Avaliação dos marcadores de estresse oxidativo e defesas antioxidantes em fígado de camundongos após infecção pelo vírus Caraparu.

Programa de Pós-Graduação em Ciências Biológicas. Núcleo de Pesquisas em Ciências Biológicas, Pró-Reitoria de Pesquisa e Pós Graduação, Universidade Federal de Ouro Preto.Submitted by Oliveira Flávia ([email protected]) on 2014-08-20T18:45:42Z No. of bitstreams: 2 license_rdf: 22190 bytes, checksum: 19e8a2b57ef43c09f4d7071d2153c97d (MD5) DISSERTAÇÃO_AvaliaçãoMarcadoresEstresse.pdf: 2490341 bytes, checksum: f36be88a6e75778aefed2902bdd89d55 (MD5)Approved for entry into archive by Gracilene Carvalho ([email protected]) on 2014-08-25T13:26:35Z (GMT) No. of bitstreams: 2 license_rdf: 22190 bytes, checksum: 19e8a2b57ef43c09f4d7071d2153c97d (MD5) DISSERTAÇÃO_AvaliaçãoMarcadoresEstresse.pdf: 2490341 bytes, checksum: f36be88a6e75778aefed2902bdd89d55 (MD5)Made available in DSpace on 2014-08-25T13:26:35Z (GMT). No. of bitstreams: 2 license_rdf: 22190 bytes, checksum: 19e8a2b57ef43c09f4d7071d2153c97d (MD5) DISSERTAÇÃO_AvaliaçãoMarcadoresEstresse.pdf: 2490341 bytes, checksum: f36be88a6e75778aefed2902bdd89d55 (MD5) Previous issue date: 2014O vírus Caraparu (CARV) é membro do grupo C do gênero Orthobunyavirus (família Bunyaviridae). Os vírus do grupo C foram primeiramente descritos na região da Amazônia Brasileira na década de 1950. O CARV foi isolado em 1956 de um macaco sentinela no estado do Pará. Nos países da América do Sul, os bunyavírus do grupo C estão entre os agentes mais comuns de doença febril em humanos e têm causado notáveis surtos nas últimas décadas. Dentre os sintomas da infecção pelo CARV no homem, destacam-se febre, cefaléia, mialgia, náuseas e fraqueza. Já em camundongos, o CARV causa hepatite, o que leva a acreditar que, algum grau de lesão hepática possa também ocorrer nas infecções humanas. Apesar da doença em humanos ser há tanto conhecida, foram poucos os estudos subsequentes pautando esse vírus no que diz respeito a sua patogenia. Estudos têm demonstrado que na patogênese de algumas doenças virais ocorre um aumento no estresse oxidativo, que resumidamente, é um distúrbio no balanço oxidante-antioxidante que pode levar a um potencial dano celular. Muitas células podem tolerar um grau de estresse oxidativo, pois elas possuem antioxidantes tais como a Glutationa, Superóxido Dismutase (SOD) e Catalase (CAT). Portanto, o objetivo deste trabalho foi investigar a patogênese hepática do CARV em camundongos e o possível envolvimento do estresse oxidativo e defesas antioxidantes nessa patologia. Após a infecção subcutânea de camundongos BALB/c, o CARV foi detectado no fígado e a histopatologia revelou hepatite aguda. Elevados níveis séricos de aspartato e alanina aminotransferases (AST/ALT) e alta expressão hepática da citocina pró-inflamatória Fator de Necrose Tumoral Alpha (TNF-α) foram encontrados nos animais infectados. A infecção pelo CARV não alterou os biomarcadores de estresse oxidativo, mas aumentou o conteúdo de Glutationa e alterou a expressão e atividade das enzimas SOD e CAT. Este trabalho é o primeiro que mostra alterações na homeostase oxidativa após infecção pelo CARV e, pode, em parte, ajudar a explicar melhor a patogênese hepática deste vírus, assim como a patogênese de outros membros da família Bunyaviridae.Caraparu virus (CARV) is a member of Group C of the Orthobunyavirus genus (Bunyaviridae family). Group C viruses were firstly described in the brazilian Amazon region during the 1950s. The CARV was first isolated in 1956 from a sentinel monkey in state of Pará. In the countries of South America, bunyavirus Group C are among the common agents of human febrile illness and have caused notable outbreaks in recent decades. Among the symptoms of CARV infection in human stand out fever, headache, myalgia, vomiting and weakness. Already in mice CARV causes hepatitis, which leads to believe that some degree of liver injury may also occur in human infections. Although the disease in humans is already known to a time, few subsequent studies basing this virus regarding its pathogenesis. Studies have shown that in the pathogenesis of some viral diseases there is an increase in oxidative stress that, briefly, is a disturbance in the oxidant-antioxidant balance leading to potential cellular damage. Most cells can tolerate a mild degree of oxidative stress because they have antioxidant molecules such as glutathione, superoxide dismutase (SOD) and catalase (CAT). Therefore, the purpose of this study was to examine the hepatic pathogenesis of CARV in mice and the possible involvement of oxidative stress and antioxidant defenses on this pathology. Following subcutaneous infection of BALB/c mice, CARV was detected in the liver and histopathology revealed acute hepatitis. Increased serum levels of aspartate and alanine aminotransferases (AST/ALT) and greater hepatic expression of the proinflammatory cytokine Tumor Necrosis Factor Alpha (TNF-α) were found in infected animals. The CARV-infection did not alter the biomarkers of oxidative stress but caused increased GSH content and altered expression and activity of SOD and CAT. This is the first report of an alteration of oxidative homeostasis upon CARV infection, which may, in part, explain the hepatic pathogenesis of this virus, as well as the pathogenesis of other Bunyaviridae members

Avalia??o do estresse oxidativo e defesas antioxidantes ap?s infec??o pelo Mayaro virus e prospec??o da atividade antiviral e antioxidante da silimarina.

Programa de P?s-Gradua??o em Ci?ncias Biol?gicas. N?cleo de Pesquisas em Ci?ncias Biol?gicas, Pr?-Reitoria de Pesquisa de P?s Gradua??o, Universidade Federal de Ouro Preto.O Mayaro virus (MAYV) ? um arbov?rus negligenciado, de car?ter emergente, que causa em humanos uma s?ndrome febril, a febre Mayaro, muitas vezes acompanhada por um quadro de artrite/artralgia incapacitante. Apesar de a febre Mayaro ser conhecida por d?cadas, n?o existe terapia ou vacina dispon?vel e, ainda, os mecanismos celulares que contribuem para a patog?nese do MAYV s?o pouco elucidados. Nesse contexto, um grande n?mero de trabalhos tem demostrado que o estresse oxidativo pode contribuir para a patog?nese de uma variedade de agentes virais. O estresse oxidativo ? causado pelo aumento de Esp?cies Reativas de Oxig?nio (ERO) e/ou uma redu??o no sistema de defesa antioxidante. Dentre os principais antioxidantes celulares destacam-se as enzimas Super?xido Dismutase (SOD) e Catalase (CAT) e, dentre os n?o enzim?ticos, a Glutationa. Assim, uma vez que pouco se sabe sobre os mecanismos pelos quais o MAYV induz danos celulares e que o estresse oxidativo pode ser fator chave na patog?nese de infec??es virais, este trabalho teve como objetivo investigar se a infec??o pelo MAYV seria capaz de induzir estresse oxidativo e/ou alterar as defesas antioxidantes. C?lulas hep?ticas humanas, HepG2, foram infectadas e em diferentes horas p?s-infec??o foram avaliados par?metros oxidantes e antioxidantes. A infec??o pelo MAYV levou a um aumento na produ??o de ERO e dos biomarcadores de estresse, malondialde?do e prote?na carbonilada, bem como uma diminui??o da raz?o glutationa reduzida/oxidada (GSH/GSSG). De maneira geral, a infec??o causou um aumento nas defesas antioxidantes. Observamos um aumento nas atividades de SOD e CAT, bem como um aumento no conte?do celular de Glutationa. Como o ambiente redox celular ? influenciado pela produ??o e remo??o de ERO, n?s hipotetizamos que, apesar do aumento nas defesas antioxidantes, uma superprodu??o de ERO foi respons?vel pelo estresse oxidativo causado pela infec??o pelo MAYV. Em seguida, n?s prospectamos se a silimarina, composto originado da planta Silybum marianum, poderia apresentar atividade anti-MAYV, bem como inibir os danos oxidativos gerados pela infec??o. Nossos resultados mostraram que a silimarina possui efetiva atividade antiviral in vitro contra o MAYV, bem como protege as c?lulas do dano oxidativo associado ? infec??o pelo v?rus. Uma vez que a doen?a causada pelo MAYV ? um problema de sa?de p?blica e de car?ter emergente, ampliar os conhecimentos sobre os aspectos relacionados ? infec??o por esse v?rus ? de primordial import?ncia. Coletivamente, os resultados aqui obtidos elucidam alguns mecanismos que contribuem para os danos celulares causados pela infec??o pelo MAYV.Mayaro virus (MAYV) is an arbovirus neglected, emergent that causes in humans a febrile syndrome, Mayaro fever, often accompanied by disabling arthritis/arthralgia. Although Mayaro fever has been known for decades, there is no therapy or vaccine available, and yet the cellular mechanisms that contribute to the pathogenesis of MAYV are poorly elucidated. In this context, a large number of studies have demonstrated that oxidative stress may contribute to the pathogenesis of a variety of viral agents. Oxidative stress is caused by increased Reactive Oxygen Species (ROS) and/or a reduction in the antioxidant defense system. Among the main cellular antioxidants are the enzymes Superoxide Dismutase (SOD) and Catalase (CAT) and, among the non-enzymes, Glutathione. Thus, since little is known about the mechanisms by which MAYV induces cellular damage and that oxidative stress may be a key factor in the pathogenesis of viral infections, this work aimed to investigate if the infection by the MAYV would be able to induce oxidative stress and/or change antioxidant defenses. Human hepatic cells, HepG2, were infected and at different post-infection hours oxidative and antioxidant parameters were evaluated. MAYV infection led to an increase in ROS production and stress biomarkers, malondialdehyde and carbonyl protein, as well as a decrease in the reduced/oxidized glutathione ratio (GSH/GSSG). In general, the infection caused an increase in antioxidant defenses. We observed an increase in the activities of SOD and CAT, as well as an increase in the cellular content of Glutathione. As the cellular redox environment is influenced by the production and removal of ROS, we hypothesized that, despite the increase in antioxidant defenses, an overproduction of ROS was responsible for the oxidative stress caused by the MAYV infection. Next, we investigated whether silymarin, a compound from the Silybum marianum plant, could present anti-MAYV activity, as well as inhibit the oxidative damage generated by the infection. Our results showed that silymarin possesses effective antiviral activity in vitro against MAYV, as well as protects cells from the oxidative damage associated with virus infection. Since the disease caused by the MAYV is a public health problem of an emerging nature, increasing knowledge about the aspects related to the virus infection is of primordial importance. Collectively, the results obtained here elucidate some mechanisms that contribute to the cellular damage caused by the MAYV infection

Avaliação dos marcadores de estresse oxidativo e defesas antioxidantes em fígado de camundongos após infecção pelo vírus Caraparu.

Programa de Pós-Graduação em Ciências Biológicas. Núcleo de Pesquisas em Ciências Biológicas, Pró-Reitoria de Pesquisa e Pós Graduação, Universidade Federal de Ouro Preto.O vírus Caraparu (CARV) é membro do grupo C do gênero Orthobunyavirus (família Bunyaviridae). Os vírus do grupo C foram primeiramente descritos na região da Amazônia Brasileira na década de 1950. O CARV foi isolado em 1956 de um macaco sentinela no estado do Pará. Nos países da América do Sul, os bunyavírus do grupo C estão entre os agentes mais comuns de doença febril em humanos e têm causado notáveis surtos nas últimas décadas. Dentre os sintomas da infecção pelo CARV no homem, destacam-se febre, cefaléia, mialgia, náuseas e fraqueza. Já em camundongos, o CARV causa hepatite, o que leva a acreditar que, algum grau de lesão hepática possa também ocorrer nas infecções humanas. Apesar da doença em humanos ser há tanto conhecida, foram poucos os estudos subsequentes pautando esse vírus no que diz respeito a sua patogenia. Estudos têm demonstrado que na patogênese de algumas doenças virais ocorre um aumento no estresse oxidativo, que resumidamente, é um distúrbio no balanço oxidante-antioxidante que pode levar a um potencial dano celular. Muitas células podem tolerar um grau de estresse oxidativo, pois elas possuem antioxidantes tais como a Glutationa, Superóxido Dismutase (SOD) e Catalase (CAT). Portanto, o objetivo deste trabalho foi investigar a patogênese hepática do CARV em camundongos e o possível envolvimento do estresse oxidativo e defesas antioxidantes nessa patologia. Após a infecção subcutânea de camundongos BALB/c, o CARV foi detectado no fígado e a histopatologia revelou hepatite aguda. Elevados níveis séricos de aspartato e alanina aminotransferases (AST/ALT) e alta expressão hepática da citocina pró-inflamatória Fator de Necrose Tumoral Alpha (TNF-α) foram encontrados nos animais infectados. A infecção pelo CARV não alterou os biomarcadores de estresse oxidativo, mas aumentou o conteúdo de Glutationa e alterou a expressão e atividade das enzimas SOD e CAT. Este trabalho é o primeiro que mostra alterações na homeostase oxidativa após infecção pelo CARV e, pode, em parte, ajudar a explicar melhor a patogênese hepática deste vírus, assim como a patogênese de outros membros da família Bunyaviridae.Caraparu virus (CARV) is a member of Group C of the Orthobunyavirus genus (Bunyaviridae family). Group C viruses were firstly described in the brazilian Amazon region during the 1950s. The CARV was first isolated in 1956 from a sentinel monkey in state of Pará. In the countries of South America, bunyavirus Group C are among the common agents of human febrile illness and have caused notable outbreaks in recent decades. Among the symptoms of CARV infection in human stand out fever, headache, myalgia, vomiting and weakness. Already in mice CARV causes hepatitis, which leads to believe that some degree of liver injury may also occur in human infections. Although the disease in humans is already known to a time, few subsequent studies basing this virus regarding its pathogenesis. Studies have shown that in the pathogenesis of some viral diseases there is an increase in oxidative stress that, briefly, is a disturbance in the oxidant-antioxidant balance leading to potential cellular damage. Most cells can tolerate a mild degree of oxidative stress because they have antioxidant molecules such as glutathione, superoxide dismutase (SOD) and catalase (CAT). Therefore, the purpose of this study was to examine the hepatic pathogenesis of CARV in mice and the possible involvement of oxidative stress and antioxidant defenses on this pathology. Following subcutaneous infection of BALB/c mice, CARV was detected in the liver and histopathology revealed acute hepatitis. Increased serum levels of aspartate and alanine aminotransferases (AST/ALT) and greater hepatic expression of the proinflammatory cytokine Tumor Necrosis Factor Alpha (TNF-α) were found in infected animals. The CARV-infection did not alter the biomarkers of oxidative stress but caused increased GSH content and altered expression and activity of SOD and CAT. This is the first report of an alteration of oxidative homeostasis upon CARV infection, which may, in part, explain the hepatic pathogenesis of this virus, as well as the pathogenesis of other Bunyaviridae members

Implications of oxidative stress on viral pathogenesis.

Reactive species are frequently formed after viral infections. Antioxidant defences, including enzymatic and non-enzymatic components, protect against reactive species, but sometimes these defences are not completely adequate. An imbalance in the production of reactive species and the body?s inability to detoxify these reactive species is referred to as oxidative stress. The aim of this review is to analyse the role of oxidative stress in the pathogenesis of viral infections and highlight some major therapeutic approaches that have gained importance, with regards to controlling virus-induced oxidative injury. Attention will be focused on DNA viruses (papillomaviruses, hepadnaviruses), RNA viruses (flaviviruses, orthomyxoviruses, paramyxoviruses, togaviruses) and retroviruses (human immunodeficiency virus). In general, viruses cause an imbalance in the cellular redox environment, which depending on the virus and the cell can result in different responses, e.g. cell signaling, antioxidant defences, reactive species, and other processes. Therefore, the modulation of reactive species production and oxidative stress potentially represents a novel pharmacological approach for reducing the consequences of viral pathogenesis

Oxidative stress in Mayaro virus infection.

Mayaro virus (MAYV) is a neglected tropical arbovirus that causes a febrile syndrome that is sometimes accompanied by incapacitating arthritis/arthralgia. The pathogenesis of MAYV has not been completely defined and oxidative stress mediated by an increase in reactive oxygen species (ROS) and/or depletion of antioxidant defences has been found to contribute to several aspects of viral disease. To investigate whether MAYV induced oxidative stress in host cells, we monitored ROS production, oxidative stress markers and antioxidant defences at different time points after infection. Our results show that MAYV induced significant oxidative stress in infected HepG2 cells, as indicated by the increase of malondialdehyde (MDA) and protein carbonyl levels, and by a significant decrease of the reduced versus oxidized glutathione (GSH/GSSG) ratio. Generally, MAYV-infected HepG2 cells also showed an increase in antioxidant defences. We observed an increase in the superoxide dismutase (SOD) and catalase (CAT) activities and the total glutathione content. To determine whether similar effects occurred in other cell types, we evaluated the ROS, MDA and SOD activity levels in J774 cells after MAYV infection. Similar to our observations in HepG2 cells, the J774 cells showed an increase in ROS, MDA and total SOD activity following MAYV infection. Thus, since the cellular redox environment is influenced by the production and removal of ROS, we hypothesize that the overproduction of ROS was responsible for the oxidative stress in response to the MAYV infection despite the increase in the antioxidant status. This study is the first report on the involvement of oxidative stress during MAYV infection. Collectively, our data shed light on some mechanisms that are operational in host cells following exposure to MAYV

Antiviral activity of silymarin against Mayaro virus and protective effect in virus-induced oxidative stress.

Mayaro virus (MAYV) is a neglected arbovirus belonging to the family Togaviridae. Its infection leads to Mayaro fever, with clinical manifestations such as fever, myalgia, headache, rash, arthralgia, vomiting, and diarrhea. The most prominent complaint from infected person is the long-lasting arthritis/arthralgia. The treatment for Mayaro fever is mainly symptom-based and there are no vaccines or antiviral drugs currently available, thus, natural products with anti-MAYV activity may provide a potential alternative. Recent evidences suggest that oxidative stress plays an important role in MAYV infection and compounds capable of modulating oxidative stress could represent a novel therapeutic approach in modulating MAYV-associated oxidative cellular damage. Silymarin is a complex extracted of Silybum marianum, or milk thistle, and its major active compound is silybin, which has a remarkable biological effect. Its antioxidant and antiviral effects, including its antiviral activity against the Chikungunya virus (CHIKV), prompted us to think whether silymarin could also reduce the replication of the MAYV and restore the pro-oxidant/antioxidant balance in the context of MAYV infection, leading to reduced cellular oxidative stress. We assessed the antiviral activity and protective effect of silymarin against oxidative stress in MAYV-infected HepG2 cells. Cytopathic effect inhibition, viral replication, and plaque reduction assays were used to determine the anti-MAYV activity of silymarin. Additionally, we determined whether silymarin could reduce MAYV-induced oxidative cell damage. Briefly, silymarin exhibited potent antiviral activity against MAYV and reduced MAYV-induced ROS formation and levels of malondialdehyde (MDA) and carbonyl protein, which are biomarkers of oxidative stress. In conclusion, the ability of silymarin to inhibit MAYV replication and attenuate MAYV-induce oxidative stress warrants further investigation of this compound as a novel therapeutic approach to Mayaro fever disease

Caraparu virus induces damage and alterations in antioxidant defenses in the liver of BALB/c mice after subcutaneous infection.

Oxidative stress is a disturbance in the oxidantantioxidant balance leading to potential cellular damage. Most cells can tolerate a mild degree of oxidative stress because they have a system that counteracts oxidation that includes antioxidant molecules such as glutathione (GSH) and superoxide dismutase (SOD). Disruption of the host antioxidant status has been recognized as an important contributor to the pathogenesis of many viruses. Caraparu virus (CARV) is a member of group C of the Bunyaviridae family of viruses. In South American countries, group C bunyaviruses are among the common agents of human febrile illness and have caused multiple notable outbreaks of human disease in recent decades; nevertheless, little is known about the pathogenic characteristics of these viruses. The purpose of this study was to examine the hepatic pathogenesis of CARV in mice and the involvement of oxidative stress and antioxidant defenses on this pathology. Following subcutaneous infection of BALB/c mice, CARV was detected in the liver, and histopathology revealed acute hepatitis. Increased serum levels of aspartate and alanine aminotransferases (AST/ALT) and greater hepatic expression of the proinflammatory cytokine tumor necrosis factor- a (TNF-a) were found in infected animals. CARV infection did not alter the biomarkers of oxidative stress but caused an increase in GSH content and altered the expression and activity of SOD. This is the first report of an alteration of oxidative homeostasis upon CARV infection, which may, in part, explain the hepatic pathogenesis of this virus, as well as the pathogenesis of other Bunyaviridae members

Mayaro virus induction of oxidative stress is associated with liver pathology in a non-lethal mouse model.

Mayaro virus (MAYV) causes Mayaro fever in humans, a self-limiting acute disease, with persistent arthralgia and arthritis. Although MAYV has a remerging potential, its pathogenic mechanisms remain unclear. Here, we characterized a model of MAYV infection in 3?4-week BALB/c mice. We investigated whether the liver acts as a site of viral replication and if the infection could cause histopathological alterations and an imbalance in redox homeostasis, culminating with oxidative stress. MAYV-infected mice revealed lower weight gain; however, the disease was self-resolving. High virus titre, neutralizing antibodies, and increased levels of aspartate and alanine aminotransferases were detected in the serum. Infectious viral particles were recovered in the liver of infected animals and the histological examination of liver tissues revealed significant increase in the inflammatory infiltrate. MAYV induced significant oxidative stress in the liver of infected animals, as well as a deregulation of enzymatic antioxidant components. Collectively, this is the first study to report that oxidative stress occurs in MAYV infection in vivo, and that it may be crucial in virus pathogenesis. Future studies are warranted to address the alternative therapeutic strategies for Mayaro fever, such as those based on antioxidant compounds