Imunobiologia e imunogenética de desordens gestacionais e transtorno do espectro autista

Esta tese apresenta artigos de dados e trabalhos teóricos envolvendo imunologia da gestação, desordens gestacionais e imunogenética do Transtorno do Espectro Autista (TEA). Este trabalho está dividido em duas partes, a primeira sendo composta por quatro capítulos, e a segunda parte, por três capítulos totalizando sete artigos diferentes. Uma introdução geral precede as duas partes principais, onde são apresentados conceitos importantes abordados ao longo dos textos. A gestação humana é um processo complexo que envolve diferentes sistemas fisiológicos que sofrem influência do sistema imune materno. Complicações gestacionais e seus possíveis gatilhos ambientais são apresentados nesta tese, com destaque para desequilíbrios nos níveis de citocinas nessas condições. Além disso, é abordado o componente genético das respostas imunes aos diferentes desafios ambientais enfrentados durante a gravidez pela mãe e pelo feto em desenvolvimento. O TEA é uma condição que impacta o neurodesenvolvimento fetal, além de afetar as habilidades cognitivas e sociais dos indivíduos acometidos, geralmente manifestando-se antes dos três anos de idade. Sabe-se que o TEA possui um forte componente genético com altas taxas de herdabilidade e de concordância entre gêmeos monozigóticos. Além disso, o TEA possui um importante componente ambiental. Sabe-se, ainda, que distúrbios imunológicos são um componente do TEA. Nesta linha, são abordados polimorfismos em genes relacionados ao sistema imunológico no contexto do TEA. Um dos fatores ambientais fortemente sugeridos como risco para TEA são algumas complicações gestacionais, principalmente aquelas com fundo inflamatório. Assim, diferentes aspectos imunológicos e ambientais durante a gravidez podem ser fatores-chave para o desenvolvimento de distúrbios gestacionais e/ou para a incidência de problemas neurológicos na prole. Na “Parte I” desta tese são abordados aspectos imunológicos da gestação humana, juntamente com discussões sobre o papel das vesículas extracelulares no contexto de gravidez bem-sucedida e de complicações gestacionais e em diferentes doenças infecciosas. Ao longo da “Parte II”, são apresentados genes relacionados ao sistema imunológico, nos quais diferentes polimorfismos, especificamente variantes genéticas pró-inflamatórias, variantes genéticas do MHC e variantes genéticas imunometabólicas, já foram estudados no contexto do autismo e TEA. Diferentes gatilhos inflamatórios durante a gravidez que já foram indicados como fatores de risco para a manifestação de TEA em crianças nascidas dessas gestações são também aqui discutidos. Nesse contexto, destacam-se as consequências da ativação imune materna (MIA) e sua influência no feto em desenvolvimento. Além disso, é proposta uma conexão mecanicista entre os principais distúrbios relacionados à inflamação na gravidez e risco para autismo considerando o “universo das vesículas extracelulares”. Por fim, são apresentados resultados de estudos imunogenéticos envolvendo a deleção do gene NKG2C e variantes nos genes NKG2D e NKG2A em pacientes com TEA e em seus respectivos pais biológicos.This thesis presents data and theoretical studies involving gestational immunology, gestational disorders, and immunogenetics of Autism Spectrum Disorder (ASD). This work is divided into two parts, each consisting of four chapters, totaling eight different articles. A general introduction precedes these two main parts, where important concepts covered throughout the texts are presented. Human pregnancy is a complex process that involves different physiological systems, which are influenced by the maternal immune system. Gestational complications and their possible environmental triggers are also presented in this thesis, highlighting imbalances in cytokine levels under these conditions. In addition, the genetic component of immune responses to the different environmental challenges faced during pregnancy by the mother and the developing fetus is addressed. ASD is a condition that impacts both fetal neurodevelopment and the cognitive and social abilities of affected individuals, usually manifesting before the age of three. ASD is known to have a strong genetic component with high heritability and agreement rates between monozygotic twins. In addition, it has an important environmental component. Immune disorders are also known to be a component of ASD. In this line, polymorphisms in genes related to the immune system in the context of ASD are addressed. Strongly suggested environmental risk factors for ASD are gestational complications, especially those with an inflammatory background. Thus, different immunological and environmental aspects during pregnancy may be key factors for the development of gestational disorders and/or for the incidence of neurological problems in the offspring. In “Part I” of this thesis, immunological aspects of human pregnancy are discussed, along with discussions about the role of extracellular vesicles in the contexts of both successful and complicated pregnancies and in different infectious diseases. “Part II” presentes genes related to the immune system, in which different polymorphisms have already been studied in the context of autism and ASD, specifically proinflammatory genetic variants, MHC genetic variants, and immunometabolic genetic variants. Different inflammatory triggers during pregnancy that were already suggested as risk factors for ASD in children born of these pregnancies are also discussed. In this context, the consequences of maternal immune activation (MIA) and its influence on the developing fetus are highlighted. In addition, a mechanistic connection between major inflammation-related disorders in pregnancy and risk for autism considering the “extracellular vesicle universe” is proposed. Finally, results of immunogenetic studies involving deletion of the NKG2C gene and variants in the NKG2D and NKG2A genes in ASD patients and their respective biological parents are presented

Increased systemic IL-6 levels point to inflammation as a determinant of renal cell carcinoma development

Introduction: Renal cell carcinoma (RCC) is one of the most prevalent kidney tumors. Inflammation is believed to be a key factor in its progression and spread since inflammatory markers are generally associated with poor prognosis in RCC patients. Cytokines are cell communication molecules involved in both healthy and pathological processes, including tumor growth and progression. Recent findings suggest that cytokine level measurements could be used for cancer monitoring and prognosis. Methods: This study characterized and compared the levels of different cytokines associated with the classical Th1, Th2, and Th17 immune responses in plasma samples from RCC patients (n = 25) and healthy controls (n = 29). Cytokine levels (IL-2, IL-4, IL-6, IL-10, TNF-α, IFN-γ, and IL-17A) were evaluated by flow cytometry using a BD Cytometric Bead Array (CBA) kit. Results: No statistical differences in systemic IL-2, IL-4, IL-10, IL-17A, TNF, and INF-γ levels were observed between RCC patients and controls (p > 0.05). However, higher systemic IL-6 levels were observed in RCC patients (p = 0.0034). Conclusions: This study highlights the importance of assessing the impact of IL-6 on RCC pathogenesis and its potential role as a biomarker of disease progression

Control and prevention of infectious diseases from a One Health perspective

The ongoing COVID-19 pandemic has caught the attention of the global community and rekindled the debate about our ability to prevent and manage outbreaks, epidemics, and pandemics. Many alternatives are suggested to address these urgent issues. Some of them are quite interesting, but with little practical application in the short or medium term. To realistically control infectious diseases, human, animal, and environmental factors need to be considered together, based on the One Health perspective. In this article, we highlight the most effective initiatives for the control and prevention of infectious diseases: vaccination; environmental sanitation; vector control; social programs that encourage a reduction in the population growth; control of urbanization; safe sex stimulation; testing; treatment of sexually and vertically transmitted infections; promotion of personal hygiene practices; food safety and proper nutrition; reduction of the human contact with wildlife and livestock; reduction of social inequalities; infectious disease surveillance; and biodiversity preservation. Subsequently, this article highlights the impacts of human genetics on susceptibility to infections and disease progression, using the SARS-CoV-2 infection as a study model. Finally, actions focused on mitigation of outbreaks and epidemics and the importance of conservation of ecosystems and translational ecology as public health strategies are also discussed

Exploring potential impacts of pregnancy-related maternal immune activation and extracellular vesicles on immune alterations observed in autism spectrum disorder

Autism Spectrum Disorder (ASD) is a set of neurodevelopmental disorders usually observed in early life, with impacts on behavioral and social skills. Incidence of ASD has been dramatically increasing worldwide, possibly due to increase in awareness/diagnosis as well as to genetic and environmental triggers. Currently, it is estimated that ~1% of the world population presents ASD symptoms. In addition to its genetic background, environmental and immune-related factors also influence the ASD etiology. In this context, maternal immune activation (MIA) has recently been suggested as a component potentially involved in ASD development. In addition, extracellular vesicles (EVs) are abundant at the maternal-fetal interface and are actively involved in the immunoregulation required for a healthy pregnancy. Considering that alterations in concentration and content of EVs have also been associated with ASD, this article raises a debate about the potential roles of EVs in the processes surrounding MIA. This represents the major differential of the present review compared to other ASD studies. To support the suggested correlations and hypotheses, findings regarding the roles of EVs during pregnancy and potential influences on ASD are discussed, along with a review and update concerning the participation of infections, cytokine unbalances, overweight and obesity, maternal anti-fetal brain antibodies, maternal fever, gestational diabetes, preeclampsia, labor type and microbiota unbalances in MIA and ASD

Beyond diversity loss and climate change : impacts of Amazon deforestation on infectious diseases and public health

Amazonian biodiversity is increasingly threatened due to the weakening of policies for combating deforestation, especially in Brazil. Loss of animal and plant species, many not yet known to science, is just one among many negative consequences of Amazon deforestation. Deforestation affects indigenous communities, riverside as well as urban populations, and even planetary health. Amazonia has a prominent role in regulating the Earth’s climate, with forest loss contributing to rising regional and global temperatures and intensification of extreme weather events. These climatic conditions are important drivers of emerging infectious diseases, and activities associated with deforestation contribute to the spread of disease vectors. This review presents the main impacts of Amazon deforestation on infectious-disease dynamics and public health from a One Health perspective. Because Brazil holds the largest area of Amazon rainforest, emphasis is given to the Brazilian scenario. Finally, potential solutions to mitigate deforestation and emerging infectious diseases are presented from the perspectives of researchers in different fields

Synthesizing the connections between environmental disturbances and zoonotic spillover

Zoonotic spillover is a phenomenon characterized by the transfer of pathogens between different animal species. Most human emerging infectious diseases originate from non-human animals, and human-related environmental disturbances are the driving forces of the emergence of new human pathogens. Synthesizing the sequence of basic events involved in the emergence of new human pathogens is important for guiding the understanding, identifi cation, and description of key aspects of human activities that can be changed to prevent new outbreaks, epidemics, and pandemics. This review synthesizes the connections between environmental disturbances and increased risk of spillover events based on the One Health perspective. Anthropogenic disturbances in the environment (e.g., deforestation, habitat fragmentation, biodiversity loss, wildlife exploitation) lead to changes in ecological niches, reduction of the dilution effect, increased contact between humans and other animals, changes in the incidence and load of pathogens in animal populations, and alterations in the abiotic factors of landscapes. These phenomena can increase the risk of spillover events and, potentially, facilitate new infectious disease outbreaks. Using Brazil as a study model, this review brings a discussion concerning anthropogenic activities in the Amazon region and their potential impacts on spillover risk and spread of emerging diseases in this region

Transcriptional profiling of a fungal granuloma reveals a low metabolic activity of Paracoccidioides brasiliensis yeasts and an actively regulated host immune response

Granulomas are important immunological structures in the host defense against the fungus Paracoccidioides brasiliensis, the main etiologic agent of Paracoccidioidomycosis (PCM), a granulomatous systemic mycosis endemic in Latin America. We have performed transcriptional and proteomic studies of yeasts present in the pulmonary granulomas of PCM aiming to identify relevant genes and proteins that act under stressing conditions. C57BL/6 mice were infected with 1x106 yeasts and after 8- and 12-weeks of infection, granulomatous lesions were obtained for extraction of fungal and murine RNAs and fungal proteins. Dual transcriptional profiling was done comparing lung cells and P. brasiliensis yeasts from granulomas with uninfected lung cells and the original yeast suspension used in the infection, respectively. Mouse transcripts indicated a lung malfunction, with low expression of genes related to muscle contraction and organization. In addition, an increased expression of transcripts related to the activity of neutrophils, eosinophils, macrophages, lymphocytes as well as an elevated expression of IL-1β, TNF-α, IFN-γ, IL-17 transcripts were observed. The increased expression of transcripts for CTLA-4, PD-1 and arginase-1, provided evidence of immune regulatory mechanisms within the granulomatous lesions. Also, our results indicate iron as a key element for the granuloma to function, where a high number of transcripts related to fungal siderophores for iron uptake was observed, a mechanism of fungal virulence not previously described in granulomas. Furthermore, transcriptomics and proteomics analyzes indicated a low fungal activity within the granuloma, as demonstrated by the decreased expression of genes and proteins related to energy metabolism and cell cycle

Blocking the CTLA-4 and PD-1 pathways during pulmonary paracoccidioidomycosis improves immunity, reduces disease severity, and increases the survival of infected mice

Immune checkpoint pathways, i.e., coinhibitory pathways expressed as feedback following immune activation, are crucial for controlling an excessive immune response. Cytotoxic T lymphocyte antigen-4 (CTLA-4) and programmed cell death protein-1 (PD-1) are the central classical checkpoint inhibitory (CPI) molecules used for the control of neoplasms and some infectious diseases, including some fungal infections. As the immunosuppression of severe paracoccidioidomycosis (PCM), a chronic granulomatous fungal disease, was shown to be associated with the expression of coinhibitory molecules, we hypothesized that the inhibition of CTLA-4 and PD-1 could have a beneficial effect on pulmonary PCM. To this end, C57BL/6 mice were infected with Paracoccidioides brasiliensis yeasts and treated with monoclonal antibodies (mAbs) α-CTLA-4, α-PD-1, control IgG, or PBS. We verified that blockade of CTLA-4 and PD-1 reduced the fungal load in the lungs and fungal dissemination to the liver and spleen and decreased the size of pulmonary lesions, resulting in increased survival of mice. Compared with PBS-treated infected mice, significantly increased levels of many pro- and anti-inflammatory cytokines were observed in the lungs of α-CTLA-4-treated mice, but a drastic reduction in the liver was observed following PD-1 blockade. In the lungs of α-CPI and IgG-treated mice, there were no changes in the frequency of inflammatory leukocytes, but a significant reduction in the total number of these cells was observed. Compared with PBS-treated controls, α-CPI- and IgG-treated mice exhibited reduced pulmonary infiltration of several myeloid cell subpopulations and decreased expression of costimulatory molecules. In addition, a decreased number of CD4+ and CD8+ T cells but sustained numbers of Th1, Th2, and Th17 T cells were detected. An expressive reduction in several Treg subpopulations and their maturation and suppressive molecules, in addition to reduced numbers of Treg, TCD4+, and TCD8+ cells expressing costimulatory and coinhibitory molecules of immunity, were also detected. The novel cellular and humoral profiles established in the lungs of α-CTLA-4 and α-PD-1-treated mice but not in control IgG-treated mice were more efficient at controlling fungal growth and dissemination without causing increased tissue pathology due to excessive inflammation. This is the first study demonstrating the efficacy of CPI blockade in the treatment of pulmonary PCM, and further studies combining the use of immunotherapy with antifungal drugs are encouraged

Imunobiologia e imunogenética de desordens gestacionais e transtorno do espectro autista

Esta tese apresenta artigos de dados e trabalhos teóricos envolvendo imunologia da gestação, desordens gestacionais e imunogenética do Transtorno do Espectro Autista (TEA). Este trabalho está dividido em duas partes, a primeira sendo composta por quatro capítulos, e a segunda parte, por três capítulos totalizando sete artigos diferentes. Uma introdução geral precede as duas partes principais, onde são apresentados conceitos importantes abordados ao longo dos textos. A gestação humana é um processo complexo que envolve diferentes sistemas fisiológicos que sofrem influência do sistema imune materno. Complicações gestacionais e seus possíveis gatilhos ambientais são apresentados nesta tese, com destaque para desequilíbrios nos níveis de citocinas nessas condições. Além disso, é abordado o componente genético das respostas imunes aos diferentes desafios ambientais enfrentados durante a gravidez pela mãe e pelo feto em desenvolvimento. O TEA é uma condição que impacta o neurodesenvolvimento fetal, além de afetar as habilidades cognitivas e sociais dos indivíduos acometidos, geralmente manifestando-se antes dos três anos de idade. Sabe-se que o TEA possui um forte componente genético com altas taxas de herdabilidade e de concordância entre gêmeos monozigóticos. Além disso, o TEA possui um importante componente ambiental. Sabe-se, ainda, que distúrbios imunológicos são um componente do TEA. Nesta linha, são abordados polimorfismos em genes relacionados ao sistema imunológico no contexto do TEA. Um dos fatores ambientais fortemente sugeridos como risco para TEA são algumas complicações gestacionais, principalmente aquelas com fundo inflamatório. Assim, diferentes aspectos imunológicos e ambientais durante a gravidez podem ser fatores-chave para o desenvolvimento de distúrbios gestacionais e/ou para a incidência de problemas neurológicos na prole. Na “Parte I” desta tese são abordados aspectos imunológicos da gestação humana, juntamente com discussões sobre o papel das vesículas extracelulares no contexto de gravidez bem-sucedida e de complicações gestacionais e em diferentes doenças infecciosas. Ao longo da “Parte II”, são apresentados genes relacionados ao sistema imunológico, nos quais diferentes polimorfismos, especificamente variantes genéticas pró-inflamatórias, variantes genéticas do MHC e variantes genéticas imunometabólicas, já foram estudados no contexto do autismo e TEA. Diferentes gatilhos inflamatórios durante a gravidez que já foram indicados como fatores de risco para a manifestação de TEA em crianças nascidas dessas gestações são também aqui discutidos. Nesse contexto, destacam-se as consequências da ativação imune materna (MIA) e sua influência no feto em desenvolvimento. Além disso, é proposta uma conexão mecanicista entre os principais distúrbios relacionados à inflamação na gravidez e risco para autismo considerando o “universo das vesículas extracelulares”. Por fim, são apresentados resultados de estudos imunogenéticos envolvendo a deleção do gene NKG2C e variantes nos genes NKG2D e NKG2A em pacientes com TEA e em seus respectivos pais biológicos.This thesis presents data and theoretical studies involving gestational immunology, gestational disorders, and immunogenetics of Autism Spectrum Disorder (ASD). This work is divided into two parts, each consisting of four chapters, totaling eight different articles. A general introduction precedes these two main parts, where important concepts covered throughout the texts are presented. Human pregnancy is a complex process that involves different physiological systems, which are influenced by the maternal immune system. Gestational complications and their possible environmental triggers are also presented in this thesis, highlighting imbalances in cytokine levels under these conditions. In addition, the genetic component of immune responses to the different environmental challenges faced during pregnancy by the mother and the developing fetus is addressed. ASD is a condition that impacts both fetal neurodevelopment and the cognitive and social abilities of affected individuals, usually manifesting before the age of three. ASD is known to have a strong genetic component with high heritability and agreement rates between monozygotic twins. In addition, it has an important environmental component. Immune disorders are also known to be a component of ASD. In this line, polymorphisms in genes related to the immune system in the context of ASD are addressed. Strongly suggested environmental risk factors for ASD are gestational complications, especially those with an inflammatory background. Thus, different immunological and environmental aspects during pregnancy may be key factors for the development of gestational disorders and/or for the incidence of neurological problems in the offspring. In “Part I” of this thesis, immunological aspects of human pregnancy are discussed, along with discussions about the role of extracellular vesicles in the contexts of both successful and complicated pregnancies and in different infectious diseases. “Part II” presentes genes related to the immune system, in which different polymorphisms have already been studied in the context of autism and ASD, specifically proinflammatory genetic variants, MHC genetic variants, and immunometabolic genetic variants. Different inflammatory triggers during pregnancy that were already suggested as risk factors for ASD in children born of these pregnancies are also discussed. In this context, the consequences of maternal immune activation (MIA) and its influence on the developing fetus are highlighted. In addition, a mechanistic connection between major inflammation-related disorders in pregnancy and risk for autism considering the “extracellular vesicle universe” is proposed. Finally, results of immunogenetic studies involving deletion of the NKG2C gene and variants in the NKG2D and NKG2A genes in ASD patients and their respective biological parents are presented