ABSTRACT
Snake envenomation is a common but neglected disease that affects millions of people around the world annually. Among venomous snake species in Brazil, the tropical rattlesnake (Crotalus durissus terrificus) accounts for the highest number of fatal envenomations and is responsible for the second highest number of bites. Snake venoms are complex secretions which, upon injection, trigger diverse physiological effects that can cause significant injury or death. The components of C. d. terrificus venom exhibit neurotoxic, myotoxic, hemotoxic, nephrotoxic, and cardiotoxic properties which present clinically as alteration of central nervous system function, motor paralysis, seizures, eyelid ptosis, ophthalmoplesia, blurred vision, coagulation disorders, rhabdomyolysis, myoglobinuria, and cardiorespiratory arrest. In this study, we focused on proteomic characterization of the cardiotoxic effects of C. d. terrificus venom in mouse models. We injected venom at half the lethal dose (LD50) into the gastrocnemius muscle. Mouse hearts were removed at set time points after venom injection (1 h, 6 h, 12 h, or 24 h) and subjected to trypsin digestion prior to high-resolution mass spectrometry. We analyzed the proteomic profiles of >1300 proteins and observed that several proteins showed noteworthy changes in their quantitative profiles, likely reflecting the toxic activity of venom components. Among the affected proteins were several associated with cellular deregulation and tissue damage. Changes in heart protein abundance offer insights into how they may work synergistically upon envenomation. Significance Venom of the tropical rattlesnake (Crotalus durissus terririficus) is known to be neurotoxic, myotoxic, nephrotoxic and cardiotoxic. Although there are several studies describing the biochemical effects of this venom, no work has yet described its proteomic effects in the cardiac tissue of mice. In this work, we describe the changes in several mouse cardiac proteins upon venom treatment. Our data shed new light on the clinical outcome of the envenomation by C. d. terrificus, as well as candidate proteins that could be investigated in efforts to improve current treatment approaches or in the development of novel therapeutic interventions in order to reduce mortality and morbidity resulting from envenomation.
ABSTRACT
Todo ano, aproximadamente 2,7 milhões de indivíduos são mordidos por serpentes peçonhentas em todo o mundo. Dentre eles, são reportados cerca de 490.000 amputações e 130.000 óbitos bem como outros problemas graves de saúde resultante da mordida por serpentes. No Brasil, a serpente Crotalus durissus popularmente conhecida como cascavel é a única espécie do gênero Crotalus representada em todo o território. Essa serpente é a mais letal e a segunda maior responsável pelos acidentes na população. A subespécie Crotalus durissus terrificus é encontrada predominantemente nas regiões Sul e Sudeste do país e possui uma peçonha conhecida por apresentar efeitos neurotóxicos, nefrotóxicos, miotóxicos, cardiotóxicos e por causar distúrbios hemolíticos e quadros de coagulopatias. Na literatura são encontrados muitos relatos que descrevem os aspectos patológicos, clínicos, toxicológicos, fisiológicos e bioquímicos dos efeitos do envenenamento em vários órgãos, contudo existem poucos registros descrevendo o efeito do envenenamento a nível molecular. Dessa forma, neste projeto foram avaliados os efeitos da peçonha da serpente C. d. terrificus no coração de camundongos de linhagem Swiss 1 h, 6 h, 12 h e 24 h após a aplicação de 0,5 DL50 da peçonha no músculo gastrocnêmio. O coração dos animais foi dissecado e lisado com tampão PTS. As proteínas cardíacas foram então extraídas, modificadas quimicamente (reduzidas e alquiladas), digeridas com tripsina e os peptídeos trípticos resultantes foram marcados com formaldeído leve (grupo controle) e pesado (grupo tratado com a peçonha) e submetidos à análise por espectrometria de massas de alta resolução. Foram identificadas mais de 1300 proteínas das quais várias delas apresentaram alterações no perfil quantitativo ao longo do tempo após o tratamento com a peçonha. A análise do perfil proteico mostrou os efeitos tóxicos da peçonha no coração do camundongo modulando várias proteínas associadas às mitocôndrias e relacionadas com cardiomiopatias hipertrófica e dilatada. As mudanças na abundância dessas proteínas sugerem como elas podem atuar sinergicamente sobre um quadro de envenenamento no tecido cardíaco por meio de distintos efeitos imunológicos e bioquímicos
Every year, approximately 2.7 million individuals are bitten by venomous snakes worldwide. Among them, about 490,000 amputations and 130,000 deaths are reported, as well as other serious health problems resulting from snake bites. In Brazil, Crotalus durissus, popularly known as rattlesnake is the only species of the Crotalus genus found in the entire country. This snake is the most lethal and the second most responsible for accidents in the population. Crotalus durissus terrificus is found predominantly in the South and Southeast of Brazil. Its venom is known for presenting neurotoxic, nephrotoxic, myotoxic and cardiotoxic effects and for causing hemolytic disorders and coagulopathies. Several reports in the literature describe the pathological, clinical, toxicological, physiological, and biochemical aspects of the effects of the envenomation in various organs, however there are few works describing the effect of the venom at the molecular level. In this project, we analyzed the effects of the C. d. terrificus snake venom in the heart of mice of the Swiss strain at 1 h, 6 h, 12 h and 24 h, after the application of 0.5 DL50 of the venom in the gastrocnemius muscle. The animals' hearts were dissected and lysed with PTS buffer. Cardiac proteins were extracted, chemically modified (reduced and alkylated), digested with trypsin and the resulting tryptic peptides were labeled with light (control group) and heavy (venom treated group) dimethyl reagent and subjected to high-resolution mass spectrometry analysis. More than 1300 proteins were identified, several of them showed changes in the quantitative profile over the time after the venom treatment. The venom modulated several proteins associated with mitochondria and related to hypertrophic and dilated cardiomyopathies. Changes in the protein abundance suggests how proteins act synergistically upon envenomation in cardiac tissue, through distinct immunological and biochemical effects.
ABSTRACT
Cancer is characterized by the development of abnormal cells that divide in an uncontrolled way and may spread into other tissues where they may infiltrate and destroy normal body tissue. Several previous reports have described biochemical anti-tumorigenic properties of crude snake venom or its components, including their capability of inhibiting cell proliferation and promoting cell death. However, to the best of our knowledge, there is no work describing cancer cell proteomic changes following treatment with snake venoms. In this work we describe the quantitative changes in proteomics of MCF7 and MDA-MB-231 breast tumor cell lines following treatment with Bothrops jararaca snake venom, as well as the functional implications of the proteomic changes. Cell lines were treated with sub-toxic doses at either 0.63 μg/mL (low) or 2.5 μg/mL (high) of B. jararaca venom for 24 h, conditions that cause no cell death per se. Proteomics analysis was conducted on a nano-scale liquid chromatography coupled on-line with mass spectrometry (nLC-MS/MS). More than 1000 proteins were identified and evaluated from each cell line treated with either the low or high dose of the snake venom. Protein profiling upon venom treatment showed differential expression of several proteins related to cancer cell metabolism, immune response, and inflammation. Among the identified proteins we highlight histone H3, SNX3, HEL-S-156an, MTCH2, RPS, MCC2, IGF2BP1, and GSTM3. These data suggest that sub-toxic doses of B. jararaca venom have potential to modulate cancer-development related protein targets in cancer cells. This work illustrates a novel biochemical strategy to identify therapeutic targets against cancer cell growth and survival.
ABSTRACT
Sea urchins live in a challenging environment that requires rapid and efficient responses against pathogens and invaders. This response may be also important in reproductive processes once males and females release their gametes into water. In addition, the gonads are organs with dual function: reproductive organ and nutrient reserve, therefore it needs efficient protective mechanisms to preserve the nutrients as well as the reproductive cells. The aim of this study was to evaluate the presence and characterize antimicrobial molecules in the male and female gonads of the sea urchin Lytechinus variegatus. Through HPLC purification, antimicrobial activity test and mass spectrometry several antimicrobial molecules were found in the gonads of both gender. Computational in silico analyses showed that they are fragments of a glycoprotein called toposome, also known as major yolk protein (MYP) which is one of the major proteins found in the gonads. Although different functions have been reported for this protein, this is the first description of a direct antimicrobial activity in Lytechinus variegatus. The results indicate that when undergoing proteolysis the toposome generates different fragments with antimicrobial activity which may indicate the importance of a rapid defense response strategy against invading microorganisms in the gonads used by both males and females sea urchins.
ABSTRACT
Snake envenomation is responsible for more than 130,000 deaths worldwide. In Brazil, the Crotalus rattlesnake is responsible for the second largest number of accidental snake bites in the country. Although there are many descriptions of the clinical and biochemical effects of Crotalus envenoming, there are few works describing the molecular events in the central nervous system of an organism due to envenomation. In this study, we analyzed the proteomic effect of Crotalus durissus terrificus snake venom on mice cerebellums. To monitor the envenomation over time, changes in the protein abundance were evaluated at 1 h, 6 h, 12 h and 24 h after venom injection by mass spectrometry. The analysis of the variation of over 4600 identified proteins over time showed a reduction in components of inhibitory synapse signaling, oxidative stress, and maintenance of neuronal cells, which paralleled increasing tissue damage and apoptosis factors. These analyses revealed the potential protein targets of the C. d. terrificus venom on the murine cerebellum, showing new aspects of the snake envenomation effect. These data may contribute to new therapeutic approaches (i.e., approaches directed at protein targets affected by the envenomation) on the treatment of envenomation by the neurotoxic C. d. terrificus snake venom. SIGNIFICANCE: Snakebites are a neglected global health problem that affects mostly rural and tropical areas of developing countries. It is estimated that over 5.4 million people are bitten by snakes each year, from which 2.7 million people are bitten by venomous snakes, resulting in disabilities such as amputations and in some cases leading to death. The C. d. terrificus snake is the most lethal snake in Brazil. Studying the molecular changes upon envenomation in a specific tissue may lead to a better understanding of the envenomation process by C. d. terrificus snakebites.
Subject(s)
Crotalid Venoms , Animals , Brazil , Cerebellum , Crotalid Venoms/toxicity , Crotalus , Mice , ProteomicsABSTRACT
Snake envenomation is responsible for more than 130,000 deaths worldwide. In Brazil, the Crotalus rattlesnake is responsible for the second largest number of accidental snake bites in the country. Although there are many descriptions of the clinical and biochemical effects of Crotalus envenoming, there are few works describing the molecular events in the central nervous system of an organism due to envenomation. In this study, we analyzed the proteomic effect of Crotalus durissus terrificus snake venom on mice cerebellums. To monitor the envenomation over time, changes in the protein abundance were evaluated at 1 h, 6 h, 12 h and 24 h after venom injection by mass spectrometry. The analysis of the variation of over 4600 identified proteins over time showed a reduction in components of inhibitory synapse signaling, oxidative stress, and maintenance of neuronal cells, which paralleled increasing tissue damage and apoptosis factors. These analyses revealed the potential protein targets of the C. d. terrificus venom on the murine cerebellum, showing new aspects of the snake envenomation effect. These data may contribute to new therapeutic approaches (i.e., approaches directed at protein targets affected by the envenomation) on the treatment of envenomation by the neurotoxic C. d. terrificus snake venom. Significance Snakebites are a neglected global health problem that affects mostly rural and tropical areas of developing countries. It is estimated that over 5.4 million people are bitten by snakes each year, from which 2.7 million people are bitten by venomous snakes, resulting in disabilities such as amputations and in some cases leading to death. The C. d. terrificus snake is the most lethal snake in Brazil. Studying the molecular changes upon envenomation in a specific tissue may lead to a better understanding of the envenomation process by C. d. terrificus snakebites.
ABSTRACT
Snake envenomation is responsible for more than 130,000 deaths worldwide. In Brazil, the Crotalus rattlesnake is responsible for the second largest number of accidental snake bites in the country. Although there are many descriptions of the clinical and biochemical effects of Crotalus envenoming, there are few works describing the molecular events in the central nervous system of an organism due to envenomation. In this study, we analyzed the proteomic effect of Crotalus durissus terrificus snake venom on mice cerebellums. To monitor the envenomation over time, changes in the protein abundance were evaluated at 1 h, 6 h, 12 h and 24 h after venom injection by mass spectrometry. The analysis of the variation of over 4600 identified proteins over time showed a reduction in components of inhibitory synapse signaling, oxidative stress, and maintenance of neuronal cells, which paralleled increasing tissue damage and apoptosis factors. These analyses revealed the potential protein targets of the C. d. terrificus venom on the murine cerebellum, showing new aspects of the snake envenomation effect. These data may contribute to new therapeutic approaches (i.e., approaches directed at protein targets affected by the envenomation) on the treatment of envenomation by the neurotoxic C. d. terrificus snake venom. Significance Snakebites are a neglected global health problem that affects mostly rural and tropical areas of developing countries. It is estimated that over 5.4 million people are bitten by snakes each year, from which 2.7 million people are bitten by venomous snakes, resulting in disabilities such as amputations and in some cases leading to death. The C. d. terrificus snake is the most lethal snake in Brazil. Studying the molecular changes upon envenomation in a specific tissue may lead to a better understanding of the envenomation process by C. d. terrificus snakebites.
