Thy-1 is a glycosylphosphatidylinositol-anchored protein that is expressed on murine T lymphocytes and is involved in T cell-mediated immune responses. In the presence of costimulatory signals, monoclonal antibody (mAb)-induced signaling through Thy-1 is associated with hallmarks of T cell activation, including IL-2 production and T cell proliferation. Thy-1-induced signaling promotes cytotoxic effector molecule expression, but is unable to trigger delivery of the lethal hit to target cells, suggesting that Thy-1 provides an incomplete T cell receptor (TcR)-like signal. However, the effect of Thy-1 signaling on cytokine production and the development of T helper (Th) cell phenotypes (Th1, Th2, Th17) remains unclear. The purpose of this work was to further our understanding of Thy-1-mediated signal transduction and the role that Thy-1 plays in the development of effector T cell responses. I found that, in the context of costimulatory signals, anti-Thy-1 mAb induced significantly less IL-2 production, CD25 expression and T cell proliferation than anti-TcR? mAb. Several key signaling molecules, including protein tyrosine kinases, zeta chain-associated protein-70 and extracellular signal-regulated kinase were activated with delayed kinetics during Thy-1-mediated T cell activation. The delayed signaling kinetics resulted in the delayed acquisition of cytotoxic effector function and also delayed delivery of the lethal hit to target cells. Interestingly, Thy-1-mediated signaling induced significantly more IL-17 and IL-4 synthesis and less IFN-? synthesis in comparison to TcR-mediated signaling. Moreover, Thy-1-activated CD4+ T cells produced high levels of IL-17 and IL-4 but minimal IFN? when restimulated with anti-Thy-1 mAb or anti-TcR? mAb with or without costimulatory signals. The unique ability of Thy-1 signaling to induce IL-17 production correlated with the expression of the Th17 lineage-specific transcription factor, retinoic orphan receptor gamma t. These observations show that Thy-1 signaling differs from TcR signaling in its ability to induce Th cell cytokines. Taken together, my findings show that Thy-1 signaling can provide the full TcR-like signal required for both the differentiation and triggering of Th cells and cytotoxic T lymphocytes, albeit with delayed kinetics in comparison to TcR signaling. They also suggest that Thy-1 signaling may be important in the development of Th2 and Th17 responses.

Thymoquinone (TQ), a volatile oil component of black seed oil (derived from Nigella sativa), has been shown to have various biological effects including disease treatment and prevention. TQ is believed to share similar properties to the benzoquinones already in use as therapeutic drugs. Based on previous reports on the anti-inflammatory properties of black seed oil and TQ, it was originally hypothesized that TQ would inhibit lipopolysaccharide (LPS)-induced cellular sialidase activity in an anti-inflammatory manner. Sialidase activity was tested on live mouse bone marrow derived primary macrophage cells, BMC-2 macrophage cells, human embryonic kidney epithelial (HEK293) cells and human fibroblast cells using an assay that measures the cleavage of the sialidase specific fluorescent substrate 2’-(4-methylumbelliferyl)-α-DN-acetylneuraminic acid (4-MUNANA). The cleavage of 4-MUNANA causes the release of free 4-methylumbelliferone, which fluoresces at 450nm (blue) after excitation at 365nm. Unexpectedly, TQ induced sialidase activation in all three cell lines and wild type primary macrophage cells. TQ was unable to induce sialidase activity in primary macrophage cells isolated from Neu4 knockout mice suggesting that the TQ activates Neu4 sialidase enzyme. TQ-induced sialidase activity in these live cells was found to occur through intermediate GPCR-associated guanine nucleotide Gαi subunit and matrix metalloproteinase 9 (MMP9) by using specific inhibitors. In addition, TQ was found to induce sialidase activity in Toll-like receptor-deficient HEK293 cells. These latter data suggested that TQ may be activating GPCR Gαi and MMP9 signaling associated with Neu4 sialidase independent of TLRs. It is proposed that TQ-induced sialidase activity may activate Toll-like receptors in macrophage cells and the subsequent production of pro-inflammatory cytokines in the absence of LPS. Immunocytochemical staining of BMC-2 cells shows that TQ induced NFκB activation. NFκB activation was confirmed with electrophoretic mobility shift assay (EMSA) and western immunoblotting techniques. Cytokine arrays were used to test the pro-inflammatory cytokine response induced in mice by 5 hour treatment of TQ, compared to LPS. Mice treated with TQ exhibited an increase in IL-1β, IL-6 and TNF-α production, similar to LPS treatment. Taken together, the findings in these studies suggest that TQ is a novel ligand for Neu4 sialidase activation which consequently induces pro-inflammatory cytokine responses. / Thesis (Master, Microbiology & Immunology) -- Queen's University, 2009-04-21 17:38:10.413

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal

Improved perinatal care during the last few years has led to higher survival rates for preterm infants. However, with higher survival rates, the number of children demonstrating long-term health disorders that result in a poorer quality of life is increasing. The most common complications in those preterm children include motion disorders, vision and hearing impairment, mental disorders, and chronic lung disease. Intrauterine infection may cause foetal infection and inflammation thus inducing the inflammatory response in foetus, defined by foetal inflammatory response syndrome (FIRS). FIRS may cause a heavy damage in foetus and newborns as well as later disorders in the infant organism, such as cerebral palsy and chronic lung disease. Speaking about researches proving relation of the perinatal inflammatory response and psychomotor development in a preterm newborn, these are few. The foetal inflammatory response syndrome is a problem that has not been examined yet in Lithuania. Researches of cytokines in umbilical cord blood to make prognoses on the health and psychomotor development in a premature infant has not been performed either. Hopefully, the study results will allow a more detail explanation of the reasons for preterm delivery, better understanding of health disorders in premature infants and prognosis of the process of a child development. The aim of the study was to evaluate the importance of FIRS for the early and later adaptation of premature infants and for... [to full text] / Dėl pagerėjusios perinatalinės priežiūros per pastaruosius metus neišnešiotų naujagimių išgyvenamumas labai pagerėjo. Tačiau, kai neišnešiotų naujagimių išgyvena vis daugiau, daugėja ir vaikų, kuriems augant, išryškėja ilgalaikiai sveikatos sutrikimai, pabloginantys jų gyvenimo kokybę. Nurodoma, kad dažniausia neišnešioto naujagimio gimimo priežastis yra intrauterinė infekcija, kuri progresuodama gali inicijuoti vaisiaus uždegiminio atsako sindromą, kurio metu vaisiuje suintensyvėja uždegiminių citokinų IL-1, IL-6, IL-8, TNF-α, augimo veiksnių gamyba. Šio sindromo pasekmė - sunkūs vaisiaus ir naujagimio pažeidimai bei vėlesni liekamieji kūdikio sveikatos sutrikimai, tokie kaip cerebrinis paralyžius ir lėtinė plaučių liga. Tyrimų, kurie rodytų perinatalinio uždegimo ir neišnešioto naujagimio psichomotorinės raidos sąsajas nėra daug. Ryšys tarp vaisiaus uždegimo ir neišnešiotų naujagimių retinopatijos atrastas neseniai, tad tyrimų šia kryptimi atlikta taip pat nedaug. Ryšys tarp perinatalinio uždegimo ir grėsmingų naujagimystės komplikacijų leidžia daryti prielaidą, kad neišnešioto naujagimio ankstyvas sveikatos vertinimas ir gyvenimo kokybės prognozė yra labai svarbūs, norint kuo anksčiau pradėti tikslinį gydymą bei ankstyvą vaiko raidos korekciją. Šio darbo tikslas buvo įvertinti vaisiaus uždegiminio atsako sindromo įtaką neišnešioto naujagimio sveikatai ir psichomotorinei raidai iki 1 metų koreguoto amžiaus. Tyrimo objektą sudarė virkštelės kraujo citokinų IL-6, bTNF-α... [toliau žr. visą tekstą]

Dėl pagerėjusios perinatalinės priežiūros per pastaruosius metus neišnešiotų naujagimių išgyvenamumas labai pagerėjo. Tačiau, kai neišnešiotų naujagimių išgyvena vis daugiau, daugėja ir vaikų, kuriems augant, išryškėja ilgalaikiai sveikatos sutrikimai, pabloginantys jų gyvenimo kokybę. Nurodoma, kad dažniausia neišnešioto naujagimio gimimo priežastis yra intrauterinė infekcija, kuri progresuodama gali inicijuoti vaisiaus uždegiminio atsako sindromą, kurio metu vaisiuje suintensyvėja uždegiminių citokinų IL-1, IL-6, IL-8, TNF-α, augimo veiksnių gamyba. Šio sindromo pasekmė - sunkūs vaisiaus ir naujagimio pažeidimai bei vėlesni liekamieji kūdikio sveikatos sutrikimai, tokie kaip cerebrinis paralyžius ir lėtinė plaučių liga. Tyrimų, kurie rodytų perinatalinio uždegimo ir neišnešioto naujagimio psichomotorinės raidos sąsajas nėra daug. Ryšys tarp vaisiaus uždegimo ir neišnešiotų naujagimių retinopatijos atrastas neseniai, tad tyrimų šia kryptimi atlikta taip pat nedaug. Ryšys tarp perinatalinio uždegimo ir grėsmingų naujagimystės komplikacijų leidžia daryti prielaidą, kad neišnešioto naujagimio ankstyvas sveikatos vertinimas ir gyvenimo kokybės prognozė yra labai svarbūs, norint kuo anksčiau pradėti tikslinį gydymą bei ankstyvą vaiko raidos korekciją. Šio darbo tikslas buvo įvertinti vaisiaus uždegiminio atsako sindromo įtaką neišnešioto naujagimio sveikatai ir psichomotorinei raidai iki 1 metų koreguoto amžiaus. Tyrimo objektą sudarė virkštelės kraujo citokinų IL-6, bTNF-α... [toliau žr. visą tekstą] / Improved perinatal care during the last few years has led to higher survival rates for preterm infants. However, with higher survival rates, the number of children demonstrating long-term health disorders that result in a poorer quality of life is increasing. The most common complications in those preterm children include motion disorders, vision and hearing impairment, mental disorders, and chronic lung disease. Intrauterine infection may cause foetal infection and inflammation thus inducing the inflammatory response in foetus, defined by foetal inflammatory response syndrome (FIRS). FIRS may cause a heavy damage in foetus and newborns as well as later disorders in the infant organism, such as cerebral palsy and chronic lung disease. Speaking about researches proving relation of the perinatal inflammatory response and psychomotor development in a preterm newborn, these are few. The foetal inflammatory response syndrome is a problem that has not been examined yet in Lithuania. Researches of cytokines in umbilical cord blood to make prognoses on the health and psychomotor development in a premature infant has not been performed either. Hopefully, the study results will allow a more detail explanation of the reasons for preterm delivery, better understanding of health disorders in premature infants and prognosis of the process of a child development. The aim of the study was to evaluate the importance of FIRS for the early and later adaptation of premature infants and for... [to full text]

Background Type 2 diabetes is a common disease and the prevalence has increased in large parts of the world. In treatment of diabetes the type of diet is of great importance considering metabolic factors such as glucose level and blood lipids. Which diet that is most beneficial to avoid diabetic complications has been heavily debated in recent decades. This thesis is based on two clinical studies designed to compare the effects of different macronutrients. Methods A clinical trial was designed to compare a low-carbohydrate diet (LCD) to a low-fat diet (LFD) in treatment of patients with type 2 diabetes. Sixty-one patients at two health care centres were included and randomized to get advice to eat a LCD or a LFD. The LCD had an energy content where 50 energy percent (E%) where from fat, 20 E% from carbohydrates and 30 E% from protein. For the LFD the nutrient composition was similar to what is traditionally recommended for treatment of type 2 diabetes in Sweden. Metabolic factors, anthropometrics and questionnaires were analysed. To study postprandial effects a trial was designed to compare three different diets. Twentyone patients with type 2 diabetes were included to in randomized order test the three types of diets on separate test days. On each test day the patients were served breakfast and lunch and blood samples were taken at six times these days. Glucose, lipids and hormones were analysed. Results There were equal weight reduction in the two groups in the first trial during the two-year study period. At six month when compliance was good according to diet-records, the glucose level (HbA1c) was lowered and the HDL-cholesterol was increased in the LCD group. The inflammatory markers IL-6 and IL-1Ra were significantly lower in the LCD group than in the LFD group. At 12 months the physical function, bodily pain and general health  scores improved within the LCD group only. In the second trial the postprandial glucose and insulin levels were lower on the LCD compared to the LFD. However, the LCD resulted in a tendency to higher postprandial triglyceride levels. The Mediterranean type of diet with all energy intake at lunch resulted in a more pronounced insulin response and a glucose level at lunch similar to that of the low-fat diet. The increase-ratio of insulin correlated to the elevation of the incretin glucose-dependent insulinotropic peptide (GIP). Conclusions In the two-year study we found benefits for the LCD group regarding glucose control and insulin doses. Furthermore, only the LCD was found to improve the subclinical inflammatory state and there were some aspects of improved well-being in this group. Aiming for 20% of energy intake from carbohydrates is safe with respect to cardiovascular risk factors  compared with the traditional LFD and this approach could constitute a treatment alternative. In the postprandial state, the LCD induced lower insulin and glucose excursions than the LFD but at the same time a tendency of higher triglycerides. The long-term significance needs to be further examined. The accumulation of caloric intake from breakfast to lunch to a single large Mediterranean-style lunch-meal in type 2 diabetes might be advantageous from a metabolic perspective.

L'interferon-γ (IFNγ) est une cytokine immunomodulatrice puissante, également dotée d'une activité antivirale. Il possède deux ligands de haute affinité : un récepteur par lequel il transmet ses signaux et des polysaccharides complexes de la famille des héparanes sulfates (HS), tous deux situés à la surface cellulaire. In vivo, la liaison aux HS permet de concentrer localement la cytokine et de réguler son activité biologique par le biais d'une protection partielle du domaine C-terminal de la protéine. Ce domaine C-terminal, caractérisé par deux domaines basiques D1 et D2, est impliqué dans la reconnaissance du récepteur et des HS. Dans ce contexte, nos travaux se sont attachés à définir les aspects structuraux de l'interaction de l'IFNg, et plus précisément de son extrémité C-terminale, avec ses deux ligands. Pour cela, de divers mutants ponctuels, multiples et de délétion de l'IFNg ont été produites, purifiées et étudiées. Leur capacité à lier les HS et le récepteur de l'IFNg est déterminée par SPR puis leur influence sur l'activité antivirale de l'IFNg est déterminée. Les paramètres thermodynamiques de l'interaction IFNg:HS-oligosaccharides sont investigués. Par ailleurs, nous avons préparé une banque oligosaccharidique dérivée d'HS. Le criblage de cette banque, pour sa capacité à lier l'IFNγ, a permis de démontrer que l'IFNg reconnaissait des motifs de sulfatation particuliers. Finalement, nous avons tenté de cristallisé le complexe IFNg:HS-oligosaccharide, jusqu'à présent sans obtention de cristaux qui diffractent. Ces différentes approches visent à élucider le mécanisme de reconnaissance d'IFNg par des HS. Ceci afin de concevoir un mime de ce site d'interaction inhibant la signalisation de l'IFNg. Enfin, une compréhension plus détaillé de l'interaction de l'IFNg avec les HS et son récepteur reste à établir afin d'entièrement comprendre comment l'IFNg migre des HS vers l'IFNgR.

Liquid chromatography tandem mass spectrometry has become a powerful tool for investigating biological systems. Herein we describe the development of both isotope dilution mass spectrometry methods and targeted metabolomics methods for the study of metabolic and cell-cell signaling applications. A putative yeast enzyme was characterized by discovery metabolite profiling, kinetic flux profiling, transcriptomics and structural biology. These experiments demonstrated that the enzyme shb17 was a sedoheptulose bisphosphatase that provides a thermodynamically dedicated step towards riboneogenesis, leading to the redefinition of the canonical pentose phosphate pathway. An extension of metabolic profiling and kinetic flux profiling methods was developed for a set of symbiotic marine microorganisms. Carbon flux from the most abundant photosynthetic organism, Prochlorococcus, to a symbiotic Alteromonas was observed in liquid coculture. These methods enable a more biologically relevant assay for marine species and will lead to a better understanding of carbon flux in the oceans. Energy taxis refers to the active migration of bacteria in response to electron transport system related signals. The second messenger cyclic-di-GMP provides a link between the metabolic signals and motility. Quantitation of c-di-GMP helped characterize the nature of this regulation. Autoinducer-2 is a small sugar produced by a large variety of bacteria that is proposed to be a universal quorum sensing signal. The quorum sensing function of autoinducer-2 is disputed because it is produced by an enzyme of the activated methyl cycle, leading to an alternate hypothesis that it is simply a metabolic byproduct. Herein a method for the detection of autoinducer-2 is developed to enable studies of its signaling role and biosynthetic regulation. These studies demonstrated that autoinducer-2 does not function as a signal in all species. Further, metabolic experiments indicated that the metabolic impact of LuxS dysfunction was small and could be mitigated by recycling oxidized glutathione. Together, these data indicate that neither hypothesis is adequate. Evidence is provided that autoinducer-2 suppresses the immune system, by the interruption of cytokine signaling, implying that autoinducer play a protective role during host colonization.

Consequences of atherosclerosis may result in a number of diseases of the cardiovascular system that represent serious health problems and major causes of morbidity and mortality worldwide. Although it is initially considered as disease of fibro-lipid and thrombus deposition in the arterial wall, it also involves an ongoing inflammatory response. Normally, the inflammatory response is considered as a protective defence mechanism of the body. However, if the inflammation gets out of proportion to the threat it is dealing with, it may then result in a sustained chronic disorder and thus may underlie the initial stage of atherogenesis. The work of this thesis focuses on the expression of cytokines/chemokines and the vascular transcriptional response to inflammation, i.e. LPS in atherosclerosis. This has mainly been studied in animal models of atherosclerosis; consequently, we set out to investigate these events using human material in vitro (human carotid lesions). Employing quantitative analysis, we were able to detect a significant induction of protein and mRNA of the cytokines IL-1β, IL-6, IL-10 and TNF-α and the chemokines IL-8 and MCP-1 by LPS in both atherosclerotic and non-atherosclerotic vessels. In contrast, LPS induction of TNF-α, IL-1β and IL-10 was solely observed in the lesions, but not in normal arteries. In addition, the impact of IL-1 gene polymorphism on the risk of myocardial infarction (MI) was estimated by DNA genotyping of 387 survivors of a first MI and 387 sex and age-matched control subjects. We found no statistically significant differences in either genotypic distribution or allelic frequencies of IL-1β (-511) or IL-1Ra (VNTR) polymorphisms between first-time survivors of myocardial infarction and their age-matched healthy controls. Incontrast, our results demonstrated a strong association between the IL-1Ra genotype and severity of angiographically determined coronary artery disease in post-MI patients. To further investigate the vascular response to inflammation, we used gene array analysis to evaluate the human vascular transcriptional response to LPS of non-atherosclerotic human renal arteries compared to carotid lesions. In LPS treated renal arteries, 54% of the transcripts gave a detectable signal, where 4% were upregulated and 3.8% down-regulated. In the LPS stimulated carotid lesions, 44% of transcripts were detected. In this latter group, 5.1% of transcripts were increased and 3.3% decreased. Interestingly, a newly identified virus-inducible antiviral protein, CMV inducible gene 5/viperin (Cig5), was among the most strongly induced gene in both normal and atherosclerotic biopsies. Single gene analysis revealed viperin in the endothelium of human atherosclerotic lesions. Further, viperin was induced in vascular cells by inflammatory stimuli and CMV infection. In conclusion we show that atherosclerotic vessels produce more proinflammatory cytokines/chemokines than normal vessels. Interestingly, our results indicate that LPS enhances the expression of cytokines/chemokines in a similar pattern both in lesions and normal arteries. However, the response is stronger in atherosclerotic lesions. Furthermore, our results suggest that genetic polymorphisms within the IL-1Ra loci may influence the severity of CAD. Finally, the CMV inducible gene 5/viperin have been identified as a putative culprit molecule in vascular inflammation and atherosclerosis.

Submitted by Ana Maria Fiscina Sampaio (fiscina@bahia.fiocruz.br) on 2012-06-05T21:05:38Z No. of bitstreams: 1 Bruno de Bezerril Andrade - 2010.pdf: 74164438 bytes, checksum: a1cccf3d1f924ff7710fc4a73a190f0a (MD5) / Made available in DSpace on 2012-06-05T21:05:38Z (GMT). No. of bitstreams: 1 Bruno de Bezerril Andrade - 2010.pdf: 74164438 bytes, checksum: a1cccf3d1f924ff7710fc4a73a190f0a (MD5) / Universidade Federal da Bahia. Centro de Pesquisas Gonçalo Moniz. Salvador, Bahia, Brasil / A malária é considerada uma das mais importantes doenças infecciosas do mxmdo. Esta doença é causada por diversas espécies do protozoário Plasmodium sp., principalmente o Plasmodium falciparum e o Plasmodium vivax, transmitido por mosquitos do gênero Anopheles. Apesar dos esforços governamentais e privados para o desenvolvimento de estratégias para o controle da doença, o panorama atual da malária está piorando, muito em razão do aparecimento de cepas de parasitas resistentes aos medicamentos. Os casos fatais são relatados principalmente na Áfiica e são causados pelo Plasmodium falciparum. Apesar de ser menos letal, a malária causada pelo Plasmodium vivax é mais amplamente distribuída e pode apresentar também alta morbidade e mortalidade. Na maioria das áreas endêmicas, estudos têm identificado vários fatores relacionados à imunidade clínica ou susceptibilidade aos parasitas. Assim, pelo menos quanto à malária causada pelo Plasmodium falciparum, idade, polimorfismos genéticos e exposição repetida ao parasita são considerados importantes determinantes da evolução da doença. Infelizmente, pouco tem sido feito na identificação de fatores preditores consistentes que poderiam ser usados para avaliação clínica. Este quadro é ainda pior para malária causada pelo Plasmodium vivax, provavelmente porque muitos pesquisadores consideram que é uma doença benigna. Além disso, como a maioria do conhecimento atual sobre a patogênese da malária não ajudou a reduzir a ocorrência da infecção e suas complicações, novas abordagens são necessárias para superar este cenário desfavorável. Esta Tese reúne um conjunto de seis manuscritos que visam identificar potenciais determinantes da gravidade da malária em uma área endêmica da Amazônia Ocidental Brasileira. Em primeiro lugar, um método preciso e eficaz para o diagnóstico da malária foi rastreado através da comparação de vários testes, incluindo um software baseado em redes neurais artificiais. O ensaio molecular mostrou-se o mais eficiente para o diagnóstico da malária sintomáticos e assintomáticos. Além disso, a utilização racional de um teste rápido para diagnóstico da malária pode ser promissora em áreas onde há dificuldade na formação continuada dos técnicos diagnósticos. A rede neural artificial indicou que o balanço de citocinas é um forte determinante do quadro clínico. Em outro estudo, uso de sorologia para mensuração de anticorpos IgG contra o sonicado de glândula salivar do vetor Anopheles darlingi mostrou-se útil para a avaliação da exposição ao Plasmodium vivax e também para estimar a imunidade clínica á malária. Em um terceiro estudo com foco na identificação de outros fatores relacionados à imunidade clínica, a exposição natural ao vírus da hepatite B mostrou-se associada à redução da gravidade clínica da malária causada tanto pelo Plasmodium vivax quanto pelo Plasmodium falciparum. No que diz respeito exclusivamente à malária vivax, os casos graves apresentaram uma intensa e desregulada resposta inflamatoria sistêmica. Nestes pacientes, a enzima antioxidante superóxido dismutase-1 surgiu como um excelente marcador da gravidade e mostrou-se envolvida na patogênese da doença grave, na qual há uma liberação de grandes quantidades de heme livre. Em conjunto, os manuscritos desta tese adicionam importantes informações no entendimento dos mecanismos determinantes da gravidade da malária, extremamente / Malaria is considered one of the most important infectious diseases that ever threaten the world. This disease is caused mainly by the infection with Plasmodium falciparum or Plasmodium vivax transmitted by Anopheles mosquitoes. Despite governmental and private efforts for the development of key strategies for the disease control, the actual panorama of the Plasmodium infection is getting worse due to the emergence of drug resistMt parasite strains. The lethal cases are reported mostly in Africa and are caused by Plasmodium falciparum. Albeit being less lethal, Plasmodium vivax infections are more widely distributed can cause high morbidity and eventually death. In most endemic areas, studies have indentified a number of factors related to clinical immunity or susceptibility to the parasites. Thus, at least regarding the falciparum malaria, age, genetic polymorphisms and repeated exposure to Plasmodium are considered most important determinants of the disease outcome. Unfortunately, little has been made in the screening of reliable predicting factors that could be ultimately used for clinical evaluations. This landscape is even worse for vivax malaria, probably because many researches consider it as a benign disease. Moreover, as most of the current knowledge about the malaria pathogenesis did not truly help to relieve the disease burden, new insights are necessary to overcome this unfavorable scenario. This thesis brings together a set of six manuscripts that aim to identify potential determinants of the disease severity linked to the immunopathogenesis in an endemic area from the western Brazilian Amazon. First, a precise and effective method for malaria diagnosis was screening by comparing multiple tests, including a software based of artificial neural networks. The molecular assay showed to be the most efficient for the diagnosis of symptomatic and asymptomatic malaria. In addition, the rational use of a rapid test for the diagnosis of malaria may be promising in areas where there is difficulty in continued training of technical human resources. The artificial neural network indicated that the cytokine balance is a strong determinant of the clinical presentation. In another study, the use of serology for measuring IgG antibodies against the sonicate salivary gland of Anopheles darlingi vector is a promising marker of exposure to Plasmodium vivax and can also estimate the clinical immunity. Intriguingly, the natural exposure to the hepatitis B virus appeared as an important factor associated with reduced clinical severity for both vivax and falciparum malaria. Concerning solely the vivax malaria, severe cases have an intense and unregulated inflammatory response. In these patients, the antioxidant enzyme superoxide dismutase-1 has emerged as an excellent marker of severity and was involved in the pathogenesis of the severe disease in which there is a release of large amounts of free heme. Together, the manuscripts of this thesis add important information in understanding the mechanisms that determine the severity of malaria.