In Vivo Effects of Bothrops jararaca Venom on Metabolic Profile and on Muscle Protein Metabolism in Rats

This study investigated the in vivo effects of the Bothrops Jararaca venom (BjV) on general metabolic profile and, specifically. oil muscle protein metabolism in rats. The crude venom (0.4 mg/kg body weight, IV) was infused in awake rats, and plasma activity of enzymes and metabolites levels were determined after 1, 2, 3, and 4 hours. BjV increased urea, lactate, and activities of creatine kinase. lactate dehydrogenase. and aspartate aminotransferase after 4 hours. The content of liver glycogen was reduced by BjV. Protein metabolism was evaluated by means of microdialysis technique and in isolated muscles. BjV induced increase in the muscle interstitial-arterial tyrosine concentration difference. indicating a high protein catabolism. The myotoxicity induced by this venom is associated with reduction of protein synthesis and increase in rates of overall proteolysis, which was accompanied by activation of lysosomal and ubiquitin-proteasome systems without changes in protein levels of cathepsins and ubiquitin-protein conjugates.

An easy method for handling the genus Phoneutria (Araneae, Ctenidae) for venom extraction

This paper describes an easy, cheap, and safe method of capturing and handling the medically important spider Phoneutria for venom extraction. The method does not injure or kill the spider and allows the extraction of pure venom.

Brown spider venom toxins interact with cell surface and are endocytosed by rabbit endothelial cells

Bites from the Loxosceles genus (brown spiders) cause severe clinical symptoms, Including dermonecrotic injury, hemorrhage, hemolysis, platelet aggregation and renal failure. Histological findings of dermonecrotic lesions in animals exposed to Loxosceles intermedia venom show numerous vascular alterations Study of the hemorrhagic consequences of the venom in endothelial cells has demonstrated that the degeneration of blood vessels results not only from degradation of the extracellular matrix molecule or massive leukocyte infiltration, but also from a direct and primary activity of the venom on endothelial cells. Exposure of an endothelial cell line in vitro to L. intermedia venom induce morphological alterations, such as cell retraction and disadhesion to the extracellular matrix. The aim of the present study was to investigate the interaction between the venom toxins and the endothelial cell surface and their possible internalization, in order to illuminate the information about the deleterious effect triggered by venom After treating endothelial cells with venom toxins, we observed that the venom Interacts with cell surface. Venom treatment also can cause a reduction of cell surface glycoconjugates When cells were permeabilized, it was possible to verify that some venom toxins were internalized by the endothelial cells The venom internalization involves endocytic vesicles and the venom was detected in the lysosomes. However, no damage to lysosomal integrity was observed, suggesting that the cytotoxic effect evoked by L interned:a venom on endothelial cells is not mediated by venom internalization (C) 2010 Elsevier Ltd. All rights reserved

Respiratory Infection, Exposure to Mouse Allergen and Breastfeeding: Role in Recurrent Wheezing in Early Life

Background: Environmental factors may influence the development of allergen sensitization and asthma. The aim of this study was to evaluate the role of endotoxin and allergen exposure in early life as a risk factor for recurrent wheezing. Methods: One hundred and four infants from low-income families, at high risk of asthma, were enrolled at birth. Dust samples were collected from the bedding and bedroom floor within 6 months after birth. Recurrent wheezing was defined as 3 or more wheezing episodes in the past year. Endotoxin was determined by Limulus amebocyte lysate assay, and major indoor allergens were quantitated by ELISA in dust extracts. IgE antibodies were measured by ImmunoCAP at 30 months of age. Results: At 30 months, 51 of the 99 infants who completed the study (51.5%) had recurrent wheezing. Respiratory infection was strongly associated with recurrent wheezing (OR 6.67, 95% CI 1.96-22.72), whereas exclusive breastfeeding for at least 1 month was a protective factor (OR 0.09, 95% CI 0.01-0.51). Exposure to high levels of mouse allergen was more frequent among non-recurrent wheezers, approaching significance (OR 0.12, 95% CI 0.01-1.13; p=0.064). None of the children were sensitized to mouse. Sensitization to mite was found in 26/90 (28.8%) children, with no association with recurrent wheezing. Conclusion: Respiratory infection was strongly associated with recurrent wheezing in the first 30 months of life, in children at high risk of asthma, living in a socially deprived community in Brazil. Copyright (C) 2009 S. Karger AG, Basel

Leukotriene B(4) is essential for selective eosinophil recruitment following allergen challenge of CD4(+) cells in a model of chronic eosinophilic inflammation

Subcutaneous heat-coagulated egg white implants (EWI) induce chronic, intense local eosinophilia in mice, followed by asthma-like responses to airway ovalbumin challenge. Our goal was to define the mechanisms of selective eosinophil accumulation in the EWI model. EWI carriers were challenged i.p. with ovalbumin and the contributions of cellular immunity and inflammatory mediators to the resulting leukocyte accumulation were defined through cell transfer and pharmacological inhibition protocols. Eosinophil recruitment required Major Histocompatibility Complex Class It expression, and was abolished by the leukotriene B4 (LTB4) receptor antagonist CP 105.696, the 5-lipoxygenase inhibitor BWA4C and the 5-lipoxygenase activating protein inhibitor MK886. Eosinophil recruitment in EWI carriers followed transfer of: a) CD4(+) (but not CD4(-)) cells, harvested from EWI donors and restimulated ex vivo; b) their cell-free supernatants, containing LTB4. Restimulation in the presence of MK886 was ineffective. CC chemokine receptor ligand (CCL)5 and CCL2 were induced by ovalbumin challenge in vivo. mRNA for CCL17 and CCL11 was induced in ovalbumin-restimulated CD4(+) cells ex vivo. MK886 blocked induction of CCL17 Pretreatment of EWI carriers with MK886 eliminated the effectiveness of exogenously administered CCL11, CCL2 and CCL5. In conclusion, chemokine-producing, ovalburnin-restimulated CD4(+) cells initiate eosinophil recruitment which is strictly dependent on LTB4 production. (C) 2008 Elsevier Inc. All rights reserved.

Anticoagulant and fibrinogenolytic properties of the venom of Polybia occidentalis social wasp

Previous studies have shown that venoms of social wasps and bees exhibit strong anticoagulant activity. The present study describes the anticoagulant and fibrinogen-degrading pharmacological properties of the venom of Polybia occidentalis social wasp. The results demonstrated that this venom presented anticoagulant effect, inhibiting the coagulation at different steps of the clotting pathway (intrinsic, extrinsic and common pathway). The venom inhibited platelet aggregation and degraded plasma fibrinogen, possibly containing metal-dependent metalloproteases that specifically cleave the B beta-chain of fibrinogen. In conclusion, fibrinogenolytic and anticoagulant properties of this wasp venom find a potential application in drug development for the treatment of thrombotic disorders. For that, further studies should be carried out in order to identify and isolate the active compounds responsible for these effects. Blood Coagul Fibrinolysis 21: 653-659 (c) 2010 Wolters Kluwer Health | Lippincott Williams & Wilkins.

Neuromuscular activity of BaTX, a presynaptic basic PLA(2) isolated from Bothrops alternatus snake venom

We have previously isolated a Lys49 phospholipase A(2) homolog (BaTX) from Bothrops alternatus snake venom using a combination of molecular exclusion chromatography and reverse phase HPLC and shown its ability to cause neuromuscular blockade. In this work, we describe a one-step procedure for the purification of this toxin and provide further details of its neuromuscular activity. The toxin was purified by reverse phase HPLC and its purity and molecular mass were confirmed by SIDS-PAGE, MALDI-TOF mass spectrometry, amino acid analysis and N-terminal sequencing. BaTX (0.007-1.4 mu M) produced time-dependent, irreversible neuromuscular blockade in isolated mouse phrenic nerve-diaphragm and chick biventer cervicis preparations (time to 50% blockade with 0.35 mu M toxin: 58 +/- 4 and 24 +/- 1 min, respectively; n = 3-8; mean +/- S.E.) without significantly affecting the response to direct muscle stimulation. In chick preparations, contractures to exogenous acetylcholine (55 and 110 mu M) or KCl (13.4 mM) were unaltered after complete blockade by all toxin concentrations. These results, which strongly suggested a presynaptic mechanism of action for this toxin, were reinforced by (1) the inability of BaTX to interfere with the carbachol-induced depolarization of the resting membrane, (2) a significant decrease in the frequency and amplitude of miniature end-plate potentials, and (3) a significant reduction (59 +/- 4%, n=12) in the quantal content of the end-plate potentials after a 60 min incubation with the toxin (1.4 mu M). In addition, a decrease in the organ bath temperature from 37 degrees C to 24 degrees C and/or the replacement of calcium with strontium prevented the neuromuscular blockade, indicating a temperature-dependent effect possibly mediated by enzymatic activity. (C) 2009 Elsevier Inc. All rights reserved.

Effect of the Absence of Interleukin-12 on Mesangial Proliferative Glomerulonephritis Induced by Habu Snake Venom

Background. Interleukin-12 (IL12) participates in the pathophysiology of various experimental types of progressive glomerulonephritis, but its role in acute mesangial glomerulonephritis (AMG) induced by habu snake venom (HSV) has not been determined. This study aims to evaluate the effect of the absence of IL12 on AMG induced by HSV. Methods. AMG was induced in IL12 knockout (IL12-/-) and C57B1/6 (IL12+/+) mice by a single i.v. administration of HSV. Vehicle was used in control animals. Mice were studied after 3, 7, and 14 days (D3, D7, and D14). Results. After treatment with HSV, IL12+/+ and -/-mice developed focal glomerular lesions, but groups of both lineages showed no statistical difference concerning albuminuria, serum creatinine, histopathology, number of cells by glomerular tuft, and glomerular tuft area. Compared to IL12+/+ mice, IL12-/-mice showed lower scores of glomerular desmin expression on D7 [1.55 (1.32; 1.65) vs. 1.12 (1.07; 1.22); p < 0.01] and D14 [1.60 (1.55; 1.75) vs. 1.20 (1.15; 1.20); p < 0.001], respectively, and lower scores of glomerular alpha-SMA expression on D14 [0.30 (0.21; 0.38) vs. 0.16 (0.26; 0.36); p < 0.001], respectively. Conclusion. The absence of IL12 reduced the activity of mesangial cells, but did not modify the course of HSV-induced AMG in mice.

Plant-antivenom: Database of anti-venom medicinal plants

Plant-antivenom is a computational Websystem about medicinal plants with anti-venom properties. The system consists of a database of these plants, including scientific publications on this subject and amino acid sequences of active principles from venomous animals. The system relates these data allowing their integration through different search applications. For the development of the system, the first surveys were conducted in scientific literature, allowing the creation of a publication database in a library for reading and user interaction. Then, classes of categories were created, allowing the use of tags and the organization of content. This database on medicinal plants has information such as family, species, isolated compounds, activity, inhibited animal venoms, among others. Provision is made for submission of new information by registered users, by the use of wiki tools. Content submitted is released in accordance to permission rules defined by the system. The database on biological venom protein amino acid sequences was structured from the essential information from National Center for Biotechnology Information (NCBI). Plant-antivenom`s interface is simple, contributing to a fast and functional access to the system and the integration of different data registered on it. Plant-antivenom system is available on the Internet at http://gbi.fmrp.usp.br/plantantivenom.

Venom-Sweet-Venom: N-Linked Glycosylation in Snake Venom Toxins

Protein glycosylation represents one of the most important post-translational events, and is a mean of diversifying a protein without recourse to the genome. The venoms produced by snakes contain an abundance of glycoproteins with N-linked carbohydrates. N-linked glycosylation can ensure the correct folding of important functional domains. Characterization of carbohydrates structures aids in development of human therapeutics by snake venom toxins.

Inflammatory mediators involved in the nociceptive and oedematogenic responses induced by Tityus serrulatus scorpion venom injected into rat paws.

The present study investigated the role of kinins, prostaglandins (PGs) and nitric oxide (NO) in mechanical hypernociception, Spontaneous nociception and paw oedema after intraplantar (ipl) injection of Tityus serrulatus venom (Tsv) in male Wistar rats. Tsv was ipl-injected in doses of 0.01-10 mu g/paw. Pre-treatment (30 min prior) with DALBK (100 nmol/paw) and icatibant (10 nmol/paw), B1 and B2 selective kinin receptor antagonists, L-NAME (50 mg/kg, i.p., a non-selective nitric oxide synthase inhibitor) or celecoxib, selective COX-2 inhibitor, was given 1 h prior per os (5 mg/kg, p.o.), significantly reduced the hypernociceptive response (Von Frey method), the spontaneous nociception (determined by counting the number of flinches) and paw oedema (plethysmometer method) induced by Tsv at doses of 1.0 and 10 mu g/paw for both nociceptive and oedematogenic responses, respectively. Nevertheless, indomethacin (5 mg/kg, i.p.. 30 min prior) was ineffective in altering all of these events. The results of the present study show that Tsv, injected ipl into the rat paw, causes a dose-dependent paw oedema, mechanical hypernociception and flinches (a characteristic biphasic response) in which kinins and NO are substantially involved. Although celecoxib was effective against the oedema and pain caused by Tsv, COX-2 does not seem to be involved in the inflammatory response caused by Tsv. (C) 2008 Elsevier Ltd. All rights reserved.

Identification and characterization of a new member of snake venom thrombin inhibitors from Bothrops insularis using a proteomic approach

Snake venom C-type lectin-like proteins (CLPs) are ubiquitously found in Viperidae snake venoms and differ from the C-type lectins as they display different biological activities but no carbohydrate-binding activity. Previous analysis of the transcriptome obtained from the Bothrops insularis venom gland showed the presence of two clusters homologous to bothrojaracin (BJC) chains a and P. In an effort to identify a new BJC-like molecule, we used an approach associated with proteomic technologies to identify the presence of the expressed protein and then to purify and characterize a new thrombin inhibitor from B. insularis venom. We also constructed homology models of this protein and BJC, which were compared with other C-type lectin-like family members and revealed several conserved features of this intriguing snake venom toxin family. (C)0 2007 Elsevier Ltd. All rights reserved.

Analysis of the Ontogenetic Variation in the Venom Proteome/Peptidome of Bothrops jararaca Reveals Different Strategies to Deal with Prey

Previous studies have demonstrated that the pharmacological activities displayed by Bothrops jararaca venom undergo a significant ontogenetic shift. Variation in the venom proteome is a well-documented phenomenon; however, variation in the venom peptidome is poorly understood. We report a comparative proteomic and peptidomic analysis of venoms from newborn and adult specimens of B. jararaca and correlate it with the evaluation of important venom features. We demonstrate that newborn and adult venoms have similar hemorrhagic activities, while the adult venom has a slightly higher lethal activity in mice; however, the newborn venom is extremely more potent to kill chicks. The coagulant activity of newborn venom upon human plasma is 10 times higher than that of adult venom. These differences were clearly reflected in their different profiles of SDS-PAGE, gelatin zimography, immunostaining using specific antibodies, glycosylation pattern, and concanavalin A-binding proteins. Furthermore, we report for the first time the analysis of the peptide fraction of newborn and adult venoms by MALDI-TOF mass spectrometry and LC-MS/MS, which revealed different contents of peptides, while the bradykinin potentiating peptides (BPPs) showed rather similar profiles and were detected in the venoms showing their canonical sequences and also novel sequences corresponding to BPPs processed from their precursor protein at sites so far not described. As a result of these studies, we demonstrated that the ontogenetic shift in diet, from ectothermic prey in early life to endothermic prey in adulthood, and in animal size are associated with changes in the venom proteome in B. jararaca species.

Antitumoural effect of an L-amino acid oxidase isolated from Bothrops jararaca snake venom

An L-amino acid oxidase (BjarLAAO-I) from Bothrops jararaca snake venom was highly purified using a stepwise sequential chromatography on Sephadex G-75, Benzamidine Sepharose and Phenyl Sepharose. Purified BjarLAAO-I showed a molecular weight around 60,000 under reducing conditions and about 125,000 in the native form, when analysed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration, respectively. BjarLAAO-I is a homodimeric acidic glycoprotein, pI similar to 5.0, and N-terminal sequence showing close structural homology with other snake venom LAAOs. The purified enzyme catalysed the oxidative deamination of L-amino acids, the most specific substrate being L-Phe. Five amino acids, L-Ser, L-Pro, L-Gly, L-Thr and L-Cys were not oxidized, clearly indicating a significant specificity. BjarLAAO-I significantly inhibited Ehrlich ascites tumour growth and induced an influx of polymorphonuclear cells, as well as spontaneous liberation of H(2)O(2) from peritoneal macrophages. Later, BjarLAAO-I induced mononuclear influx and peritoneal macrophage spreading. Animals treated with BjarLAAO-I showed higher survival time.

Food aversion: A critical balance between allergen-specific IgE levels and taste preference

Animals sensitized to allergens change their feeding behavior and avoid drinking the otherwise preferred sweetened solutions containing the allergens. This phenomenon, known as food aversion, appears to be mediated by allergen-specific IgE antibodies. Here we investigated food aversion in BALB/c and C57BL/6 mice, which differ in their allergic responses to the allergen ovalbumin as well as in their preference for sweet taste. BALB/c mice present higher levels of IgE and a natural lower preference for sweet flavors when compared to C57BL/6 mice. Specifically, we studied a conflicting situation in which animals simultaneously experienced the aversive contact with the allergen and the attractive sweet taste of increasing concentrations of sucrose. We found that BALB/c mice were more prone to develop food aversion than C57BL/6 mice and that this aversive behavior could be abolished in both strains by increasing the palatability of the solution containing the allergen. In both strains food aversion was positively correlated with the levels of allergen-specific IgE antibodies and inversely correlated with their preference for sucrose sweetened solutions. (C) 2009 Elsevier Inc. All rights reserved.