Suppression of cardiomyocyte functions by ß-CTX isolated from the Thai king cobra (Ophiophagus hannah) venom via an alternative method

Beta-cardiotoxin (ß-CTX), the three-finger toxin isolated from king cobra (Ophiophagus hannah) venom, possesses ß-blocker activity as indicated by its negative chronotropy and its binding property to both ß-1 and ß-2 adrenergic receptors and has been proposed as a novel ß-blocker candidate. Previously, ß-CTX was isolated and purified by FPLC. Here, we present an alternative method to purify this toxin. In addition, we tested its cytotoxicity against different mammalian muscle cell types and determined the impact on cardiac function in isolated cardiac myocyte so as to provide insights into the pharmacological action of this protein. Methods: ß-CTX was isolated from the crude venom of the Thai king cobra using reverse-phased and cation exchange HPLC. In vitro cellular viability MTT assays were performed on mouse myoblast (C2C12), rat smooth muscle (A7r5), and rat cardiac myoblast (H9c2) cells. Cell shortening and calcium transient dynamics were recorded on isolated rat cardiac myocytes over a range of ß-CTX concentration. Results: Purified ß-CTX was recovered from crude venom (0.53% w/w). MTT assays revealed 50% cytotoxicity on A7r5 cells at 9.41 ± 1.14 µM (n = 3), but no cytotoxicity on C2C12 and H9c2 cells up to 114.09 µM. ß-CTX suppressed the extend of rat cardiac cell shortening in a dose-dependent manner; the half-maximal inhibition concentration was 95.97 ± 50.10 nM (n = 3). In addition, the rates of cell shortening and re-lengthening were decreased in ß-CTX treated myocytes concomitant with a prolongation of the intracellular calcium transient decay, indicating depression of cardiac contractility secondary to altered cardiac calcium homeostasis. Conclusion: We present an alternative purification method for ß-CTX from king cobra venom. We reveal cytotoxicity towards smooth muscle and depression of cardiac contractility by this protein. These data are useful to aid future development of pharmacological agents derived from ß-CTX.(AU)

Suppression of cardiomyocyte functions by ß-CTX isolated from the Thai king cobra (Ophiophagus hannah) venom via an alternative method

Beta-cardiotoxin (ß-CTX), the three-finger toxin isolated from king cobra (Ophiophagus hannah) venom, possesses ß-blocker activity as indicated by its negative chronotropy and its binding property to both ß-1 and ß-2 adrenergic receptors and has been proposed as a novel ß-blocker candidate. Previously, ß-CTX was isolated and purified by FPLC. Here, we present an alternative method to purify this toxin. In addition, we tested its cytotoxicity against different mammalian muscle cell types and determined the impact on cardiac function in isolated cardiac myocyte so as to provide insights into the pharmacological action of this protein. Methods: ß-CTX was isolated from the crude venom of the Thai king cobra using reverse-phased and cation exchange HPLC. In vitro cellular viability MTT assays were performed on mouse myoblast (C2C12), rat smooth muscle (A7r5), and rat cardiac myoblast (H9c2) cells. Cell shortening and calcium transient dynamics were recorded on isolated rat cardiac myocytes over a range of ß-CTX concentration. Results: Purified ß-CTX was recovered from crude venom (0.53% w/w). MTT assays revealed 50% cytotoxicity on A7r5 cells at 9.41 ± 1.14 µM (n = 3), but no cytotoxicity on C2C12 and H9c2 cells up to 114.09 µM. ß-CTX suppressed the extend of rat cardiac cell shortening in a dose-dependent manner; the half-maximal inhibition concentration was 95.97 ± 50.10 nM (n = 3). In addition, the rates of cell shortening and re-lengthening were decreased in ß-CTX treated myocytes concomitant with a prolongation of the intracellular calcium transient decay, indicating depression of cardiac contractility secondary to altered cardiac calcium homeostasis. Conclusion: We present an alternative purification method for ß-CTX from king cobra venom. We reveal cytotoxicity towards smooth muscle and depression of cardiac contractility by this protein. These data are useful to aid future development of pharmacological agents derived from ß-CTX.(AU)

Plantas cardiotóxicas para ruminantes no Brasil / Cardiotoxic plants affecting ruminants in Brazil

Esta revisão atualiza informações sobre plantas cardiotóxicas que afetam os ruminantes no Brasil. Atualmente, sabe-se que existem pelo menos 131 plantas tóxicas pertencentes a 79 gêneros. Vinte e cinco espécies afetam o funcionamento do coração. As plantas que contêm monofluoroacetato de sódio (Palicourea spp., Psychotria hoffmannseggiana, Amorimia spp., Niedenzuella spp., Tanaecium bilabiatum e Fridericia elegans) causam numerosos surtos de intoxicação, principalmente em bovinos, mas búfalos, ovinos e caprinos são ocasionalmente afetados. A intoxicação por Palicourea marcgravii continua a ser a mais importante devido à ampla distribuição desta planta no Brasil. Novas espécies do gênero Palicourea contendo monofluoracetato de sódio, como Palicourea amapaensis, Palicourea longiflora, Palicourea barraensis, Palicourea macarthurorum, Palicourea nigricans, Palicourea vacillans e Palicourea aff. juruana foram descritas na região amazônica. Na região nordeste, a planta tóxica mais importante para bovinos é Amorimia septentrionalis. No Centro-Oeste, surtos de intoxicação por Niedenzuella stannea foram relatados em bovinos na região do Araguaia e a doença precisa ser melhor investigada quanto à sua ocorrência e importância. Tetrapterys multiglandulosa e Tetrapterys acutifolia, duas plantas que causam fibrose cardíaca, também contêm monofluoracetato de sódio e foram reclassificadas para o gênero Niedenzuella. Essas duas espécies e Ateleia glazioveana, outra planta que causa fibrose cardíaca, continuam sendo importantes no Sul e Sudeste do Brasil. Outras espécies menos importantes e que ocasionamente provocam surtos acidentais de intoxicação são as plantas que contém glicosídeos cardiotóxicos, tais como Nerium oleander e Kalanchoe blossfeldiana. Recentemente, várias metodologias experimentais foram empregadas para evitar intoxicações por plantas que contêm monofluoroacetato de sódio. Estas metodologias incluem a indução de aversão condicionada utilizando cloreto de lítio, a utilização de doses repetidas não tóxicas de folhas para induzir resistência, o uso de acetamida para prevenir as intoxicações e a inoculação intraruminal de bactérias degradantes de monofluoroacetato de sódio.(AU)

This review updates information about cardiotoxic plants affecting ruminants in Brazil. Currently it is known that there are at least 131 toxic plants belonging to 79 genera. Twenty five species affect the heart function. Plants that contain sodium monofluoroacetate (Palicourea spp., Psychotria hoffmannseggiana, Amorimia spp., Niedenzuella spp., Tanaecium bilabiatum and Fridericia elegans) cause numerous outbreaks of poisoning, mainly in cattle, but buffaloes, sheep and goats are occasionally affected. Poisoning by Palicourea marcgravii remains the most important due to the wide distribution of this plant in Brazil. New species of the genus Palicourea containing sodium monofluoracetate, such as Palicourea amapaensis, Palicourea longiflora, Palicourea barraensis, Palicourea macarthurorum, Palicourea nigricans, Palicourea vacillans and Palicourea aff. juruana were described in the amazon region. In the northeast region, the most important toxic plant for cattle is Amorimia septentrionalis. In the midwest, outbreaks of Niedenzuella stannea poisoning have been reported in cattle in the Araguaia region and the disease needs to be better investigated for its occurrence and importance. Tetrapterys multiglandulosa and Tetrapterys acutifolia, two plants causing cardiac fibrosis also contain sodium monofluoroacetate and were reclassified to the genus Niedenzuella. These two plants and Ateleia glazioveana, other plant that causes cardiac fibrosis continues to be important in the southeastern and south of Brazil. Other less important are the plants that contain cardiotoxic glycosides, such as Nerium oleander and Kalanchoe blossfeldiana, in wich poisonings are generally accidental. Recently, several experimental methodologies were successfully employed to avoid poisonings by sodium monofluoroacetate containing plants. These methodologies include the induction of food avertion using lithium chloride, the ministration of repeatedly non-toxic doses of leaves to induce resistance, the use of acetamide to prevent poisonings and the intraruminal inoculation of sodium monofluoroacetate degrading bacteria.(AU)

Plantas cardiotóxicas para ruminantes no Brasil / Cardiotoxic plants affecting ruminants in Brazil

Esta revisão atualiza informações sobre plantas cardiotóxicas que afetam os ruminantes no Brasil. Atualmente, sabe-se que existem pelo menos 131 plantas tóxicas pertencentes a 79 gêneros. Vinte e cinco espécies afetam o funcionamento do coração. As plantas que contêm monofluoroacetato de sódio (Palicourea spp., Psychotria hoffmannseggiana, Amorimia spp., Niedenzuella spp., Tanaecium bilabiatum e Fridericia elegans) causam numerosos surtos de intoxicação, principalmente em bovinos, mas búfalos, ovinos e caprinos são ocasionalmente afetados. A intoxicação por Palicourea marcgravii continua a ser a mais importante devido à ampla distribuição desta planta no Brasil. Novas espécies do gênero Palicourea contendo monofluoracetato de sódio, como Palicourea amapaensis, Palicourea longiflora, Palicourea barraensis, Palicourea macarthurorum, Palicourea nigricans, Palicourea vacillans e Palicourea aff. juruana foram descritas na região amazônica. Na região nordeste, a planta tóxica mais importante para bovinos é Amorimia septentrionalis. No Centro-Oeste, surtos de intoxicação por Niedenzuella stannea foram relatados em bovinos na região do Araguaia e a doença precisa ser melhor investigada quanto à sua ocorrência e importância. Tetrapterys multiglandulosa e Tetrapterys acutifolia, duas plantas que causam fibrose cardíaca, também contêm monofluoracetato de sódio e foram reclassificadas para o gênero Niedenzuella. Essas duas espécies e Ateleia glazioveana, outra planta que causa fibrose cardíaca, continuam sendo importantes no Sul e Sudeste do Brasil. Outras espécies menos importantes e que ocasionamente provocam surtos acidentais de intoxicação são as plantas que contém glicosídeos cardiotóxicos, tais como Nerium oleander e Kalanchoe blossfeldiana. Recentemente, várias metodologias experimentais foram empregadas para evitar intoxicações por plantas que contêm monofluoroacetato de sódio. Estas metodologias incluem a indução de aversão condicionada utilizando cloreto de lítio, a utilização de doses repetidas não tóxicas de folhas para induzir resistência, o uso de acetamida para prevenir as intoxicações e a inoculação intraruminal de bactérias degradantes de monofluoroacetato de sódio.(AU)

This review updates information about cardiotoxic plants affecting ruminants in Brazil. Currently it is known that there are at least 131 toxic plants belonging to 79 genera. Twenty five species affect the heart function. Plants that contain sodium monofluoroacetate (Palicourea spp., Psychotria hoffmannseggiana, Amorimia spp., Niedenzuella spp., Tanaecium bilabiatum and Fridericia elegans) cause numerous outbreaks of poisoning, mainly in cattle, but buffaloes, sheep and goats are occasionally affected. Poisoning by Palicourea marcgravii remains the most important due to the wide distribution of this plant in Brazil. New species of the genus Palicourea containing sodium monofluoracetate, such as Palicourea amapaensis, Palicourea longiflora, Palicourea barraensis, Palicourea macarthurorum, Palicourea nigricans, Palicourea vacillans and Palicourea aff. juruana were described in the amazon region. In the northeast region, the most important toxic plant for cattle is Amorimia septentrionalis. In the midwest, outbreaks of Niedenzuella stannea poisoning have been reported in cattle in the Araguaia region and the disease needs to be better investigated for its occurrence and importance. Tetrapterys multiglandulosa and Tetrapterys acutifolia, two plants causing cardiac fibrosis also contain sodium monofluoroacetate and were reclassified to the genus Niedenzuella. These two plants and Ateleia glazioveana, other plant that causes cardiac fibrosis continues to be important in the southeastern and south of Brazil. Other less important are the plants that contain cardiotoxic glycosides, such as Nerium oleander and Kalanchoe blossfeldiana, in wich poisonings are generally accidental. Recently, several experimental methodologies were successfully employed to avoid poisonings by sodium monofluoroacetate containing plants. These methodologies include the induction of food avertion using lithium chloride, the ministration of repeatedly non-toxic doses of leaves to induce resistance, the use of acetamide to prevent poisonings and the intraruminal inoculation of sodium monofluoroacetate degrading bacteria.(AU)

Effects of thymoquinone against cisplatin-induced cardiac injury in rats

PURPOSE: To investigate the possible protective effect of thymoquinone (TQ) in cisplatin (CP) induced myocardial injury.METHODS:A total of 28 adult male Wistar-Albino rats were randomly and equally divided into four groups as follows: Group 1 (control), Group 2 (CP at 15 mg/kg dose), Group 3 (TQ 40 mg/kg/day for two days prior to CP injection and on third day, CP at 15 mg/kg dose was intraperitoneally administered and TQ treatment continued until fifth day) and Group 4 (TQ at 40mg/kg/day dose for five days).RESULTS:There was a significant increment in CP group in terms of congestion, edema and pycnotic nuclei in myocardial fibers, comparing with other groups. TQ group exhibited significant increase in expression of antiapoptotic protein Bcl-2, comparing with CP group (p<0.05). In only CP administered group, expression of antiapoptotic protein Bcl-2 was lowest comparing with other groups.CONCLUSION:Established data indicate that cisplatin is cardiotoxic and thymoquinone may be useful in treating CP-induced cardiac injury.(AU)

Amiodarone may prevent the tilmicosin-caused lethal toxicity

Background: Tilmicosin is widely used in veterinary medicine and its accidental overdose by injection may cause death viacausing negative inotropy and positive chronotropy in both the treated animal and the veterinarian. In addition, there is noany antidote against to tilmicosin-caused death. Amiodarone blocks some channels in the heart, but it has much complexeffect including vagotonic, bradycardic etc on the heart. Considering vagotonic and bradycardic effects of amiodarone, ithas been hypothesised that amiodarone may prevent tilmicosin-caused death. The aim of this study was to determine theeffect of amiodarone on the survival rate of rats in tilmicosin-caused lethal toxicity.Materials, Methods & Results: Twenty female Wistar rats (body weight: 288 ± 33.8 g, age: 7-8 months) were used in thisstudy. The study protocol was approved by the Ethical Committee. Rats received food and water ad libitum. The rats weredivided into two groups containing 10 rats each. Rats in Group 1 were administered 360 mg/kg of tilmicosin in a singlesubcutaneous injection. Rats in Group 2 were administered 25 mg/kg of amiodarone via the tail vein at 8. min after thesingle subcutaneous injection of tilmicosin in a dose of 360 mg/kg. After the injections, deaths were recorded at 0, 2, 6, 10,12 and 24 h. At the end of the 24-h period, survival/death ratio was analysed by the Chi-square test. The level of statisticalsignifi cance was set at P < 0.05. The survival rate of Group 2 (40%) was statistically signifi cantly (P < 0.025) higher thanthat of Group 1 (0.0%). In control group all rats died at 10 h after subcutaneously tilmicosin injection. In Group 2 wereadministered 25 mg/kg of amiodarone (intravenously) at 8 min after the single subcutaneous injection of tilmicosin in adose of 360 mg/kg, and 2 rats died at 2 h and 4...(AU)

Amiodarone may prevent the tilmicosin-caused lethal toxicity

Background: Tilmicosin is widely used in veterinary medicine and its accidental overdose by injection may cause death viacausing negative inotropy and positive chronotropy in both the treated animal and the veterinarian. In addition, there is noany antidote against to tilmicosin-caused death. Amiodarone blocks some channels in the heart, but it has much complexeffect including vagotonic, bradycardic etc on the heart. Considering vagotonic and bradycardic effects of amiodarone, ithas been hypothesised that amiodarone may prevent tilmicosin-caused death. The aim of this study was to determine theeffect of amiodarone on the survival rate of rats in tilmicosin-caused lethal toxicity.Materials, Methods & Results: Twenty female Wistar rats (body weight: 288 ± 33.8 g, age: 7-8 months) were used in thisstudy. The study protocol was approved by the Ethical Committee. Rats received food and water ad libitum. The rats weredivided into two groups containing 10 rats each. Rats in Group 1 were administered 360 mg/kg of tilmicosin in a singlesubcutaneous injection. Rats in Group 2 were administered 25 mg/kg of amiodarone via the tail vein at 8. min after thesingle subcutaneous injection of tilmicosin in a dose of 360 mg/kg. After the injections, deaths were recorded at 0, 2, 6, 10,12 and 24 h. At the end of the 24-h period, survival/death ratio was analysed by the Chi-square test. The level of statisticalsignifi cance was set at P < 0.05. The survival rate of Group 2 (40%) was statistically signifi cantly (P < 0.025) higher thanthat of Group 1 (0.0%). In control group all rats died at 10 h after subcutaneously tilmicosin injection. In Group 2 wereadministered 25 mg/kg of amiodarone (intravenously) at 8 min after the single subcutaneous injection of tilmicosin in adose of 360 mg/kg, and 2 rats died at 2 h and 4...

Biochemical and toxinological characterization of Naja sumatrana (Equatorial spitting cobra) venom

The lethal and enzymatic activities of venom from Naja sumatrana (Equatorial spitting cobra) were determined and compared to venoms from three other Southeast Asian cobras (Naja sputatrix, Naja siamensis and Naja kaouthia). All four venoms exhibited the common characteristic enzymatic activities of Asiatic cobra venoms: low protease, phosphodiesterase, alkaline phosphomonoesterase and L-amino acid oxidase activities, moderately high acetylcholinesterase and hyaluronidase activities and high phospholipase A2. Fractionation of N. sumatrana venom by Resource® S cation exchange chromatography (GE Healthcare, USA) yielded nine major protein peaks, with all except the acidic protein peak being lethal to mice. Most of the protein peaks exhibit enzymatic activities, and L-amino acid oxidase, alkaline phosphomonoesterase, acetylcholinesterase, 5'-nucleotidase and hyaluronidase exist in multiple forms. Comparison of the Resource® S chromatograms of the four cobra venoms clearly indicates that the protein composition of N. sumatrana venom is distinct from venoms of the other two spitting cobras, N. sputatrix (Javan spitting cobra) and N. siamensis (Indochinese spitting cobra). The results support the revised systematics of the Asiatic cobra based on multivariate analysis of morphological characters. The three spitting cobra venoms exhibit two common features: the presence of basic, potentially pharmacologically active phospholipases A2 and a high content of polypeptide cardiotoxin, suggesting that the pathophysiological actions of the three spitting cobra venoms may be similar.(AU)

Dosagem de marcadores cardíacos CK-MB e TnIc e eletrólitos no envenenamento experimental por veneno de sapo em cães / Determination of cardiac markers and CK-MB TnIc and electrolyte in experimental poisoning by poison frog in dogs

Dentre os sinais sistêmicos causados pelo envenenamento por veneno de sapo (bufotoxina) em cães, os efeitos cardiotóxicos são um dos mais importantes. O objetivo deste estudo foi avaliar as potenciais alterações no músculo cardíaco de cães envenenados experimentalmente por veneno de sapo e observar as alterações eletrolíticas que podem ocorrer nesse tipo de envenenamento. Utilizaram-se 20 cães divididos em grupo controle (n=5) e grupo envenenado (n=15). O veneno de sapo foi extraído por meio de compressão manual das glândulas paratóides. Após anestesia geral, os cães do grupo controle receberam placebo (solução fisiológica) e os do grupo envenenado uma alíquota do veneno por sonda orogástrica. As colheitas de sangue para dosagem dos marcadores cardíacos foram realizadas seis e 24 horas após o envenenamento. As colheitas de sangue para dosagem dos eletrólitos foram realizadas antes e duas, quatro, seis e 12 horas após o envenenamento. A análise estatística empregada foi o teste não-paramétrico de Mann-Withney (P<0,05). Os cães envenenados por veneno de sapo apresentaram elevação dos níveis dos marcadores cardíacos CK-MB e TnIc, confirmando a cardiotoxicidade do veneno. Hipocalemia e hipocalcemia foram também observadas nos cães envenenados.(AU)

Among the systemic signs of toad venom (bufotoxin) poisoning in dogs, the cardiotoxic effects are one of the most important. Thus, the objective of this experiment was to evaluate potential changes in the cardiac muscle in dogs poisoned experimentally by toad venom and to observe the eletrolyte alterations which may occur in this condition. Twenty dogs divided into control group (n=5) and poisoned group (n=15) were utilized. The toad venom was extracted by manual compression of the paratoidic glands. After general anesthesia, dogs in the control group received placebo and dogs in the poisoned group received the venom by orogastric catheter. Samples for dosage were collected 6 hours and 24 hours after poisoning and 0, 2, 4, 6 and 12 hours after poisoning for electrolytes dosage. The Man-Withney test was used for statistical analysis (P<0.05). The poisoned dogs showed (saline) elevated levels of cardiac markers CK-MB and TnIc, confirming the cardiotoxic effect of the bufotoxin. Hypokalemia and hypocalcemia were also observed.(AU)