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Immunological assessment of mice hyperimmunized with native and Cobalt-60-irradiated Bothrops venoms
Ferreira Junior, R. S.;Nascimento, N.;Martinez, J. C.;Alves, J. B.;Meira, D. A.;Barraviera, B.;
Journal of Venomous Animals and Toxins including Tropical Diseases , 2005, DOI: 10.1590/S1678-91992005000400006
Abstract: elisa was used to evaluate, accompany, and compare the humoral immune response of swiss mice during hyperimmunization with native and cobalt-60-irradiated (60co) venoms of bothrops jararaca, bothrops jararacussu and bothrops moojeni. potency and neutralization were evaluated by in vitro challenges. after hyperimmunization, immunity was observed by in vivo challenge, and the side effects were assessed. the animals immunization with one ld50 of each venom occurred on days 1, 15, 21, 30, and 45, when blood samples were collected; challenges happened on the 60th day. results showed that elisa was efficient in evaluating, accompanying and comparing mouse immune response during hyperimmunization. serum titers produced with natural venom were similar to those produced with irradiated venom. immunogenic capacity was maintained after 60co-irradiation. the sera produced with native venom showed neutralizing potency and capacity similar to those of the sera produced with irradiated venom. all antibodies were able to neutralize five ld50 from these venoms. clinical alterations were minimum during hyperimmunization with irradiated venom, however, necrosis and death occurred in animals inoculated with native venom.
Pulsed ultrasound therapy accelerates the recovery of skeletal muscle damage induced by Bothrops jararacussu venom
Saturnino-Oliveira, J.;Tomaz, M.A.;Fonseca, T.F.;Gaban, G.A.;Monteiro-Machado, M.;Strauch, M.A.;Cons, B.L.;Calil-Elias, S.;Martinez, A.M.B.;Melo, P.A.;
Brazilian Journal of Medical and Biological Research , 2012, DOI: 10.1590/S0100-879X2012007500033
Abstract: we studied the effect of pulsed ultrasound therapy (ust) and antibothropic polyvalent antivenom (pav) on the regeneration of mouse extensor digitorum longus muscle following damage by bothrops jararacussu venom. animals (swiss male and female mice weighing 25.0 ± 5.0 g; 5 animals per group) received a perimuscular injection of venom (1 mg/kg) and treatment with ust was started 1 h later (1 min/day, 3 mhz, 0.3 w/cm2, pulsed mode). three and 28 days after injection, muscles were dissected and processed for light microscopy. the venom caused complete degeneration of muscle fibers. ust alone and combined with pav (1.0 ml/kg) partially protected these fibers, whereas muscles receiving no treatment showed disorganized fascicules and fibers with reduced diameter. treatment with ust and pav decreased the effects of the venom on creatine kinase content and motor activity (approximately 75 and 48%, respectively). sonication of the venom solution immediately before application decreased the in vivo and ex vivo myotoxic activities (approximately 60 and 50%, respectively). the present data show that ust counteracts some effects of b. jararacussu venom, causing structural and functional improvement of the regenerated muscle after venom injury.
Rabbit antivenom efficacy against myotoxic and neurotoxic activities of Bothrops jararacussu venom and bothropstoxin-I
Oshima-Franco, Y.;Leite, G. B.;Valério, A. A.;Hyslop, S.;Andriao-Escarso, S.;Giglio, J. R.;Prado-Franceschi, J.;Cruz-H?fling, M. A.;Rodrigues-Simioni, L.;
Journal of Venomous Animals and Toxins , 2002, DOI: 10.1590/S0104-79302002000200004
Abstract: bothrops jararacussu venom and its major toxin bothropstoxin-i (bthtx-i) possess myotoxic and neurotoxic properties. the efficacy of a rabbit antivenom raised against b. jararacussu venom in the neutralization of physiological, biochemical, and morphological changes induced by the venom and its major toxin bthtx-i was studied in mouse isolated phrenic nerve-diaphragm (pnd) and extensor digitorum longus (edl) preparations. the times required for 50% neuromuscular blockade in pnd and edl preparations for venom were 70+11.5 (s.e.m., n=5) min and 58+8 (n=16) (50 m g/ml), and for bthtx-i 31+6 (n=3) min and 30+3 (n=5) min (20 m g/ml), respectively. after 120 min incubation, creatine kinase (ck) concentrations in solution containing the edl preparations were 3464+346 u/l after exposure to venom (50 m g/ml, n=5) and 3422+135 u/l to bthtx-i (20m g/ml, n=4), respectively. rabbit antivenom dose-dependently neutralized venom and toxin-induced neuromuscular blockade in both preparations and effectively prevented venom and toxin-induced ck release from edl. histological analysis showed that rabbit antivenom neutralized morphological damage caused by b. jararacussu venom and bthtx-i in edl preparations. these results indicate that rabbit antivenom effectively neutralized the biological activities of b. jararacussu venom and bthtx-i.
Acción del veneno de Bothrops jararacussu de Argentina sobre la coagulación sanguínea Activity of the venom of Bothrops jararacussu of Argentina on blood coagulation  [cached]
S.L. Maru?ak,R.M. Ruíz de Torrent,G.P. Teibler,C.C. Gay
InVet , 2006,
Abstract: La intoxicación por mordeduras de serpientes del género Bothrops (yarará) de Argentina se caracteriza por causar efectos locales y sistémicos. Entre los sistémicos pueden generar alteraciones en la coagulación de la sangre, hemorragias, shock, insuficiencia renal. En este trabajo se estudiaron los efectos que causa el veneno de Bothrops jararacussu de Misiones, Argentina, sobre la coagulación de la sangre. Se utilizaron métodos in vitro e in vivo para evaluar la actividad coagulante, fibrinolítica y defibrinante, como así también la capacidad de degradar el fibrinógeno y la letalidad del veneno. El veneno mostró ser α-fibrinogenolítico y los resultados para las actividades ensayadas fueron: concentración coagulante mínima (CCM) 18,5 μg/ml, concentración fibrinolítica mínima (CFM) 9,5 mg/ml, dosis defibrinante mínima (DDM) 1,56 μg, DL50 43,52 μg/ratón, los que difieren de los obtenidos para otras especies de Bothrops e incluso dentro de la misma especie, pero distribuídas en otras regiones sudamericanas. Se concluye que el veneno de B. jararacussu de Argentina afecta marcadamente el sistema hemostático, y que las actividades coagulantes y anticoagulantes exhibidas por esta secreción probablemente influyan en la letalidad del veneno, dado que contribuirían a la aparición de hemorragias que, de ser severas, conducen a fallo renal y muerte de la víctima. The envenomation by snakes of the genus Bothrops of Argentina causes local and systemic signs. Among the systemic damages, bothropic venoms induce a wide variety of effects on blood coagulation, haemorrhage, shock and renal failure. In this paper the effects of Bothrops jararacussu venom from Argentina on the hemostatic system were determined. In order to estimate blood-clotting, fibrinolytic and defibrinating activities of the venom, in vitro and in vivo tests were employed. The ability to degrade fibrinogen and the lethal dosis (DL50) also were determined. The B. jararacussu venom appears to be fibrinogenolytic and the values obtained for assayed activities were minimum blood-clotting concentration 18,5 μg/ml, minimum fibrinolytic dose 9,5 mg/ml, minimum defibrinating dose 1,56 μg, LD50 43,52 μg/mouse. These results differ from those obtained for other species of genus Bothrops, or from the same species that inhabits in different South American regions. It is concluded that Bothrops jararacussu venom from Argentina largely affects blood coagulation system, and that the procoagulant and anticoagulant activities exhibited by this secretion may contribute to the lethality of the venom, since they could promo
Heparin and commercial bothropic antivenom against the paralyzing effect of Bothrops jararacussu snake venom
Rostelato-Ferreira, S;Rodrigues-Simioni, L;Oshima-Franco, Y;
Journal of Venomous Animals and Toxins including Tropical Diseases , 2010, DOI: 10.1590/S1678-91992010005000008
Abstract: the crude venom of bothrops jararacussu (bjssu) is known to induce muscular paralysis in vitro. many studies have shown that various substances, including heparin, neutralize the damage caused by snake venom. in the present study, the ability of heparin (hep) and commercial bothropic antivenom (cba) to neutralize neuromuscular effects of bjssu venom, at different time-points, was analyzed. mouse phrenic nerve-diaphragm preparation was used through a conventional myographic technique, following five different protocols: group 1 was incubated with bjssu (40 μg/ml) without any other treatment; groups 2 and 3 were pretreated with heparin (1 μl/ml) and cba (120 μl/ml), respectively, for 15 minutes before venom addition; group 4 after 50% neuromuscular blockade induced by bjssu crude venom received 1 μl/ml of heparin while group 5 received a mixture of hep:cba:bjssu. control preparations (tyrode) were treated with hep and cba (mean ± sem; n = 3-6). after 120 minutes of venom incubation, group 1 preparations presented twitch-tension of 12 ± 2%. however, in groups 2 and 3, the neutralizations were 92 ± 1.9% and 81 ± 6%, respectively. the heparin addition, after 50% neuromuscular blockade by bjssu, produced 40 ± 6% muscular response after 120 minutes of incubation. hep:cba:bjssu mixture displayed a protective effect of 84 ± 10% against venom action. in conclusion, heparin and commercial bothropic antivenom efficiently neutralized the neurotoxic effects caused by b. jararacussu crude venom, even at different incubation time-points.
Histological and biochemical effects induced by sublethal doses of Bothrops jararacussu venom in mice
Zeni, A. L. B.;Becker, A.;Krug, M.;Albuquerque, C. A. C.;
Journal of Venomous Animals and Toxins including Tropical Diseases , 2007, DOI: 10.1590/S1678-91992007000300009
Abstract: snake venom is characterized by hemorrhagic, coagulant, proteolytic and myotoxic activities which in bothrops jaracussu venom are related to intraspecific variations. in the present study, female swiss mice were divided into two groups: treated with 25μg or 50μg venom. these were subdivided into three groups of six animals each, according to blood collection: 2, 4 or 24h after venom injection. animals were anesthetized using diethyl-ether inhalation and 1ml of blood was collected by heart puncture. then, the following organs were removed: spleen, skeletal muscle, kidneys, liver and lungs; histological sections were obtained and stained with hematoxylin-eosin (he). the following biochemical parameters were analyzed: aspartate aminotransferase (ast/got), alanine aminotransferase (alt/gpt), total lactate dehydrogenase (ldh), glucose, creatinine and urea levels, and total protein content. results showed significant alterations in ast, ldh, glucose and urea levels, and total protein content, as well as important tissue alterations in the liver, kidneys and lungs. it could be concluded that, even using sublethal doses of venom, there were significant changes in almost all the tested biochemical parameters as well as tissue alterations in the kidneys and lungs.
Low-level laser therapy decreases local effects induced by myotoxins isolated from Bothrops jararacussu snake venom
Barbosa, AM;Villaverde, AB;Guimar?es-Sousa, L;Soares, AM;Zamuner, SF;Cogo, JC;Zamuner, SR;
Journal of Venomous Animals and Toxins including Tropical Diseases , 2010, DOI: 10.1590/S1678-91992010000300014
Abstract: the prominent myotoxic effects induced by bothrops jararacussu crude venom are due, in part, to its polycationic myotoxins, bthtx-i and bthtx-ii. both myotoxins have a phospholipase a2 structure: bthtx-ii is an active enzyme asp-49 pla2, while bthtx-i is a lys-49 pla2 devoid of enzymatic activity. in this study, the effect of low-level laser therapy (lllt), 685 nm laser at a dose of 4.2 j/cm2 on edema formation, leukocyte influx and myonecrosis caused by bthtx-i and bthtx-ii, isolated from bothrops jararacussu snake venom, was analyzed. bthtx-i and bthtx-ii caused a significant edema formation, a prominent leukocyte infiltrate composed predominantly by neutrophils and myonecrosis in envenomed gastrocnemius muscle. lllt significantly reduced the edema formation, neutrophil accumulation and myonecrosis induced by both myotoxins 24 hours after the injection. lllt reduced the myonecrosis caused by bthtx-i and bthtx-ii, respectively, by 60 and 43%; the edema formation, by 41 and 60.7%; and the leukocyte influx, by 57.5 and 51.6%. in conclusion, lllt significantly reduced the effect of these snake toxins on the inflammatory response and myonecrosis. these results suggest that lllt should be considered a potential therapeutic approach for treatment of local effects of bothrops species venom.
Bothrops jararacussu venom-induced neuromuscular blockade inhibited by Casearia gossypiosperma Briquet hydroalcoholic extract
Camargo, TM;Nazato, VS;Silva, MG;Cogo, JC;Groppo, FC;Oshima-Franco, Y;
Journal of Venomous Animals and Toxins including Tropical Diseases , 2010, DOI: 10.1590/S1678-91992010000300009
Abstract: the hydroalcoholic extract of casearia gossypiosperma briquet (flacourtiaceae) was standardized for the first time through quality control procedures including pharmacognostic methods, fingerprint chromatograms, defined amounts of marker substances and physicochemical characteristics. the pharmacological activity of c. gossypiosperma (cg) hydroalcoholic extract was assayed by a traditional in vitro test, which involved irreversible neuromuscular blockade induced by bothrops jararacussu (bjssu) venom (60 μg/ml) in mouse phrenic nerve-diaphragm preparations. bjssu venom blocked muscle activity for 26 (± 2.0) minutes (n = 6). cg extract (0.1 mg/ml) induced changes on the baseline muscle activity without impairing the muscle function and inhibited 87.6% (± 1.8) (n = 6) of the bjssu venom-induced blockade. both flavonoids (0.624 g%) and polyphenols (4.63 g%) from the extract were spectrophotometrically quantified. therefore, the present study confirms the antibothropic activity of cg extract, supporting the ethnomedical use of casearia sp. in the treatment of snakebite victims.
Effect of perimuscular injection of Bothrops jararacussu venom on plasma creatine kinase levels in mice: influence of dose and volume
Calil-Elias, S.;Thattassery, E.;Martinez, A.M.B.;Melo, P.A.;
Brazilian Journal of Medical and Biological Research , 2002, DOI: 10.1590/S0100-879X2002001000018
Abstract: the effect of dose and volume of a perimuscular injection of bothrops jararacussu venom on myonecrosis of skeletal muscle was studied in mice. an increase of the venom dose (0.25 to 2.0 μg/g) at a given volume (50 μl) resulted in an increase in plasma creatine kinase (ck) levels 2 h after injection. plasma ck activity increased from the basal level of 129.27 ± 11.83 (n = 20) to 2392.80 ± 709.43 iu/l (n = 4) for the 1.0 μg/g dose. histological analysis of extensor digitorum longus muscle 4 h after injection showed lesion of peripheral muscle fibers, disorganization of the bundles or the complete degeneration of muscle fibers. these lesions were more extensive when higher doses were injected. furthermore, an increase in volume (12.5 to 100 μl) by dilution of a given dose (0.5 μg/g) also increased plasma ck levels from 482.31 ± 122.79 to 919.07 ± 133.33 iu/l (n = 4), respectively. these results indicate that care should be taken to standardize volumes and sites of venom injections.
Effect of perimuscular injection of Bothrops jararacussu venom on plasma creatine kinase levels in mice: influence of dose and volume  [cached]
Calil-Elias S.,Thattassery E.,Martinez A.M.B.,Melo P.A.
Brazilian Journal of Medical and Biological Research , 2002,
Abstract: The effect of dose and volume of a perimuscular injection of Bothrops jararacussu venom on myonecrosis of skeletal muscle was studied in mice. An increase of the venom dose (0.25 to 2.0 μg/g) at a given volume (50 μl) resulted in an increase in plasma creatine kinase (CK) levels 2 h after injection. Plasma CK activity increased from the basal level of 129.27 ± 11.83 (N = 20) to 2392.80 ± 709.43 IU/l (N = 4) for the 1.0 μg/g dose. Histological analysis of extensor digitorum longus muscle 4 h after injection showed lesion of peripheral muscle fibers, disorganization of the bundles or the complete degeneration of muscle fibers. These lesions were more extensive when higher doses were injected. Furthermore, an increase in volume (12.5 to 100 μl) by dilution of a given dose (0.5 μg/g) also increased plasma CK levels from 482.31 ± 122.79 to 919.07 ± 133.33 IU/l (N = 4), respectively. These results indicate that care should be taken to standardize volumes and sites of venom injections.
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