Involvement of mast cells and histamine in edema Scolopendra viridicornis centipede venom

Bites caused by Scolopendra viridicornis centipede are mainly characterized by burning pain, paresthesia and edema. On this regard, the aim of this work was to study the involvement of mast cells and histamine in edema induced by Scolopendra viridicornis (Sv) centipede venom. The edema was analyzed on mice paws. The mice were pretreated with cromolyn (mast cell degranulation inhibitor) and antagonists of histamine receptors, such as promethazine (H1R), cimetidine (H2R) and thioperamide (H-3/H4R). The analyses were carried out at different times after the injection of Sv venom (15 mu g) or PBS in the footpad of mice. Our results showed a significant inhibition of the edema induced by Sv venom injection in mice previously treated: cromolyn (38-91%), promethazine (50-59%) and thioperamide (around 30%). The treatment with cimetidine did not alter the edema induced by Sv venom. Histopathological analysis showed that Sv venom injection (15 mu g) induced edema, leukocyte recruitment and mast cells degranulation, when compared with the PBS-injected mice. Direct effects of the Sv venom on mast cells were studied in PT-18 line (mouse mast cell) and RBL-2H3 cells (rat mast cells). The data showed that higher doses (3.8 and 7.5 mu g) of Sv venom were cytotoxic for both cell lineages and induced morphological changes. However, lower doses of the venom induced degranulation of both mast cell lines, as well as the secretion of MCP-1, IL-6 and IL-1 beta. The production of PGD(2) was only observed in the RBL-2H3 line incubated with Sv venom. Taking our results together, we demonstrated that upon Sv venom exposure, mast cells and histamine are crucial for the establishment of the local inflammatory reaction. (C) 2016 Elsevier Ltd. All rights reserved.

The cutaneous secretion of the casque-headed tree frog Corythomantis greeningi: Biochemical characterization and some biological effects

Corythomantis greeningi is a tree-frog endemic of the Brazilian semi-arid (Caatinga), mainly characterized by the flat, mineralized and spiny head, which is associated with phragmotic habits. It is already known that the skin secretion of this amphibian from both head and body is quite toxic and is used as an efficient chemical defence against predators. However, the biochemical characteristics and pharmacological effects of this secretion are still very little studied. We have tested the crude skin secretion, as well as the ten major fractions obtained by RP-HPLC for nociceptive and edema activity and for in vitro cytotoxicity using murine models. SDS-PAGE analyses demonstrated that the majority of proteins ranging through the gel lie between 55 and 30 kDa. LC-MS analysis showed multiple low molecular mass molecules (200-500 Da), which are consistent with masses of alkaloids and steroids. The crude skin secretion was able to induce fast and persistent edema accompanied by intense dose-dependent nociception. From the 10 tested fractions, five induced both edema and nociception, six fractions were able to induce only edema (80-170% control), and seven fractions induced only nociception (15-30 times compared to control). In addition, inhibition of cell growth (IC50) was demonstrated in murine fibroblasts and melanoma cells. From the data obtained, we confirmed that the skin secretion of C. greeningi is very toxic and is rich in compounds able to directly provoke local inflammation and nociception. Such characteristics are important as part of the chemical defensive repertory of this species. (C) 2016 Elsevier Ltd. All rights reserved.

Biochemical and biological characterization of Bothriechis schlegelii snake venoms from Colombia and Costa Rica

Snakebites inflicted by the arboreal viperid snake Bothriechis schlegelii in humans are characterized by pain, edema, and ecchymosis at the site of the bite, rarely with blisters, local necrosis, or defibrination. Herein, a comparative study of Bothriechis schlegelii snake venoms from Colombia (BsCo) and Costa Rica (BsCR) was carried out in order to compare their main activities and to verify the efficacy of Bothrops antivenom produced in Brazil to neutralize them. Biochemical (SDS-PAGE and zymography) and biological parameters (edematogenic, lethal, hemorrhagic, nociceptive, and phospholipase A(2) activities) induced by BsCo and BsCR snake venoms were evaluated. The presence of antibodies in Bothrops antivenom that recognize BsCo and BsCR snake venoms by enzyme-linked immunosorbent assay and Western blotting, as well as the ability of this antivenom to neutralize the toxic activities were also verified. SDS-PAGE showed differences between venoms. Distinctive caseinolytic and hyaluronidase patterns were detected by zymography. BsCo and BsCR showed similar phospholipase A(2) activity. Strong cross-reactivity between BsCo and BsCR was detected using Bothrops antivenom with many components located between 150 and 35kDa. BsCR was more edematogenic and almost fourfold more hemorrhagic than BsCo, and both venoms induced nociception. BsCR (LD50 5.60mg/kg) was more lethal to mice than BsCo (LD50 9.24mg/kg). Bothrops antivenom was effective in the neutralization of lethal and hemorrhagic activities of BsCo and BsCR and was partially effective in the neutralization of edematogenic and nociceptive activities. In conclusion, geographic distribution influences the composition and activities of Bothriechis schlegelii venoms. Bothrops antivenom cross-reacted with these venoms and was partially effective in neutralizing some toxic activities of BsCo and BsCR

Platelet participation in the pathogenesis of dermonecrosis induced by Loxosceles gaucho venom

Loxosceles gaucho spider venom induces in vitro platelet activation and marked thrombocytopenia in rabbits. Herein, we investigated the involvement of platelets in the development of the dermonecrosis induced by L. gaucho venom, using thrombocytopenic rabbits as a model. L. gaucho venom evoked a drop in platelet and neutrophil counts 4 h after venom injection. Ecchymotic areas at the site of venom inoculation were noticed as soon as 4 h in thrombocytopenic animals but not in animals with initial normal platelet counts. After 5 days, areas of scars in thrombocytopenic animals were also larger, evidencing the marked development of lesions in the condition of thrombocytopenia. Histologically, local hemorrhage, collagen fiber disorganization, and edema were more severe in thrombocytopenic animals. Leukocyte infiltration, predominantly due to polymorphonuclears, was observed in the presence or not of thrombocytopenia. Thrombus formation was demonstrated by immunohistochemistry at the microvasculature, and it occurred even under marked thrombocytopenia. Taken together, platelets have an important role in minimizing not only the hemorrhagic phenomena but also the inflammatory and wound-healing processes, suggesting that cutaneous loxoscelism may be aggravated under thrombocytopenic conditions

Ontogenetic Variation in Biological Activities of Venoms from Hybrids between Bothrops erythromelas and Bothrops neuwiedi Snakes

Lance-headed snakes are found in Central and South America, and they account for most snakebites in Brazil. The phylogeny of South American pitvipers has been reviewed, and the presence of natural and non-natural hybrids between different species of Bothrops snakes demonstrates that reproductive isolation of several species is still incomplete. The present study aimed to analyze the biological features, particularly the thrombin-like activity, of venoms from hybrids born in captivity, from the mating of a female Bothrops erythromelas and a male Bothrops neuwiedi, two species whose venoms are known to display ontogenetic variation. Proteolytic activity on azocoll and amidolytic activity on N-benzoyl-DL-arginine-p-nitroanilide hydrochloride (BAPNA) were lowest when hybrids were 3 months old, and increased over body growth, reaching values similar to those of the father when hybrids were 12 months old. The clotting activity on plasma diminished as hybrids grew; venoms from 3- and 6-months old hybrids showed low clotting activity on fibrinogen (i.e., thrombinlike activity), like the mother venom, and such activity was detected only when hybrids were older than 1 year of age. Altogether, these results point out that venom features in hybrid snakes are genetically controlled during the ontogenetic development. Despite the presence of the thrombin-like enzyme gene(s) in hybrid snakes, they are silenced during the first six months of life.