Proteomic analysis reveals rattlesnake venom modulation of proteins associated with cardiac tissue damage in mouse hearts.

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.

Proteomic analysis reveals rattlesnake venom modulation of proteins associated with cardiac tissue damage in mouse hearts

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.

Bothrops Jararaca Snake Venom Modulates Key Cancer-Related Proteins in Breast Tumor Cell Lines.

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.

Bothrops Jararaca snake venom modulates key cancer-related proteins in breast tumor cell lines

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.

The impact of rattlesnake venom on mice cerebellum proteomics points to synaptic inhibition and tissue damage.

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.

The impact of rattlesnake venom on mice cerebellum proteomics points to synaptic inhibition and tissue damage

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.

Variações proteômicas do efeito da peçonha de Bothrops jararaca em linhagens celulares tumorais de mama. / Proteomic variations of the effect of Bothrops jararaca snake venom on breast tumor cells

Cancer is characterized by unnatural cell growth that can result from genetic mutations escaping the apoptosis process. Cancer cells invade adjacent or distant tissues and, added to the process of angiogenesis and metastasis, they clump together to form the tumor. There are currently some papers describing the antitumorigenic biochemical characteristics of snake venom that claim that this type of venom is capable of inhibiting cell proliferation and promoting cell death by different means. However, there is no work characterizing the proteomic molecular effect of the tumor cell lines treatment with snake venom. In this work, we characterized the comparative, functional and quantitative differential proteome of the MCF-7 and MDA-MB231 tumor cell lines treated with Bothrops jararaca venom at different concentrations. Based on data obtained from the cytotoxic assays of B. jararaca venom treatment in these cells we subjected these lineages to low (0.63 μg / mL) and high / subcitotoxicity (2.5 μg / mL) doses of venom for 24 h. The cells were subjected to ice-cold 8M urea cell lysis, reduction and alkylation, trypsin digestion and desalination for analysis by high resolution liquid chromatography coupled to mass spectrometry (nLC-MS / MS). In order to identify, quantify, characterize and compare the functional and biochemical proteins profile in which abundance changed significantly after venom treatment, we used MaxQuant, Perseus and PantherDB, Reactome and String software for data analysis. These analyzes show the differential expression of various proteins, such as histone H3, SNX3, HEL-S-156an, MCC2 and GSTM3. Several of these proteins play important cancer-related roles, such as cell proliferation, invasion, metastasis, apoptosis and stress response. Therefore, these data show that the venom or some of its components have a potential use for cancer therapy and may induce a homeostatic imbalance in cancer cells.

O câncer é caracterizado pelo crescimento não natural de células que podem resultar de mutações genéticas escapando do processo de apoptose. Células cancerosas invadem tecidos adjacentes ou distantes e, somado ao processo de angiogênese e metástase, elas se aglomeram umas sobre as outras formando o tumor. Atualmente, existem alguns trabalhos descrevendo as características bioquímicas anti-tumorigênicas de peçonha de serpentes que afirmam que este tipo de peçonha é capaz de inibir a proliferação celular e promover a morte celular por diferentes meios. Porém, não existe nenhum trabalho caracterizando o efeito molecular proteômico do tratamento de linhagens celulares tumorais com peçonha de serpentes. Neste trabalho, caracterizamos o proteoma diferencial comparativo, funcional e quantitativo das linhagens celulares tumorais MCF-7 e MDA-MB231, tratadas com peçonha de Bothrops jararaca em diferentes concentrações. Baseado nos dados obtidos dos ensaios citotóxicos do tratamento com a peçonha de B. jararaca nestas células, submetemos essas linhagens a doses baixas (0,63 μg/mL) e altas / sub-citotóxica (2,5 μg/mL) da peçonha por 24 h. As células foram submetidas à lise celular com ureia 8M gelada, redução e alquilação, digestão com tripsina e dessalinização para análise por nano-cromatografia líquida de alta resolução acoplada à espectrometria de massas (nLC-MS/MS). De modo a identificar, quantificar, caracterizar e comparar o perfil funcional e bioquímico das proteínas em que a abundância mudou significativamente após o tratamento com a peçonha, foram utilizados os softwares MaxQuant, Perseus e as ferramentas PantherDB, Reactome e String para análise dos dados. Essas análises mostram a expressão diferencial de diversas proteínas, tais como a histona H3, SNX3, HEL-S-156an, MCC2 e GSTM3. Várias dessas proteínas desempenham papéis importantes relacionados ao câncer, como proliferação celular, invasão, metástase, apoptose e resposta ao estresse. Portanto, esses dados mostram que a peçonha ou alguns de seus componentes possuem um potencial de uso para a terapia do câncer, podendo induzir um desequilíbrio homeostático nas células cancerosas.