronario agudo / The kinetics of metalloproteinase-9: the significance of the light-dark cycle in metalloproteinase-9 in acute coronary syndrome

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Cinética de la metaloproteasa 9: importancia del ritmo luz-oscuridad de la metaloproteasa 9 en el síndrome coronario agudo. Respuesta / The kinetics of metalloproteinase-9: the significance of the light-dark cycle in metalloproteinase-9 in acute coronary syndrome. Response

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Are adrenomedullin positive modulators novel matrix metalloproteinase inhibitors?

Las metaloproteasas de la matriz (MMPs) pertenecen a la familia de enzimasque contienen zinc y juegan un papel predominante en la degradación del tejidoconectivo. Por ello se consideran dianas terapéuticas para procesos de inflamacióny enfermedades malignas y degenerativas. Por otro lado, se ha demostrado recientementeque un miembro de esta familia, MMP-2, una colagenasa de tipo IV tambiénconocida como gelatinasa A, es capaz de degradar un péptido angiogénico denominadoadrenomedulina (AM) (1). AM es una hormona peptídica que desarrolla unpapel importante en diversas patologías como diabetes, hipertensión y cáncer. Seha identificado mediante un cribado de alto rendimiento (HTS) de la colección decompuestos del Instituto Nacional del Cáncer (NCI), una serie de moduladores coninteresante actividad hipotensora (2). El mecanismo de acción de estos moduladoreses desconocido y nosotros proponemos que pueden afectar a la biodisponibilidadde la AM en el torrente sanguíneo por medio de la inhibición de la actividad dela MMP-2. En este trabajo presentamos un estudio teórico que hace uso de técnicascomo mecánica molecular, docking y Cribado Virtual con el objetivo de demostraresa hipótesis. A continuación del estudio computacional se llevó a cabo la evaluaciónbiológica de algunos compuestos, permitiéndonos proponer un nuevo tipo deZBG que puede ser interesante para el diseño de nuevos inhibidores de MMPS, coninterés como agentes anticancerosos y antiangiogénicos

Matrix metalloproteinases (MMPs), are a family of structurally related zinccontaining enzymes that play a major role in the breakdown of connective tissueand therefore, are targets for therapeutic inhibitors in many inflammatory,malignant, and degenerative diseases. On the other hand, it has been recentlydemonstrated that one of these enzymes, MMP-2, a type IV collagenase, termedgelatinase A, cleaves the angiogenic peptide adrenomedullin (AM) (1). AM is apeptide hormone that plays a critical role in several diseases such as diabetes,hypertension and cancer. In a High Throughput Screening (HTS) carried out at theNational Cancer Institute (NCI), a series of AM modulators were identified, with aninteresting hypotensive activity (2). In order to shed light into the mechanism ofaction of these interesting compounds, we have hypothesized that they may be affecting the biodisponibility of AM in the blood stream by inhibiting the MMP-2protease activity. In the present work, we present a theoretical study, making useof molecular mechanics, docking and virtual screening techniques, with the aim ofdemonstrating this hypothesis. Biological evaluation of MMP-2 inhibition by someselected compounds, followed the computational work, leading us to propose astructurally new type of MMP-2 inhibitors, with possible interest as anticancer andantiangiogenic agents

Metaloproteinasas y cáncer vesical: estado actual / Metaloproteinases and bladder cancer: state of the art

Durante los últimos diez años el papel de las metaloproteinasas en el desarrollo tumoral haalcanzado una importante relevancia. El descubrimiento de nuevas metaloproteinasas ha demostrado, no sólo su implicación en la disrupción de la matriz extracellular y la consecuente diseminación tumoral sino que diversos estudios han desvelado su papel en procesos deiniciación, promoción tumoral y apoptosis.Recientes trabajos desvelan también la relación entre niveles elevados de ciertas metaloproteinasas y sus inhibidores, sobre todo MMP2, MMP9, 17MPl Y 17MP2 en tejidoparafinado, orina y suero de pacientes con carcinoma vesical con características clínicopatológicasdesfavorables en cuanto a estadio y grado, por lo que en un futuro podríanconvertirse en buenas predictoras del comportamiento del cáncer vesical

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Secretion of haemolysins and proteases by Aeromonas hydrophila EO63: separation and characterization of the serine protease (caseinase) and the metalloprotease (elastase).

AIMS: To determine the haemolysins and proteases excreted by the virulent strain EO63 of Aeromonas hydrophila grown in complex media and to then fractionate and characterize them, in particular those with elastolytic activity. METHODS AND RESULTS: The amount of haemolytic and proteolytic activity in EO63 culture supernatants was dependent on the culture media used. In all media, haemolysins appeared during the phase of active growth and haemolytic activity decreased quickly thereafter, as previously described for aerolysin. In contrast, proteases were mainly released during the stationary phase. Serine protease activity in EO63 culture supernatants was four times greater than that caused by metalloproteases. Two main proteases were partially purified from EO63 culture supernatants by isoelectrophoresis: a serine protease (68 kDa) active against casein; a mixture of different protein bands (60, 44 and 31 kDa) representing a thermostable metalloprotease active against elastin and casein. This metallo-elastase was also inhibited by dithiothreitol and showed a pH optimum of 8.0. Both exoenzymes were toxic for eels at LD50 doses of 1.1 and 3.5 microg (g fish)(-1), respectively. CONCLUSIONS: A serine caseinase and a metallo-elastase may play a role in the pathogenicity of EO63 for eels. These toxins are excreted in vitro by EO63 in the ratio of 4:1 during the stationary phase of growth. Strain EO63 also produced beta-haemolysins in vitro which could correspond to aerolysin. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on the purification of a metallo-elastase excreted by a wild-type A. hydrophila strain.