A sepse é a Síndrome da Resposta Inflamatória Sistêmica decorrente de uma infecção por gram positivos/negativos, fungos ou vírus. É caracterizada por alta liberação de mediadores inflamatórios podendo levar à morte. As células B-1 são encontradas em cavidades peritoneal e pleural de camundongos e sua origem e função ainda não são completamente conhecidas. Apresentam marcadores de superfície de linhagem mielóide e linfóide e migram para focos inflamatórios comportando-se como macrófagos. Objetivo: investigar o papel da célula B-1 na resposta inflamatória após estímulo com lipopolissacáride (LPS) in vitro e in vivo. Métodos: TNF-, IL-6, IL-10 (ELISA) e nitrito (Griess) foram dosados em sobrenadante de cultura celular (106 cel./ml). As células em cultura receberam por 24h de estímulo com 10 g/mL de LPS de Escherichia coli (026:B6 Sigma®). Foram realizados os seguintes grupos cultura de célula B-1 (Balb/c), cultura de macrófagos de linhagem (RAW 264.7) coculturas (macrófagos de linhagem RAW 264.7 e células B-1 (Balb/c, C57BL/6 e C57BL/6 IL-10 -/-), e células peritoneais de camundongos Balb/c e Balb/Xid (imunodeficiente em célula B-1) A endotoxemia foi induzida com injeção de LPS 15 mg/kg (i.p.) em camundongos Balb/c e Balb/Xid. Foram quantificados, TNF-, IL-6, IL-10 e nitrito em soro, pulmão e intestino dos animais após 1,5, 4 e 6 horas após a injeção de LPS. Ensaios de inoculação de células B-1 (Balb/c) em camundongos Balb/Xid foram realizados, e curva de sobrevida foi analisada após indução de endotoxemia. Resultados: Após o estímulo com LPS, células B-1 produziram IL-10 e a presença destas células em cocultura com macrófago promoveu a diminuição na produção de TNF-, IL-6, Nitrito e aumento de IL-10. Contudo, célula B-1 (IL-10 -/-) em cocultura com macrófagos, não inibem a produção de mediadores pro inflamatórios. Análise com macrófagos peritoneais de camundongo Balb/Xid e Balb/c após estímulo com LPS em cultura mostrou reprodução do fenômeno encontrado com os experimentos com cultura de célula imortalizada, isto é, maior produção de TNF-, IL-6 e NO em Balb/Xid (B-1 deficiente). Os estudos in vivo mostraram 60% de mortalidade em camundongo Balb/Xid comparando com Balb/c (0%) após 16 horas de injeção de LPS. Nos animais Balb/Xid encontramos padrão pro inflamatório exacerbado com maiores concentrações de TNF-, IL-6 e menores concentrações de IL-10 no plasma e tecidos quando comparamos com Balb/c. Conclusões: Nossos dados mostraram que a presença de células B-1 promoveram diminuição de mediadores pro inflamatórios e aumento de IL-10 em coculturas com macrófagos e que a modulação da resposta inflamatória pode ser devida a secreção de IL-10 pela célula B-1. Este padrão de resposta pro inflamatória se repete in vivo e é a possível causadora da maior taxa de mortalidade em camundongos da linhagem Balb/Xid. / Sepsis syndrome is caused by inappropriate immune activation due to bacteria and bacterial components released during infection. This syndrome is the leading cause of death in intensive care units. Specialized B-lymphocytes located in the peritoneal and pleural cavities are known as B-1 cells. These cells produce IgM and IL-10, both of which are potent regulators of cell-mediated immunity. It has been suggested that B-1 cells modulate the systemic inflammatory response in sepsis. In this study, we conducted in vitro and in vivo experiments in order to investigate a putative role of B-1 cells in a murine model of LPS-induced sepsis. Macrophages and B-1 cells were studied in monocultures and in co-cultures. The B-1 cells produced the anti-inflammatory cytokine IL-10 in response to LPS. In the B-1 cell-macrophage co-cultures, production of proinflammatory mediators (TNF-, IL-6 and nitrite) was lower than in the macrophage monocultures, whereas that of IL-10 was higher in the co-cultures. Co-culture of B-1 IL-10/ cells and macrophages did not reduce the production of the proinflammatory mediators (TNF-, IL-6 and nitrite). After LPS injection, the mortality rate was higher among Balb/Xid mice, which are B-1 cell deficient, than among wild-type mice (65.0% vs. 0.0%). The Balb/Xid mice also presented a proinflammatory profile of TNF-, IL-6 and nitrite, as well as lower levels of IL-10. In the early phase of LPS stimulation, B-1 cells modulate the macrophage inflammatory response, and the main molecular pathway of that modulation is based on IL-10-mediated intracellular signaling.

No description available.

There are currently an estimated 33.2 million people living with human immunodeficiency virus (HIV) worldwide. For these individuals, long-term use of combination antiretroviral therapy (cART) is not feasible for a variety of reasons including major adverse complications, multi-drug resistance, poor adherence, and high cost. Hence, development of novel therapeutic strategies that can reduce the life-long dependency on cART is highly desired. In order to develop effective therapeutic strategies such as a therapeutic vaccine, we need to have a greater understanding of the immune correlates of viral control in chronic HIV infection. In this thesis, we used treatment interruption (TI) as a tool to test the efficacy of several therapeutic approaches and immune parameters for their association with effective control of viral replication. / In Chapter 2 we showed that cART intensification and Remune vaccination resulted in reduced viral load (VL) plateau during sequential TIs. Although HIV-specific immune responses measured by interferon-gamma (IFN-gamma) enzyme-linked immunospot assay (ELISPOT) increased in the same time frame, neither their breadth nor magnitude correlated with the decrease in VL plateau. In Chapter 3 the effect of ALVAC-vCP1425 plus Remune vaccination on HIV proteome-wide HIV-specific responses was monitored using a dual color IFN-gamma/interleukin-2 (IL-2) ELISPOT assay. We observed an increase in the magnitude of HIV-specific IFN-gamma/IL-2 responses, as well as in the breadth of Gag-specific IFN-gamma responses in the vaccinated groups compared to placebo groups. A shift towards an increased contribution of Gag-specific responses to total HIV-specific vaccine induced immune response was associated with longer delay to viral rebound during TI. In Chapters 4 and 5, we examined baseline pre-TI immune parameters and their association with viral rebound and CD4 count change during TI in HIV-infected individuals in the chronic phase of infection experiencing virologic failure before TI (Chapter 4) or with different levels of VL control while on therapy prior to TI (Chapter 5). We saw that chronic antigen stimulation from persistent viremia as well as co-infections such as with cytomegalovirus are associated with T-cell senescence, which may result in less favourable clinical outcomes during TI. / Consequently, results from this thesis contribute to further understanding of immune correlates of viral control in chronic HIV infection. New therapeutic vaccines and interventions should induce polyfunctional HIV-specific immune responses, broad Gag-specific immune responses, as well as reducing chronic antigen stimulation to prevent irreversible T-cell exhaustion. Taken together, these insights could potentially lead to the development of novel treatment interventions that could effectively control viral replication off cART.

The peripheral naive CD4 T cell pool is homeostatically regulated through a balance of thymic production, delivery of survival signals and homeostatic proliferation. CD4 recent thymic emigrants (RTEs) have a high T cell receptor excision circle (TREC) content and express high levels of CD31. We report premature thymic involution in RRMS, initiated by reduced numbers of naive CD4 T cells and various naive CD4 T cell subsets in peripheral blood. Further, CXCR4, a receptor involved in emigration from the thymus, and CD127 and Bcl-2 (survival signals) are upregulated in various naive CD4 T cell subsets in RRMS. As a compensatory process, naive CD4 T cells undergo homeostatic proliferation. This proliferation is a form of peripheral positive selection through self-MHC/self-antigen interaction and thus can contribute to the expansion of autoreactive T cells and predispose to development of RRMS.

Diss. (sammanfattning) Stockholm : Karol. inst. / Härtill 5 uppsatser.

No description available.

No description available.

Os exercícios com pesos (EP) representam importante estratégia para diminuir a sarcopenia e melhorar a capacidade funcional para realizar as atividades da vida diária de pessoas idosas. Além disso, os EP têm também sido recentemente sugeridos para restaurar os efeitos da imunossenescência. Portanto, este estudo teve como objetivo determinar o efeito de doze meses de um programa de exercícios com pesos em parâmetros imunológicos funcionais e quantitativos de mulheres idosas clinicamente saudáveis. As voluntárias foram selecionadas de acordo com o protocolo SENIEUR e 38 mulheres clinicamente saudáveis e fisicamente inativas (60 a 77 anos de idade) foram randomicamente divididas em um programa de exercícios com pesos de baixa intensidade (GE: 67,74 + 5,28 anos [n: 28]) ou em um grupo controle (CG: 68,69 + 2,98 anos [n: 14]). O programa de EP foi constituído de 3 séries de 12 repetições a 54,87 + 2,37% do teste de uma repetição máxima (1-RM) para cinco diferentes exercícios (supino reto sentado, puxada alta, remada, extensão de joelhos e leg press) realizados três vezes por semana durante 12 meses. A atividade citotóxica das células natural killer (NKCA), resposta linfoproliferativa à fito-hemaglutinina (PHA) e ao OKT3, quantificação de linfócitos (CD3+, CD3-CD19+, CD3-CD16+CD56+), subpopulações linfocitárias (CD4+, CD8+, CD56dim, CD56bright), assim como de moléculas de expressão celular (CD25+, CD28+, CD45RA+, CD45RO+, CD69+, CD95+, HLA-DR+) foram determinadas por ensaios imunológicos. As variáveis foram mensuradas a cada 6 meses durante período de um ano (pré-programa [PRÉ], 6 meses [6M] e 12 meses [12M]). A análise estatística demonstrou que o GE incrementou a força muscular em 44,2% e 48,1% após 6 e 12 meses, respectivamente (p<0,05); mas que houve aumento no consumo de oxigênio de pico (VO2pico) após 6 meses (14,7%, p<0.05). Não houve diferença significativa entre os grupos (exceto para 20:1 em 12M) ou em função do tempo (exceção para 40:1 em GE) na NKCA assim como na resposta proliferativa independente do mitógeno empregado. Foi observado decréscimo significativo (p<0,05) em GE para a contagem total de linfócitos (PRÉ x 12M), CD3+ (PRÉ x 12M), CD3+CD4+ (PRÉ x 12M), CD3-CD19+ (PRÉ x 6M), CD3+CD45RA+ (PRÉ x 6M; PRÉ x 12M), CD3+CD45RO+ (PRÉ x 12M; 6M x 12M), CD4+CD45RA+ (PRÉ x 6M), CD4+CD45RO+ (PRÉ x 12M; 6M x 12M), CD3+CD95+CD28+ (PRÉ x 6M), CD4+CD95+CD28+ (PRÉ x 6M), CD8+CD95+CD28+ (PRÉ x 12M) e para CD56dimCD25+HLA-DR+ (6M x 12M). O GC também demonstrou diminuição significativa (p<0,05) na contagem total de linfócitos (PRÉ x 12M), CD3-CD19+ (PRÉ x 6M; PRE x 12M), CD3-CD16+CD56+ (PRÉ x 12M; 6M x 12M), CD3+CD45RO+ (PRÉ x 12M) e para CD56dim (PRÉ x 12M). É possível que outra variável independente, que não o programa de exercícios com pesos e/ou alguma tendência sazonal tenham influenciado os resultados devido ao fato de ambos os grupos terem apresentado menores níveis de expressão celular durante o período do estudo. Os resultados deste estudo permitem concluir que doze meses de um programa de exercícios com pesos de leve intensidade são suficientes para incrementar a força muscular assim como o consumo de oxigênio de pico, mas não para melhorar parâmetros imunológicos funcionais e quantitativos de mulheres idosas clinicamente saudáveis. Portanto, possivelmente sugerindo que o \'limiar de efeito\' nos parâmetros imunológicos de mulheres idosas clinicamente saudáveis seja dependente do estímulo e/ou maior do que o necessário para incrementar a força muscular e/ou o VO2pico. / Resistance training program represents an important strategy to reduce sarcopenia, improving muscle strength and mass, and consequently, functional capacity to perform activities of daily living in elderly people. Additionally, resistance training program has been also recently suggested to restore the deleterious effects of aging process on immune system. Thus, the aim of this study was to determine the effect of twelve months of light resistance training program on functional and quantitative immunological paremeters of clinically healthy elderly women. Volunteers were selected by SENIEUR protocol and thirty-eight clinically healthy untrained females (aged 60-77 year-old) were randomly assigned to either a light resistance training program (RTP: 67.74 + 5.28 year-old [n: 28]) or a control group (CG: 68.69 + 2.98 year-old [n: 14]). The RTP consisted of three sets of twelve repetitions at 54.87 ± 2.37% of one repetition maximum test (1-RM) for five different exercises (seated bench press, lattissimus pull down, seated row, leg extension and leg press) performed three times per week during twelve months. Natural killer cell cytotoxic activity (NKCA), lymphoproliferative response to the phytohemaglutinin (PHA) and OKT3, and quantification of the lymphocytes (CD3+, CD19+, CD3-CD16+CD56+) and subpopulations (CD4+, CD8+, CD56dim, CD56bright) as well as cellular expression molecules (CD25+, CD28+, CD45RA+, CD45RO+, CD69+, CD95+, HLA-DR+) were determined by immunological assays. Variables were measured each 6 months during one year (pre-program [PRE], 6 months [6M] and 12 months [12M]). Statistical analysis showed that RTP volunteers increased muscle strength in 44.2% and 48.1% after 6 and 12 months, respectivelly (p<0.05), whilst there was increased in maximal oxygen peak (VO2peak) after only 6 months (14.7%, p<0.05). There were no statistically significant differences between both groups (unless 12M for the 20:1) or according to the time (unless 40:1 for RTP) when NKCA was analyzed. RTP group preserved lymphoproliferative response, while CG increased significantly the lymphoproliferative response to the PHA and OKT3. There were statistically significant decrease (p<0.05) for RTP volunteers to the total lymphocytes (PRE x 12M), CD3+ (PRE x 12M), CD3+CD4+ (PRE x 12M), CD3-CD19+ (PRE x 6M), CD3+CD45RA+ (PRE x 6M; PRE x 12M), CD3+CD45RO+ (PRE x 12M; 6M x 12M), CD4+CD45RA+ (PRE x 6M), CD4+CD45RO+ (PRE x 12M; 6M x 12M), CD3+CD95+CD28+ (PRE x 6M), CD4+CD95+CD28+ (PRE x 6M), CD8+CD95+CD28+ (PRE x 12M), and to the CD56dimCD25+HLA-DR+ (6M x 12M). GC volunteers also showed statistically significant decrease (p<0.05) to the total lymphocytes (PRE x 12M), CD3-CD19+ (PRE x 6M; PRE x 12M), CD3-CD16+CD56+ (PRE x 12M; 6M x 12M), CD3+CD45RO+ (PRE x 12M), and to the CD56dim (PRE x 12M). It is possible that the other independent variable, which not the RTP, and/or some seasonal tendency have influenced the results because to the fact of both groups had presented lower cell expression levels during the period of the study. The results of this study permit to conclude that twelve months of light RTP were sufficient to increase muscle strength and maximal oxygen peak, but not to improve functional and quantitative immunological parameters of clinically healthy elderly women. Thus, possibly suggesting that the \'threshold of effects\' on immunological paremeters on clinically healthy elderly women would be dose-response dependent and/or could be rather than that to increase muscle strength and/or maximal oxygen peak.

Development of a suitable animal model of AIDS is much needed in AIDS research to study infection and pathogenesis as well as to evaluate methods of prevention and treatment of HIV infection. Small animals such as rodents are attractive candidates for AIDS research due to the availability of various inbred and genetically engineered strains, extensive knowledge or their immune system, especially in mice, and the relative ease of breeding and maintaining animal colonies. Transgenic small animal models carrying entire HIV genome or selected genes have been instrumental to understand functions of HIV genes in vivo and their role in HIV pathogenesis. The type of cells in which HIV genes are expressed seems to be an import prerequisite for the study of HIV gene functions in transgenic mice. Mice constitutively expressing the entire HIV-1 genome or HIV-1 nef gene in CD4 + T cells and in the cells of macrophage/dendritic lineage develop an AIDS-like disease very similar to AIDS disease in humans. Similarly, expression of Nef in adult mice, using inducible system, results in the AIDS-like disease. This disease is characterized by thymic atrophy, impaired thymocyte maturation, loss of CD4+ T cells, increased activation and turnover of T cells, which can occur in the absence of lymphypenia, and non-lymphoid organ disease involving the lungs and kidneys. Susceptibility of adult mice to the pathological effects of Nef suggests that the AIDS-like disease in the constitutively expressing Nef Tg mice is not due to developmental defects caused by early expression of Nef. This model highlights the important role of Nef in HIV-1 pathogenesis. The high similarity in the disease in these Tg mice with human AIDS strongly suggest that these mice are a relevant model to study AIDS. This study further evidence that mouse cells can support functions of Nef and these Tg mice represent a unique model to study Nef functions in vivo in the context of the primary immune system. Moreover, the inducible Nef Tg model has given us the ability to control the level and time of expression of Nef which was impossible to do in the previously reported constitutive Nef Tg mouse models. These mice will be useful to study immune reconstitution since Nef expression can be turned off after withdrawal from dox.

Throughout history, human populations have continuously been challenged by new and emerging infectious diseases. For the past 26 years, sub-Saharan Africa and other countries around the world have been dealing with a pandemic caused by a relatively new pathogen called the human immunodeficiency virus (HIV). Although antiretroviral (ARV) therapies effectively reduce morbidity and mortality rates, the long term use in those who have access to treatment inevitably leads to drug-related toxicity and resistance. Even with a strong commitment from governments to expand and finance prevention and treatment programs, transmission rates continue to outpace the benefits of these efforts. Therefore to effectively eradicate the disease, research is focusing on the design of protective and therapeutic vaccines. The first major step in designing these alternative therapies is to define correlates of immune protection. / The research presented in this thesis focuses on characterizing the quantitative and qualitative features of T cell immune responses in individuals who spontaneously control viral replication and exhibit a benign course of disease while remaining off ARV therapy. A comprehensive analysis of HIV-specific IFN-gamma secreting immune responses revealed that neither the breadth nor the magnitude of responses directed against the entire HIV proteome accurately predicts the viral load or rate of CD4 decline. Subsequent analyses showed that the preferential targeting of Gag was associated with reduced rates of CD4 decline and was later confirmed in a cohort of individuals in primary infection whereby the relative breadth and magnitude of Gag p24 was inversely correlated with viral load set point. / The maintenance of polyfunctional immune responses in HIV-infected subjects with a benign course of disease prompted us to develop a method that could comprehensively assess the breadth, magnitude and specificity of three functionally distinct subsets of HIV-specific lymphocytes (single IFN-gamma, single IL-2 and dual IFN-gamma/IL-2 secretors). Survey of immune responses in chronically infected individuals revealed that only the breadth and magnitude of dual IFN-gamma/IL-2 secreting lymphocytes correlated with reduced viral loads and increased CD4 counts suggesting that secretion of IFN-gamma alone was a poor correlate of protection. We also showed that the contribution of polyfunctional lymphocytes to the total response was greater for epitopes restricted by major histocompatibility complex (MHC) class I alleles associated with slow disease progression compared to those restricted by alleles associated with rapid or neutral rates of HIV disease progression. / Taken together, this work supports the view that immune monitoring of infected and vaccinated individuals should include methodologies capable of detecting both IFN-gamma and IL-2 secretion from responding T lymphocytes. The studies presented here have furthered our understanding of what constitutes protection from disease progression emphasizing that both specificity and polyfunctionality are features of effective control of viral replication.