Antiophidic activity of the extract of the Amazon plant Humirianthera ampla and constituents.

ETHNOPHARMACOLOGICAL RELEVANCE: Although serotherapy against snakebite has been discovered more than one hundred years ago, antivenom is not available all over Brazil. The use of plants from folk medicine is common mainly in the Brazilian Amazon area. One of these plants is named Humirianthera ampla (HA). MATERIALS AND METHODS: We have investigated HA extract and constituents' antiophidic activity in different experimental protocols against some Bothrops snake venoms (Bothrops jararacussu, Bothrops atrox and Bothrops jararaca). The protocols investigated include phospholipase, proteolytic, pro-coagulant, hemorrhagic, edematogenic and myotoxic activities induced by these venoms in Swiss mice. RESULTS: All the venoms caused an increase in the rate of creatine kinase (CK) release from isolated muscles, indicating damage to the sarcolemma. The crude extract of HA decreased the myotoxic activity in a concentration-dependent fashion. The presence of HA 300 µg/mL decreased up to 96% of Bothrops jararacussu and 94% of Bothrops atrox myotoxicity after 90 min of exposure. In vivo myotoxicity of Bothrops atrox venom was decreased in 75% when the venom was preincubated with HA 500 mg/kg. Similar results were observed with lupeol against Bothrops jararacussu and Bothrops atrox venoms. The hemorrhagic activity was evaluated by intradermal injection of Bothrops atrox venom. Preincubation and oral pre- and posttreatment with HA decreased hemorrhage by 100%, 45% and 45%, respectively. Bothrops atrox venom also induced formation of edema, which was significantly inhibited by pre- and posttreatment with HA. All the venoms showed extensive pro-coagulating properties, and these activities were inhibited by up to 90% with HA, which presented concentration-dependent inhibition. Finally, proteolytic and phospholipase activities of the venoms were all inhibited by increasing concentrations of HA, lupeol and sitosterol. The inhibition of these activities might help explain the actions against in vivo myotoxicity and the in vivo effects observed, i.e., edema, myotoxicity, pro-coagulation and hemorrhage. CONCLUSIONS: Altogether, our results give support for the popular use of HA extracts in cases of accidents with snakes, suggesting that it can be used as an adjunct in the management of venomous snakebites.

Preclinical assessment of the neutralizing capacity of antivenoms produced in six Latin American countries against medically-relevant Bothrops snake venoms.

Species of the genus Bothrops induce the vast majority of snakebite envenomings in Latin America. A preclinical study was performed in the context of a regional network of public laboratories involved in the production, quality control and development of antivenoms in Latin America. The ability of seven polyspecific antivenoms, produced in Argentina, Brazil, Peru, Bolivia, Colombia and Costa Rica, to neutralize lethal, hemorrhagic, coagulant, defibrinogenating and myotoxic activities of the venoms of Bothrops neuwiedi (diporus) (Argentina), Bothrops jararaca (Brazil), B. neuwiedi (mattogrossensis) (Bolivia), Bothrops atrox (Peru and Colombia) and Bothrops asper (Costa Rica) was assessed using standard laboratory tests. Despite differences in the venom mixtures used in the immunization of animals for the production of these antivenoms, a pattern of extensive cross-neutralization was observed between these antivenoms and all the venoms tested, with quantitative differences in the values of effective doses. This study reveals the capacity of these antivenoms to neutralize, in preclinical tests, homologous and heterologous Bothrops venoms in Central and South America, and also highlight quantitative differences in the values of Median Effective Doses (ED50s) between the various antivenoms.

Effect of low-level laser therapy in the myonecrosis induced by Bothrops jararacussu snake venom.

OBJECTIVE: The aim of this work was to investigate the capacity of low-level laser therapy (LLLT) alone or in combination with antivenom (AV) to reduce myonecrosis induced by Bothrops jararacussu snake venom. BACKGROUND DATA: Myonecrosis is the most pronounced local effect caused by B. jararacussu venom. AV therapy and other first-aid treatments do not reverse these local effects. MATERIAL AND METHODS: Male Swiss mice were used. Myonecrosis was induced by injection of 0.6 mg/kg of B. jararacussu venom in the right gastrocnemius muscle and was evaluated at 3 or 24 h after venom injection. The site of venom administration was irradiated for 29 s with a low power semiconductor laser (685 nm) at a dose of 4.2 J/cm(2). Intravenous AV therapy (0.5 mL dose) was administered at different times: 30 min before venom injection or 0, 1, or 3 h afterward. Both AV therapy and LLLT treatments were duplicated in mice groups killed at 3 or 24 h. RESULTS: B. jararacussu venom caused a significant myonecrotic effect 3 and 24 h after venom injection. LLLT significantly reduced myonecrosis by 83.5% at 24 h (p < 0.05) but not at 3 h, and AV therapy alone was ineffective for reducing myonecrosis at 3 and 24 h. CONCLUSION: Only LLLT significantly reduced myonecrosis of the envenomed muscle, suggesting that LLLT is a potentially therapeutic approach for treating the local effects of B. jararacussu venom.

Effect of low-level laser therapy in the inflammatory response induced by Bothrops jararacussu snake venom.

This article reports the effect of low-level laser therapy (LLLT) on the edema formation and leukocyte influx caused by Bothrops jararacussu snake venom as an alternative treatment for Bothrops snakebites. The inflammatory reaction was induced by injection of 0.6 mg/kg of B. jararacussu venom, in gastrocnemius muscle. Cell influx and edema were evaluated at 3 or 24h after venom injection. Mice were irradiated at the site of injury by a low-level laser (685 nm) with a dose of 4.2J/cm(2). A therapy that combines LLLT and antivenom was also studied. B. jararacussu venom caused a significant edema formation 3 and 24h after its injection, and a prominent leukocyte infiltrate composed predominantly of neutrophils at 24h after venom inoculation. LLLT significantly reduced edema formation by 53% and 64% at 3 and 24h, respectively, and resulted in a reduction of neutrophils accumulation (P<0.05). The combined therapy showed to be more efficient than each therapy acting separately. In conclusion, LLLT significantly reduced the edema and leukocyte influx into the envenomed muscle, suggesting that LLLT should be considered as a potentially therapeutic approach for the treatment of the local effects of Bothrops species.

Anticoagulant and antifibrinogenolytic properties of the aqueous extract from Bauhinia forficata against snake venoms.

The aqueous extract from aerial parts of Bauhinia forficata was able to neutralize the clotting activity induced by Bothrops and Crotalus crude venoms. The clotting time, upon human plasma, induced by B. moojeni venom was significantly prolonged. Clotting and fibrinogenolytic activities induced by isolated thrombin-like enzyme from Bothrops jararacussu were totally inhibited after incubation at different ratios. The extract was not able to neutralize the hemorrhagic activity induced by an Bothrops venoms, but it efficiently inhibited the edema induced by Crotalus durissus terrificus venom and isolated PLA2s. In addition, it did not inhibited the phospholipase A2 activity of Bothrops snake venoms. Interaction studies between Bauhinia forficata extract and snake venoms, when analyzed by SDS-PAGE, did not reveal any apparent degradation of the venom proteins. This extract is a promising source of natural inhibitors of serine-proteases involved in blood clotting disturbances induced by snake venoms.

Effects of the Ga-As laser irradiation on myonecrosis caused by Bothrops Moojeni snake venom.

BACKGROUND AND OBJECTIVES: Viper snake envenoming induces in the victims systemic coagulopathy, and severe local tissue damage such as edema, hemorrhage, intense pain, and myonecrosis. Serumtherapy and other first-aid managements are ineffective in neutralizing these local effects. The effects of the gallium-arsenide (Ga-As) laser irradiation on mice gastrocnemius injected intramuscularly (i.m.) with Bothrops moojeni snake venom were investigated. STUDY DESIGN/MATERIALS AND METHODS: Macroscopical, histopatological, and myonecrosis quantification through serum creatine kinase (CK) evaluation was done at 3, 12, and 24 hours (two, five, and eight irradiation sessions, 4 J/cm(2), 1 minute 32 seconds per period, respectively), were done after the venom or saline injection, and in venom-unirradiated mice. RESULTS: In unirradiated gastrocnemius, the venom induced massive hemorrhage, vascular congestion, time-progressing myonecrosis, edema, abundant inflammatory infiltrate, and high CK serum levels. Ga-As irradiation significantly decreased the amount of myonecrosis in all the periods tested (P < 0.05). CONCLUSIONS: The laser treatment significantly inhibited the ability of B. moojeni venom to rapidly disrupt the integrity of the plasma membrane.

Reduction of rattlesnake-venom-induced myonecrosis in mice by hyperbaric oxygen therapy.

Hyperbaric oxygen therapy (HBOT) at 1, 2, and 2.75 atmospheres absolute (ATA) was used to treat rattlesnake (Crotalus atrox) venom-induced tissue damage and edema in thigh muscles of mice. Tissue damage was evaluated by double-blind histopathologic examination: tissue edema was determined by measuring tissue water content. A total of 10 intermittent exposures to oxygen over a period of 4 days at 2 and 2.75 ATA did not influence the resolution of venom-induced tissue edema, whereas tissue damage was significantly ameliorated as compared to air-treated envenomated controls. HBOT also promoted healing in the venom-injected mice as evidenced by the presence of regenerating muscle cells. It is concluded that HBOT may limit rattlesnake venom-induced myonecrosis and promote healing in a dose-response relationship without reducing venom-induced edema.