Placental vessels and the umbilical circulatory network function to carry oxygen and nutrients to the fetus. It is at this level that placental lesions such as villitis, obliterative vasculopathy, and thrombotic vasculopathy have been observed in association with fetal inflammatory response syndrome (FIRS) and cerebral palsy. We used human umbilical vein endothelial cells (HUVECs) as a model to study the regulation of inflammation and thrombosis in fetal vessels by microbial products. In this thesis we measured interleukin-8 (IL-8) and tissue factor (TF) expression by HUVECs treated with lipopolysaccharide (LPS), poly (I:C) (PIC), and peptidoglycan (PG). Our results show a profound induction of IL-8 by PIC, a TLR-3 ligand. We also show a moderate induction of tissue factor expression in PIC-treated HUVECs. These results show that HUVECs are exquisitely sensitive to PIC and suggests an important role for viral infection in umbilical vessel inflammation. We additionally treated HUVECs with dexamethasone (DEX), an anti-inflammatory steroid, and melatonin (MT), a pineal gland product with immunomodulatory and anti-oxidant properties. DEX reduced the level of both IL-8 and TF expression in PIC-treated cells. MT, however, further enhanced IL-8 expression in PIC-treated cells. Our results indicate a potential role for glucocorticoid therapy in reducing placental vessel inflammation and thrombosis. Thus, intervention with GC in pregnancies with FIRS may reduce the severity of placental lesions associated with cerebral palsy.

One unique feature of cancer cells that can be exploited for anticancer drug discovery is their dependence on their own cellular stress responses to survive the stressful acidotic, hypoxic and nutrient-deprived conditions within the tumor. Reasoning that desert organisms surviving under stressful conditions may have evolved to produce small molecule metabolites capable of modulating heat shock protein 90 (Hsp90) function, and/or other cell stress responses, we employed the cellular heat shock response in a moderate-throughput phenotypic assay. This strategy has resulted in the isolation and characterization of a number of small molecule natural products with heat shock induction activity from these organisms. Three such natural products are the subject of this study.In a limited structure-activity relationship (SAR) study, a previously known Hsp90 inhibitor radicicol (RAD), and several structurally related molecules including the fungal metabolite monocillin 1 (MON) were found to interact with Hsp90. In addition, RAD and MON were shown to lead to the degradation of Hsp90 client proteins involved in the cancer cell survival the estrogen receptor (ER) and the insulin-like growth factor receptor 1 (IGF-1R).We further characterized MON and showed that by targeting the molecular chaperone Hsp90, this compound induces components of the heat shock response at the transcriptional and translational levels, and leads to the acquisition of a thermotolerant phenotype in seedlings of the plant Arabidopsis thaliana. These findings support our hypothesis that there is ecological significance to the elaboration of small molecules that target stress responses.A number of extracts active in our phenotypic assay contained small molecules with no apparent Hsp90 activity. One such extract afforded terrecyclic acid A (TCA) with significant anti-tumor activity against a panel of human cancer cell lines. To characterize the biological activities of TCA we examined three key stress responsesthe heat shock, oxidative, and inflammatory responsesand show that TCA destabilizes these pathways associated with cancer cell survival through induction of oxidative stress (ROS), and inhibition of NF-kappaB transactivation.The isolation of RAD, MON and TCA from Sonoran desert organisms provides proof of principle that we have developed an effective strategy for the discovery of small molecule modulators of cellular stress responses that can serve as leads for the development of new anticancer drugs with novel mechanisms of action.

No description available.

Burn injuries represent a major cause of mortality and morbidity as well as a significant drain on limited resources, particularly in the developing world. Advances in resuscitation, critical care, protective ventilatory strategies, earlier complete debridement complemented by more aggressive treatment of burn wound sepsis have reduced the mortality of thermal injuries. There has also been a move to focus on education and prevention campaigns, which have borne fruit and resulted in some reduction in the incidence of burns in the paediatric population. Burn care, once a Cinderella specialty, has become a well focused multi-disciplinary specialty in its own right. Burn injury is dissimilar to many forms of trauma. In major burns, the initial trauma may be limited to the skin alone or skin and lungs, but all organ systems are rapidly involved as the physiological derangement becomes systemic. The burden of this multi-system insult is substantial. Globally, the World Health Organisation estimated that fire-associated burns alone directly resulted in over 320,000 deaths in 2002, and in the USA, approximately 1million children sustain burns each year. Australasian guidelines suggest that all adult burns with greater than 15% total body surface area (TBSA) and >10% TBSA burn in children will require fluid resuscitation and possibly critical care support at some point in their hospitalisation. Mortality in these patients with severe burns follows a bimodal pattern of early and late deaths. Causes of early death comprise refractory shock, inability to obtain a safe airway or provide adequate oxygenation, co-existent trauma, non-survivable carboxyhaemoglobin poisoning, and decisions that injuries are non-compatible with recovery, leading to therapy withdrawal. With improved resuscitation strategies, 95% of patients survive the early resuscitation phase. Late deaths are secondary to sepsis normally associated with wound infection and multiple organ failure. The morbidity associated with burn injury continues well after the acute hospital admission, frequently for up to several years post burn injury, as is witnessed by prolonged rise in basal metabolic rate and worsening scar tissue1, 2. It has been calculated that in children approximately 60% of the cost of burn care occurs post wound closure; that is, dealing with the disabling and disfiguring contractures associated with hypertrophic scar3, 4. Hence, research and new modalities are being aimed at reducing cost of treatment and improving quality of life for survivors of burn injury. Thus, this PhD aims to reflect the multidisciplinary nature of modern day burn care, with the inclusion of seven published papers and one book chapter covering prevention and education relevant to paediatric burns, treatment and minimisation of wound infection , and scar minimisation, along with the first study into the relevance of fetal wound healing post burn injury.

Conselho Nacional de Desenvolvimento Científico e Tecnológico / Sono e imunidade parecem apresentar uma relação de reciprocidade. A ativação do sistema imune altera o padrão de sono e distúrbios do sono podem afetar a função imune. Além disso, é bem descrito que a privação de sono paradoxal (PSP) leva à hiperalgesia e o tratamento com fármacos clássicos, como opióides ou antidepressivos tricíclicos, não é capaz de reverter este quadro. Neste trabalho, avaliamos se a PSP afetaria a resposta inflamatória e a sobrevida em ratos e se o tratamento com um análogo sintético de lipoxinas (ATL-1) seria capaz de reverter a hiperalgesia induzida pela PSP. Todos os protocolos experimentais foram previamente aprovados pelo Comitê de Ética para o Uso de Animais, da UERJ (CEUA/032/2010). Ratos Wistar machos foram submetidos a 96 h de PSP, induzidas pelo método de plataforma única (PU) ou de múltiplas plataformas modificado (MPM). Após 96 h de PSP os animais foram submetidos ao modelo da bolha de ar ou pleurisia utilizando-se a carragenina como agente flogístico, ou ainda a PSP foi aplicada antes ou após a indução de um modelo de ligação e perfuração do ceco (CLP). Quatro horas após a injeção de carragenina os animais apresentaram um aumento no recrutamento de leucócitos para a cavidade da bolha, porém não houve diferença entre animais PSP e controles. O número total de leucócitos no plasma não se alterou após a injeção de carragenina. Na pleurisia, os animais PSP apresentaram um aumento nos níveis de IL-6, IL-1β e TNF-α no plasma, enquanto apenas IL-1β e IL-6 estavam aumentados no exsudato pleural dos animais que receberam carragenina. O padrão de recrutamento de leucócitos para o local da injúria foi bastante semelhante entre os animais controle e PSP 2 h, 4 h e 24 h após a injeção de carragenina. Houve um aumento progressivo com o tempo, apresentando um pico em 24 h, no entanto, não foi observada diferença significativa na resposta dos grupos PSP. A PSP aplicada antes ou após a indução do CLP reduziu a sobrevida dos animais, mas não alterou o acúmulo de neutrófilos, nos dois protocolos. Quando a PSP foi aplicada antes do CLP, os níveis séricos de IL-6 estavam aumentados nos grupos PSP e PSPCLP, porém quando a PSP foi aplicada após o CLP, ambas IL-6 e IL-1β estavam aumentadas nos grupo PSPCLP. O efeito do tratamento com ATL-1 (10 g/kg, i.v.) na hiperalgesia induzida pela PSP foi determinado através do teste da formalina. O análogo reduziu o número de comportamentos relacionados à dor em animais PSP e controles na fase inflamatória do teste. Nossos resultados demonstraram que a PSP por 96 h aumentou os níveis plasmáticos de citocinas, reduziu a sobrevida dos animais, contudo não foi capaz de alterar o recrutamento de leucócitos frente a um estímulo inflamatório ou infeccioso. O aumento de mediadores inflamatórios observado nesses animais pode estar relacionado à hiperalgesia em animais PSP, uma vez que o tratamento com o ATL-1 reverteu esse efeito, possivelmente através de mecanismos envolvendo sua ação anti-inflamatória. / Sleep and immunity show a reciprocal relationship. Immune system activation alters sleep pattern and sleep disturbances can affect immune function. Moreover, it is well known that paradoxical sleep deprivation (PSD) leads to hyperalgesia and the treatment with classical drugs like opioids or tricyclic antidepressants is not able to reverse this hyperalgesia. In this work we investigated whether PSD could affect inflammatory response and survival in rats and if ATL-1 treatment would be able to reverse the hyperalgesia induced by PSD. All experimental protocols were previously approved by The Animal Care Ethical Committee of UERJ (CEUA/032/2010). Male Wistar rats were submitted to 96 h of PSD by single platform or modified multiple platforms methods. After 96 h of PSD animals were submitted to carrageenan-induced air pouch or pleurisy, or PSD was induced prior or after cecal ligation and puncture model (CLP). Animals presented an increase in leukocyte recruitment to the pouch cavity 4 h after carrageenan injection, however there was no difference between PSD and controls. The number of plasma leukocyte did not change after carrageenan injection. PSD animals submitted to pleurisy showed an increase in IL-6, IL-1β e TNF-α plasma levels, while IL-1β and IL-6 were increased in pleural exsudate of animals that received carrageenan. Leukocyte recruitment pattern to the site of injury was similar between controls and PSD 2 h, 4 h and 24 h after carrageenan injection. There was a progressive increase with time, reaching the maximum point at 24 h, but no differences were observed in PSD groups. PSD induced prior or after CLP decreased animals survival, however no difference was observed on neutrophil accumulation in both protocols. When PSD was induced prior CLP, IL-6 plasma levels were increased in PSD e PSDCLP groups, when PSD was induced after CLP, IL-6 and IL-1β plasma levels were increased in PSDCLP group. The effect of ATL-1 treatment (10 g/kg, i.v.) on hyperalgesia induced by PSD was determined through formalin test. The treatment reduced the number of pain related behaviors in PSD animals and controls on inflammatory phase. Our results show that ATL-1 was able to revert the hyperalgesia induced by PSD possibly through its anti-inflammatory action. Furthermore, PSD for 96 h increased cytokine plasma levels and reduced survival, however it was not able to modify leukocyte recruitment when challenged by an inflammatory or infectious stimulus. However the inflammatory mediators increase observed in PSD animals could be related to hyperalgesia since treatment with ATL-1 reverted this effect, possibly through anti-inflammatory mechanisms.

Made available in DSpace on 2014-06-11T19:31:29Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-02-06Bitstream added on 2014-06-13T18:42:09Z : No. of bitstreams: 1 cavalheiro_js_dr_botfm.pdf: 355047 bytes, checksum: d0af4a219752a076dbe64c1332627767 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Universidade Estadual Paulista (UNESP) / A Paracoccidioidomicose é uma micose sistêmica humana causada por um fungo imperfeito e dimórfico denominado Paracoccidioides brasilienis (P.brasiliensis), sendo os conídeos sua forma infectante. Camundongos geneticamente selecionados para Resposta Inflamatória Aguda Máxima (AIRmax) e Mínima (AIRmin), foram usados como modelo experimental da paracoccidioidomicose. Os animais foram inoculados pela via intraperitoneal com P. brasiliensis (cepa 18) e sacrificados após 6, 12 e 24 horas e 3, 7 e 14 dias de infecção. Nestes períodos foram removidos fragmentos de baço, fígado e pulmão para avaliação do grau de infecção de células fúngicas nestes órgãos, avaliação da atividade macrofágica por macrófagos peritoneais e esplênicos pela determinação das concentrações de óxido nítrico e produção de citocinas pró e anti-inflamatórias de sobrenadantes de homogenatos de pulmão e baço. No presente trabalho observamos que as linhagens AIRmax apresentaram maior contenção do processo infeccioso que as linhagens AIRmin nos compartimentos analisados. Quanto a produção de NO observamos que animais AIRmax produziram mais deste metabólito em períodos tardios, o que pode ter ajudado esta linhagem na maior contenção do processo infeccioso nos órgãos analisados. Quanto a produção de citocinas observamos que a produção de INF- , TNF- , IL-1, IL- 6, IL-8, IL-12, estão aumentadas nas linhagens AIRmax na maioria dos órgãos e períodos analisados, contribuindo para a maior resistência destas linhagens frente à infecção pelo fungo em relação aos AIRmin, com exceção da IL-4 e IL-10 que apresentaram sua produção diminuída na linhagem AIRmax e aumentada na linhagem AIRmin reproduzindo seu efeito biológico supressor. Frente aos resultados observamos que a linhagem AIRmax apresentou maior contenção do processo infeccioso, com importante envolvimento... / Paracoccidioidomycosis is a systemic human mycosis caused by Paracoccidioides brasiliensis (P. brasiliensis), an imperfect dimorphic fungus whose conidia are its infectant form.Mice genetically selected for Maximum (AIRmax) and Minimum (AIRmin) Acute Inflammatory Response were used as an experimental paracoccidioidomycosis model. The animals were intraperitoneally inoculated with P. brasiliensis (strain 18) and sacrificed after 6, 12 and 24 hours and after 3, 7 and 14 days of infection. In these periods, fragments from their spleen, liver and lung were removed for evaluation of the level of infection by fungal cells in those organs, evaluation of macrophagic activity by peritoneal and splenic macrophages through the determination of the concentrations of nitric oxide (NO) and production of pro- and anti-inflammatory cytokines of lung and spleen homogenate supernatants. In the present study, it was observed that the AIRmax lineages presented greater control of the infectious process than did the AIRmin lineages in the analyzed compartments. As regards NO production, it was found that AIRmax animals produced more of such metabolite in late periods, which may have helped that lineage with the greater control of the infectious process in the analyzed organs. Concerning cytokine production, it was observed that the production of INF- , TNF- , IL-1, IL-6, IL-8 and IL-12 were increased for the AIRmax lineages in most analyzed organs and periods, thus contributing to the greater resistance exhibited by such lineages against infection with the fungus in relation to AIRmin, except for IL-4 and IL-10, which showed decreased production for the AIRmax lineage, reproducing its suppressive biological effect. From these results, it was observed that the AIRmax lineage showed greater control over the infectious process with an important involvement of the analyzed cytokines. These findings... (Complet abstract click electronic access below)

Submitted by FERNANDA CRISTINA WINCKLER null (fwinckler@fmb.unesp.br) on 2016-11-30T19:00:44Z No. of bitstreams: 1 DISSERTAÇÃO FINAL 30.11.16.pdf: 468887 bytes, checksum: 372ef2c27b73313e5608616205ed1d2e (MD5) / Approved for entry into archive by Felipe Augusto Arakaki (arakaki@reitoria.unesp.br) on 2016-12-02T16:28:26Z (GMT) No. of bitstreams: 1 winckler_fc_me_bot.pdf: 468887 bytes, checksum: 372ef2c27b73313e5608616205ed1d2e (MD5) / Made available in DSpace on 2016-12-02T16:28:26Z (GMT). No. of bitstreams: 1 winckler_fc_me_bot.pdf: 468887 bytes, checksum: 372ef2c27b73313e5608616205ed1d2e (MD5) Previous issue date: 2016-10-19 / A hepatite C é uma doença infecciosa que torna-se crônica em cerca de 85% dos infectados que poderão desenvolver cirrose e carcinoma hepato celular. O tratamento antiviral em muitos dos pacientes não é eficaz, principalmente quando estes portam o genótipo 1 e fibrose avançada, a resposta inflamatória também desempenha seu papel sobre a resposta virológica sustentada (RVS) durante o tratamento com Interferon Peguilado (PegIFN) associado a Ribavirina (RBV). Nesse estudo nosso objetivo principal foi avaliar a influência da resposta inflamatória através de células e citocinas/quimiocinas sobre a resposta virológica do paciente em tratamento antiviral com terapia tripla. Incluimos pacientes com RNA VHC+, nunca tratados (naive), portadores do genótipo 1, ambos os sexos e com fibrose avançada F3 (n=6); F4 (n=21) candidatos ao tratamento em regime triplo. Os pacientes tiveram suas amostras coletadas e analizadas nas semanas 0 e 12 do tratamento e os seguintes parâmetros foram analisados: IL-2, IL-4, IL-6, IL-8, IL-10, IL-17A, TNF-α, IFN-γ, RANTES, MCP-1, MIG, IP- 10, através de citometria de fluxo (método CBA). Foram incluídos 15 voluntários saudáveis (grupo controle) e 27 pacientes que foram separados em G1(RVS) e G2 (não RVS), a taxa de RVS foi de 63%. Os pacientes com hepatite C crônica tiveram os níveis circulantes de IP10, MCP-1, MIG, RANTES, IL-8 e IL-6 mais elevados quando comparados com voluntários saudáveis, quando comparados G1xG2 os níveis de RANTES (p=0,04) e IL-6 (p=0,02) foram associadas com a RVS na semana 0, seus níveis eram mais baixos em G1, na semana 12 os níveis de RANTES (p=0,04) e IL-8 (n=0,01) foram associados com a RVS, seus níveis são mais elevados em G2, a comparação entre as semanas 0 e 12 mostrou que em G1 os níveis de IL6 (p= 0,02) e MCP-1 (p=0,001) apresentam associação com o tratamento e em G2 os parâmetros associados ao tratamento foram RANTES (p=0,05) e MCP-1 (p=0,01). Os resultados sugerem que, a citocina IL-6 e a quimiocina RANTES estão associadas com a RVS na semana 0. Na semana 12, RANTES assim como IL-8 influenciam na RVS durante terapia antiviral em regime triplo. Quando comparado semana 0 e 12 em pacientes RVS, a citocina IL-6 está associada ao tratamento. Em pacientes não RVS, RANTES esta associada ao tratamento e MCP-1 está associada ao tratamento independente da resposta obtida. / Hepatitis C is an infectious disease which becomes chronic in about 85% of infected people who can develop cirrhosis and hepatocellular carcinoma. Antiviral therapy isn’t effective in many patients, especially when these patients are genotype 1 and have advanced fibrosis, the inflammatory response also plays a role on sustained virological response (SVR) during treatment with Pegylated (PegIFN) plus Ribavirin (RBV). The aim of this study was evaluate the influence of the inflammatory response by cells and cytocines/chemokines on the virologic response of the patients under antiviral treatment with triple therapy. We included patients with HCV RNA+, naive, genotype 1, both male and female and with advanced fibrosis F3 (n=6); F4 (n=21) for triple treatment regimen. Patients had their samples collected and analyzed at weeks 0 and 12 of treatment and the following parameters were analyzed: IL- 2, IL-4, IL-6, IL-8, IL-10, IL-17A, TNF-α, IFN-γ RANTES, MCP-1, MIG, IP-10 by flow cytometry (CBA method). Control group of 15 healthy volunteers and 27 patients, who were separated into GI (SVR) and G2 (not SRV), were included, the SVR rate was 63%. Patients with chronic hepatitis C had higher circulating levels of IP10, MCP-1, MIG, RANTES, IL-8 and IL-6 compared with healthy volunteers, when G1xG2 were compared, levels of RANTES (p=0,040 and IL-6 (n=0,02) were associated with a SVR at week 0 and its levels were lower in G1; at week 12, levels of RANTES (p=0,04) and IL-8 (p=0,01) were associated with a SVR and its levels were higher in G2. The comparison between weeks 0 and 12 showed that, in G1, the IL6 levels (p = 0.02) and MCP-1 (p = 0.001) were associated with the treatment and in G2, the parameters associated with the treatment were RANTES (p = 0.05) and MCP-1 (p = 0.01). The results suggest that the cytocine IL-6 and chemokine RANTES are associated with SVR at week 0. At week 12, RANTES as well as IL-8 influence in SVR during antiviral therapy in triple regimen. When weeks 0 and 12 in patients SVR are compared, the cytocine IL-6 is associated with treatment. In non-SVR patients, RANTES is associated with treatment and MCP-1 is associated with independent treatment of the patient's response.

Orientador: Silvio Luis de Oliveira / Banca: Maria Terezinha Serrão Peraçoli / Banca: Eduardo Bagagli / Banca: Luis Florêncio Franco Margatho / Banca: Carlos Alberto Sorgi / Resumo: A Paracoccidioidomicose é uma micose sistêmica humana causada por um fungo imperfeito e dimórfico denominado Paracoccidioides brasilienis (P.brasiliensis), sendo os conídeos sua forma infectante. Camundongos geneticamente selecionados para Resposta Inflamatória Aguda Máxima (AIRmax) e Mínima (AIRmin), foram usados como modelo experimental da paracoccidioidomicose. Os animais foram inoculados pela via intraperitoneal com P. brasiliensis (cepa 18) e sacrificados após 6, 12 e 24 horas e 3, 7 e 14 dias de infecção. Nestes períodos foram removidos fragmentos de baço, fígado e pulmão para avaliação do grau de infecção de células fúngicas nestes órgãos, avaliação da atividade macrofágica por macrófagos peritoneais e esplênicos pela determinação das concentrações de óxido nítrico e produção de citocinas pró e anti-inflamatórias de sobrenadantes de homogenatos de pulmão e baço. No presente trabalho observamos que as linhagens AIRmax apresentaram maior contenção do processo infeccioso que as linhagens AIRmin nos compartimentos analisados. Quanto a produção de NO observamos que animais AIRmax produziram mais deste metabólito em períodos tardios, o que pode ter ajudado esta linhagem na maior contenção do processo infeccioso nos órgãos analisados. Quanto a produção de citocinas observamos que a produção de INF- , TNF- , IL-1, IL- 6, IL-8, IL-12, estão aumentadas nas linhagens AIRmax na maioria dos órgãos e períodos analisados, contribuindo para a maior resistência destas linhagens frente à infecção pelo fungo em relação aos AIRmin, com exceção da IL-4 e IL-10 que apresentaram sua produção diminuída na linhagem AIRmax e aumentada na linhagem AIRmin reproduzindo seu efeito biológico supressor. Frente aos resultados observamos que a linhagem AIRmax apresentou maior contenção do processo infeccioso, com importante envolvimento... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Paracoccidioidomycosis is a systemic human mycosis caused by Paracoccidioides brasiliensis (P. brasiliensis), an imperfect dimorphic fungus whose conidia are its infectant form.Mice genetically selected for Maximum (AIRmax) and Minimum (AIRmin) Acute Inflammatory Response were used as an experimental paracoccidioidomycosis model. The animals were intraperitoneally inoculated with P. brasiliensis (strain 18) and sacrificed after 6, 12 and 24 hours and after 3, 7 and 14 days of infection. In these periods, fragments from their spleen, liver and lung were removed for evaluation of the level of infection by fungal cells in those organs, evaluation of macrophagic activity by peritoneal and splenic macrophages through the determination of the concentrations of nitric oxide (NO) and production of pro- and anti-inflammatory cytokines of lung and spleen homogenate supernatants. In the present study, it was observed that the AIRmax lineages presented greater control of the infectious process than did the AIRmin lineages in the analyzed compartments. As regards NO production, it was found that AIRmax animals produced more of such metabolite in late periods, which may have helped that lineage with the greater control of the infectious process in the analyzed organs. Concerning cytokine production, it was observed that the production of INF- , TNF- , IL-1, IL-6, IL-8 and IL-12 were increased for the AIRmax lineages in most analyzed organs and periods, thus contributing to the greater resistance exhibited by such lineages against infection with the fungus in relation to AIRmin, except for IL-4 and IL-10, which showed decreased production for the AIRmax lineage, reproducing its suppressive biological effect. From these results, it was observed that the AIRmax lineage showed greater control over the infectious process with an important involvement of the analyzed cytokines. These findings... (Complet abstract click electronic access below) / Doutor

Conselho Nacional de Desenvolvimento Científico e Tecnológico / Sono e imunidade parecem apresentar uma relação de reciprocidade. A ativação do sistema imune altera o padrão de sono e distúrbios do sono podem afetar a função imune. Além disso, é bem descrito que a privação de sono paradoxal (PSP) leva à hiperalgesia e o tratamento com fármacos clássicos, como opióides ou antidepressivos tricíclicos, não é capaz de reverter este quadro. Neste trabalho, avaliamos se a PSP afetaria a resposta inflamatória e a sobrevida em ratos e se o tratamento com um análogo sintético de lipoxinas (ATL-1) seria capaz de reverter a hiperalgesia induzida pela PSP. Todos os protocolos experimentais foram previamente aprovados pelo Comitê de Ética para o Uso de Animais, da UERJ (CEUA/032/2010). Ratos Wistar machos foram submetidos a 96 h de PSP, induzidas pelo método de plataforma única (PU) ou de múltiplas plataformas modificado (MPM). Após 96 h de PSP os animais foram submetidos ao modelo da bolha de ar ou pleurisia utilizando-se a carragenina como agente flogístico, ou ainda a PSP foi aplicada antes ou após a indução de um modelo de ligação e perfuração do ceco (CLP). Quatro horas após a injeção de carragenina os animais apresentaram um aumento no recrutamento de leucócitos para a cavidade da bolha, porém não houve diferença entre animais PSP e controles. O número total de leucócitos no plasma não se alterou após a injeção de carragenina. Na pleurisia, os animais PSP apresentaram um aumento nos níveis de IL-6, IL-1β e TNF-α no plasma, enquanto apenas IL-1β e IL-6 estavam aumentados no exsudato pleural dos animais que receberam carragenina. O padrão de recrutamento de leucócitos para o local da injúria foi bastante semelhante entre os animais controle e PSP 2 h, 4 h e 24 h após a injeção de carragenina. Houve um aumento progressivo com o tempo, apresentando um pico em 24 h, no entanto, não foi observada diferença significativa na resposta dos grupos PSP. A PSP aplicada antes ou após a indução do CLP reduziu a sobrevida dos animais, mas não alterou o acúmulo de neutrófilos, nos dois protocolos. Quando a PSP foi aplicada antes do CLP, os níveis séricos de IL-6 estavam aumentados nos grupos PSP e PSPCLP, porém quando a PSP foi aplicada após o CLP, ambas IL-6 e IL-1β estavam aumentadas nos grupo PSPCLP. O efeito do tratamento com ATL-1 (10 g/kg, i.v.) na hiperalgesia induzida pela PSP foi determinado através do teste da formalina. O análogo reduziu o número de comportamentos relacionados à dor em animais PSP e controles na fase inflamatória do teste. Nossos resultados demonstraram que a PSP por 96 h aumentou os níveis plasmáticos de citocinas, reduziu a sobrevida dos animais, contudo não foi capaz de alterar o recrutamento de leucócitos frente a um estímulo inflamatório ou infeccioso. O aumento de mediadores inflamatórios observado nesses animais pode estar relacionado à hiperalgesia em animais PSP, uma vez que o tratamento com o ATL-1 reverteu esse efeito, possivelmente através de mecanismos envolvendo sua ação anti-inflamatória. / Sleep and immunity show a reciprocal relationship. Immune system activation alters sleep pattern and sleep disturbances can affect immune function. Moreover, it is well known that paradoxical sleep deprivation (PSD) leads to hyperalgesia and the treatment with classical drugs like opioids or tricyclic antidepressants is not able to reverse this hyperalgesia. In this work we investigated whether PSD could affect inflammatory response and survival in rats and if ATL-1 treatment would be able to reverse the hyperalgesia induced by PSD. All experimental protocols were previously approved by The Animal Care Ethical Committee of UERJ (CEUA/032/2010). Male Wistar rats were submitted to 96 h of PSD by single platform or modified multiple platforms methods. After 96 h of PSD animals were submitted to carrageenan-induced air pouch or pleurisy, or PSD was induced prior or after cecal ligation and puncture model (CLP). Animals presented an increase in leukocyte recruitment to the pouch cavity 4 h after carrageenan injection, however there was no difference between PSD and controls. The number of plasma leukocyte did not change after carrageenan injection. PSD animals submitted to pleurisy showed an increase in IL-6, IL-1β e TNF-α plasma levels, while IL-1β and IL-6 were increased in pleural exsudate of animals that received carrageenan. Leukocyte recruitment pattern to the site of injury was similar between controls and PSD 2 h, 4 h and 24 h after carrageenan injection. There was a progressive increase with time, reaching the maximum point at 24 h, but no differences were observed in PSD groups. PSD induced prior or after CLP decreased animals survival, however no difference was observed on neutrophil accumulation in both protocols. When PSD was induced prior CLP, IL-6 plasma levels were increased in PSD e PSDCLP groups, when PSD was induced after CLP, IL-6 and IL-1β plasma levels were increased in PSDCLP group. The effect of ATL-1 treatment (10 g/kg, i.v.) on hyperalgesia induced by PSD was determined through formalin test. The treatment reduced the number of pain related behaviors in PSD animals and controls on inflammatory phase. Our results show that ATL-1 was able to revert the hyperalgesia induced by PSD possibly through its anti-inflammatory action. Furthermore, PSD for 96 h increased cytokine plasma levels and reduced survival, however it was not able to modify leukocyte recruitment when challenged by an inflammatory or infectious stimulus. However the inflammatory mediators increase observed in PSD animals could be related to hyperalgesia since treatment with ATL-1 reverted this effect, possibly through anti-inflammatory mechanisms.

Made available in DSpace on 2014-07-29T15:30:37Z (GMT). No. of bitstreams: 1 dissertacao_hidelbertoM.pdf: 803099 bytes, checksum: 6b3724faf96c2ed68508b711948c6a27 (MD5) Previous issue date: 2011-07-21 / Neurocysticercosis (NCC) is caused by Taenia solium cysticercus and is the main cerebral infection caused by a parasite which can produce seizures and hydrocephaly. Besides the considerable economic losses it is also responsible for over than 50 thousand annual deaths. The use of experimental models has become a good method to the study of human cysticercosis in several organs including the central nervous system. The most used parasite is T. crassiceps cysticerci due to its rapid cycle of development and the antigenic similarity to T. solium and its intraperitoneal model is the most diffused. The inoculation of Mesocestoides corti cysticerci is the most common model of NCC used. Objectives: present a new experimental model to NCC studies by using T. crassiceps cysticerci inoculated into four different mice lineages (BALB/c, BALB/c KO-IL-4, C57BL/6 e C57BL/6 KO-IFNγ) and evaluate the inflammatory response of these animals to the infection during 7, 30, 60 and 90 days after the infection (DAI). This new model showed itself useful into reproducing the disease in animals resulting into lesions and alterations similar to the ones found in humans and without significant damages in the control groups, which enables the study. We observed that BALB/c mice are the less resistant to the infection, presenting greater ventriculomegaly, inflammatory response with the prevalence of acute phase cells and late destruction of the cysticerci when compared to C57BL/6 mice. The latter showed inflammation with prevalence of mononuclear cells and greater efficiency in the destruction of the parasites. When comparing the inflammatory response of conventional BALB/c mice to KO-IL-4 ones it was possible to observe that the absence of IL-4 induces greater parasitism which favors the cysticerci survival, less intensity of the inflammatory response, ventriculomegaly and perivasculitis. Both lineages destroyed the cysticerci at the end of the late phase of the inflammation process. When comparing the inflammatory response of conventional C57BL/6 mice to KO-IFNγ ones, the presence of IFNγ induced greater ventriculomegaly, chronification of the inflammation, microgliosis and precocious destruction of the cysticerci. In the absence of IFNγ the inflammatory response showed less intensity and late destruction of the cysticerci. / A Neurocisticercose (NCC) causada pelo cisticerco da Taenia solium é a principal infecção parasitária cerebral que pode resultar em sintomas clínicos como crises convulsivas e hidrocefalia. Além de consideráveis perdas econômicas, sendo responsável por mais de 50 mil mortes anuais. Os modelos experimentais tornaram-se ótimas ferramentas para o estudo da cisticercose humana em diversos órgãos, incluindo sua foma mais grave a NCC. O parasito mais utilizado nos modelos experimentais da cisticercose é a Taenia crassiceps, devido ao seu rápido ciclo de desenvolvimento e similaridade antigênica com T. solium, sendo o modelo intraperitoneal o mais difundido. A inoculação intracranial de cisticercos de Mesocestoides corti, é o modelo experimental mais utilizado no estudo da NCC. Os objetivos deste trabalho foram apresentar um novo modelo experimental para o estudo da NCC utilizando cisticercos de T. crassiceps, inoculados intracranialmente em quatro linhagens diferentes de camundongos (BALB/c, BALB/c KO-IL-4, C57BL/6 e C57BL/6 KO-IFNγ), bem como avaliar a resposta inflamatória desses animais frente à neuroinfecção, aos 7, 30, 60 e 90 dias após a infecção (DAI). Os animais foram pesados, anestesiados com solução de xilazina/cetamina, realizada a antissepsia e tricotomia local e em seguida realizado o orifício de trepanação para a inoculação dos cisticercos. O orifício era então fechado com massa plástica estéril, suturada a incisão e os animais eutanasiados de acordo com o dia experimental com dose subletal de anestésico para a retirada do encéfalo. O novo modelo apresentado mostrou-se útil em reproduzir a doença nos animais, resultando em lesões e alterações semelhantes às que ocorrem nos seres humanos, baixa perda de animais e sem danos significantes nos controles, facilitando o estudo da doença. Observou-se que os animais BALB/c são menos resistentes à infecção, apresentando uma tendência maior à ventriculomegalia, resposta inflamatória com predomínio de células da fase aguda e destruição tardia dos cisticercos, em relação aos animais C57BL/6. Estes, mostraram uma inflamação com uma tendência ao predomínio de células mononucleares e maior eficiência na destruição dos parasitos. Comparando a resposta inflamatória nos animais BALB/c convencionais e KO-IL-4, ficou evidenciado que a ausência da IL-4 induziu maior parasitismo, favorecendo a sobrevida dos cisticercos, menor intensidade na resposta inflamatória, ventriculomegalia e perivasculite. Ambas as linhagens destruiram os cisticercos no final da fase tardia da inflamação. Na comparação da resposta inflamatória dos camundongos C57BL/6 convencionais e KO-IFNγ, a presença do IFNγ induziu maior ventriculomegalia, cronificação da inflamação, microgliose e destruição precoce dos cisticercos. Na ausência do IFNγ, a resposta inflamatória apresentou menor intensidade e destruição tardia dos cisticercos.