Apesar de uma diversidade de estudos científicos, em que dezenas de milhares de pacientes foram analisados e tratados, a sepse continua sendo um grande desafio para a medicina contemporânea. Investigações bem conduzidas levaram a uma reavaliação do modelo clássico da sepse, tradicionalmente vista como um processo descontrolado de hiperinflamação sistêmica, uma vez que se observou a existência de uma atividade antiinflamatória ao longo do seu curso evolutivo. Nesse contexto, o comportamento do sistema imune inato se assemelha ao de indivíduos idosos submetidos ao fenômeno da imunossenescência, interseção ainda mais relevante ao considerarmos o crescente incremento na faixa etária média dos pacientes internados em UTI. O presente estudo visou a estabelecer a epidemiologia da sepse em um hospital público de um país de renda media, como é o caso do Brasil. Ademais, através de citometria de fluxo, buscamos definir a cinética da expressão monocitária de moléculas HLA-DR e CD64 ao longo do processo de envelhecimento humano. Comparamos essas observações com o comprometimento do sistema imune inato visto na sepse visando discriminar as alterações da senescência do sistema imune associada ao envelhecimento daquelas associadas ao fenômeno da imunoparalisia da sepse. Na investigação epidemiológica, nós encontramos uma taxa de ocorrência de 5,9 casos de sepse por 100 pacientes e uma densidade de incidência de 6,4 casos por 1000 pacientes-dia. Documentamos ainda sua associação com uma elevada incidência de sepse e documentamos sua associação com uma elevada taxa de comordidades crônicas. A sepse foi diagnosticada tardiamente (72% dos casos após 12 horas de evolução) e em estágio avançado como atestado pelos elevados escores de gravidade de doença e de disfunção orgânica. O presente estudo identificou vários obstáculos à efetiva implementação das recomendações da Surviving Sepsis Campaign. No segundo estudo, observamos correlações negativas significativas entre idade e intensidade de expressão dos biomarcadores avaliados. Durante a sepse, a expressão de mHLA-DR foi ainda mais reduzida, enquanto a expressão de CD64 foi majorada. Para melhor discriminar imunossenescência de imunoparalisia, valores de cutoff de 9.700 m/c para mCD64 e de 7.000 m/c para mHLA-DR forneceram as melhores combinações de sensibilidade e especificidade. / Despite a multiplicity of trials enrolling several thousands of treated patients, sepsis still represents a major challenge to contemporary medicine. Since several well-conducted investigations have consistently indicated the existence of an anti-inflammatory activity during sepsis evolution, there has been a reappraisal of the classic sepsis model in which the condition was traditionally seen as an uncontrolled systemic hyper-inflammatory process. During sepsis, the innate immune system behavior has some similarities to the immunosenescence process experienced by elderly people; something that is even more relevant when considering the progressive increase of the mean age of the patients admitted to ICUs along the last decades. This study investigated the epidemiology of sepsis in a public university hospital from a middle-income country, as is the case of Brazil. Also, applying a flow cytometry methodology, we tried to establish the kinetics of the molecular expression of HLA-DR and CD64 on monocytes during human aging. To put it in context, we compared such findings with those obtained from septic patients aiming at the discrimination between changes related to the normal aging process from those of the immunoparalysis phenomenon seen in sepsis. In the epidemiological arm of this study, we found an occurrence rate of 5.9 cases of sepsis per 100 patients, and an incidence density of 6.4 cases per 1000 patient-days. We documented the association of sepsis with an elevated rate of chronic comorbid conditions. Sepsis was detected late (72% evolving for more than 12 hours) and in advanced stages as indicated by severity and organ failure scores. The study detected several obstacles to the effective implementation of the Surviving Sepsis Campaign recommendations. In the second study, we observed significant negative correlations between age and the levels of HLA-DR and of CD64 expression on monocytes. During sepsis, HLA-DR expression decreased further while CD64 expression was upregulated. To better discriminate immunosenescence from immunoparalysis, cutoff values of 9,700 m/c for mCD64 and of 7,000 m/c for HLA-DR provided the best combination of sensitivity and specificity.

Apesar de uma diversidade de estudos científicos, em que dezenas de milhares de pacientes foram analisados e tratados, a sepse continua sendo um grande desafio para a medicina contemporânea. Investigações bem conduzidas levaram a uma reavaliação do modelo clássico da sepse, tradicionalmente vista como um processo descontrolado de hiperinflamação sistêmica, uma vez que se observou a existência de uma atividade antiinflamatória ao longo do seu curso evolutivo. Nesse contexto, o comportamento do sistema imune inato se assemelha ao de indivíduos idosos submetidos ao fenômeno da imunossenescência, interseção ainda mais relevante ao considerarmos o crescente incremento na faixa etária média dos pacientes internados em UTI. O presente estudo visou a estabelecer a epidemiologia da sepse em um hospital público de um país de renda media, como é o caso do Brasil. Ademais, através de citometria de fluxo, buscamos definir a cinética da expressão monocitária de moléculas HLA-DR e CD64 ao longo do processo de envelhecimento humano. Comparamos essas observações com o comprometimento do sistema imune inato visto na sepse visando discriminar as alterações da senescência do sistema imune associada ao envelhecimento daquelas associadas ao fenômeno da imunoparalisia da sepse. Na investigação epidemiológica, nós encontramos uma taxa de ocorrência de 5,9 casos de sepse por 100 pacientes e uma densidade de incidência de 6,4 casos por 1000 pacientes-dia. Documentamos ainda sua associação com uma elevada incidência de sepse e documentamos sua associação com uma elevada taxa de comordidades crônicas. A sepse foi diagnosticada tardiamente (72% dos casos após 12 horas de evolução) e em estágio avançado como atestado pelos elevados escores de gravidade de doença e de disfunção orgânica. O presente estudo identificou vários obstáculos à efetiva implementação das recomendações da Surviving Sepsis Campaign. No segundo estudo, observamos correlações negativas significativas entre idade e intensidade de expressão dos biomarcadores avaliados. Durante a sepse, a expressão de mHLA-DR foi ainda mais reduzida, enquanto a expressão de CD64 foi majorada. Para melhor discriminar imunossenescência de imunoparalisia, valores de cutoff de 9.700 m/c para mCD64 e de 7.000 m/c para mHLA-DR forneceram as melhores combinações de sensibilidade e especificidade. / Despite a multiplicity of trials enrolling several thousands of treated patients, sepsis still represents a major challenge to contemporary medicine. Since several well-conducted investigations have consistently indicated the existence of an anti-inflammatory activity during sepsis evolution, there has been a reappraisal of the classic sepsis model in which the condition was traditionally seen as an uncontrolled systemic hyper-inflammatory process. During sepsis, the innate immune system behavior has some similarities to the immunosenescence process experienced by elderly people; something that is even more relevant when considering the progressive increase of the mean age of the patients admitted to ICUs along the last decades. This study investigated the epidemiology of sepsis in a public university hospital from a middle-income country, as is the case of Brazil. Also, applying a flow cytometry methodology, we tried to establish the kinetics of the molecular expression of HLA-DR and CD64 on monocytes during human aging. To put it in context, we compared such findings with those obtained from septic patients aiming at the discrimination between changes related to the normal aging process from those of the immunoparalysis phenomenon seen in sepsis. In the epidemiological arm of this study, we found an occurrence rate of 5.9 cases of sepsis per 100 patients, and an incidence density of 6.4 cases per 1000 patient-days. We documented the association of sepsis with an elevated rate of chronic comorbid conditions. Sepsis was detected late (72% evolving for more than 12 hours) and in advanced stages as indicated by severity and organ failure scores. The study detected several obstacles to the effective implementation of the Surviving Sepsis Campaign recommendations. In the second study, we observed significant negative correlations between age and the levels of HLA-DR and of CD64 expression on monocytes. During sepsis, HLA-DR expression decreased further while CD64 expression was upregulated. To better discriminate immunosenescence from immunoparalysis, cutoff values of 9,700 m/c for mCD64 and of 7,000 m/c for HLA-DR provided the best combination of sensitivity and specificity.

L’immunité cellulaire T est probablement la plus affectée par le vieillissement de l’être humain avec un retentissement sur les déficiences immunitaires au niveau humoral comme au niveau cellulaire. Elle contribue à une augmentation de la susceptibilité des sujets âgés aux maladies infectieuses, à l’auto-immunité, au cancer et à la baisse de la réponse vaccinale mais son intrication avec les pathologies notamment chroniques est mal connue.Notre travail s’est intéressé aux temps précoces de l’activation cellulaire T et à la mise au point d’un modèle permettant l’évaluation de la capacité d’activation des lymphocytes T vis-à-vis de surfaces recouvertes d’anticorps activateurs (Ac antiCD3) ou non (Ac antiHLA-ABC), observés en microscopie IRM.Ce modèle nous permet de montrer que l’étalement des lymphocytes T sur les surfaces est précédé d’ondulations verticales de la membrane générant des contacts transitoires durant environ 1 minute et que la cinétique d’étalement est ensuite sept à huit fois plus rapide sur les surfaces supposées activatrices. D’autre part, avec des dilutions successives des Ac antiCD3 sur les lamelles, l’aire moyenne de la zone de contact des lymphocytes T est fortement corrélée aux réponses prolifératives mesurées avec le test de prolifération lymphocytaire standard au CFSE.Ce modèle permet également l’observation en microscopie à fluorescence de différentes protéines phosphorylées des cascades de signalisation. L’étude de la fluorescence des cellules T purifiées, fixées et marquées par l’Ac anti-phospho-Erk en fonction des dilutions d’Ac antiCD3 sur la lamelle permet d’objectiver une décroissance de la moyenne de fluorescence avec les dilutions.Notre modèle a l’avantage d’étudier les temps précoces de l’activation. Il pourrait s’appliquer à l’étude de l’immunosénescence. Ainsi l’étude du contact entre une cellule T et des surfaces activatrices pourrait s’avérer être une méthode simple et utile d’exploration précoce de l’activation de lymphocytes T en pratique clinique.L’application de notre modèle à l’étude de l’immunité du sujet âgé est en cours dans le cadre d’une étude prospective comparative de sujets âgés malades et de sujets âgés « sains » recrutés selon les critères du « senieur protocole ».En conclusion, il s’agit d’un travail d’approche mixte à la fois fondamentale et clinique permettant d’étudier les phénomènes d’immunosénescence et leur lien avec la pathologie. / Our work was focused on early events in T cell activation and has proposed a model system for the Interference reflection microscopy-based monitoring of T lymphocytes activation by antibodies coated-surfaces, either with activating anti-CD3 or control anti-HLA-ABC.Our system has allowed to observe that T cell spreading on surfaces is preceded by vertical membrane undulations, generating transient contacts with the underlying surface for approximately 1 minute, and that spreading kinetics of are seven to eight times faster when activating antibodies are involved. At increasing dilutions of antiCD3 Ab on surfaces, the contact area formed between T lymphocytes and activating surfaces is linearly correlated to proliferative responses, as measured by CFSE labelling. Our system allowed also the direct observation, by fluorescent microscopy, of signaling-dependent protein phosphorylation. T cells exposed to decreasing concentrations of antiCD3 had lower phospho-Erk levels, as shown upon fixation and anti-phospho-Erk labeling. Allowing the study early events in T cell activation, our model could be applied to the study of immunosenescence. To monitor contacts between T cells and activating surfaces could be a simple and useful method for early exploration of T lymphocytes activation in clinical practice.We are currently carrying out a comparative prospective study on old sick and healthy subjects, recruited according to " senior protocol " criteria. In conclusion, our work represents a fundamental and clinical mixed approach for the study of immunosenescence-related phenomena and their link with pathologies.

Herpesviruses have co-speciated with several vertebrate and invertebrate animals throughout the history of evolution. In the immunocompetent human host, primary infection is usually benign, whereafter the virus is brought into life-long latency. Viral reactivation can however cause severe disease in immunocompromised, and rarely also in immunocompetent, patients. The overall aim of this thesis was to study the immunologic effects of cytomegalovirus (CMV) and herpes simplex type 1 (HSV-1) infection in neurocognitive disorders. CMV is known to promote T-cell differentiation towards a more effector-oriented phenotype, similar to what is seen in the elderly. We have addressed the frequency of CMV-specific CD8+ T-cells in Alzheimer's disease (AD). Furthermore, we have investigated whether AD patients present with a different CMV-specific immune profile, overall CD8 phenotype or inflammatory cytokine response to anti-CD3/CD28 beads, CMV pp65 and amyloid beta. Subjects with AD presented with a lower proportion of CMV-specific CD8+ T-cells compared to non-demented (ND) controls, but no differences in overall CD8 differentiation were seen. Overall, AD subjects presented with a more pro-inflammatory peripheral blood mononuclear cell (PBMC) phenotype. When PBMCs were challenged with CD3/CD28-stimulation, CMV seropositive AD subjects presented with more IFN-γ release than both CMV seronegative AD subjects and CMV seropositive ND controls. For effective screening of humoral herpesvirus immunity, both in research and in clinical practice, efficient immunoassays are needed. We have addressed the methodology of multiplex herpesvirus immunoassays and related bioinformatics and investigated antibody levels in AD patients and ND controls. Subjects with AD presented with lower levels of human herpesvirus 6 (HHV-6) IgG. However, there was no difference in HHV-6 DNA levels in PBMCs between the groups. Herpes simplex encephalitis (HSE) is a devastating disease, where antiviral treatment has greatly decreased mortality but not eliminated the associated long-term neurocognitive morbidity. We have investigated the correlation between N-Methyl-D-Aspartate Receptor (NMDAR) autoimmunity and recovery of neurocognitive functions after HSE. Approximately one quarter of all HSE cases developed NMDAR autoantibodies within 3 months after onset of disease. Antibody development was associated with an impaired neurocognitive recovery during the two year follow-up and could become an important therapy guiding factor in the future.

Data de aprova??o ausente. / Submitted by Jos? Henrique Henrique (jose.neves@ufvjm.edu.br) on 2017-02-13T18:51:06Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) tatiana_fernandes_araujo_almeida.pdf: 2531556 bytes, checksum: 11a2583c0414dc55d6f907a2566a4de5 (MD5) / Approved for entry into archive by Rodrigo Martins Cruz (rodrigo.cruz@ufvjm.edu.br) on 2017-03-06T12:21:52Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) tatiana_fernandes_araujo_almeida.pdf: 2531556 bytes, checksum: 11a2583c0414dc55d6f907a2566a4de5 (MD5) / Made available in DSpace on 2017-03-06T12:21:52Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) tatiana_fernandes_araujo_almeida.pdf: 2531556 bytes, checksum: 11a2583c0414dc55d6f907a2566a4de5 (MD5) Previous issue date: 2016 / O carcinoma espinocelular oral (CECO) ? uma neoplasia maligna que acomete principalmente idosos. O desencadeamento da doen?a ? relacionado ? exposi??o aos fatores de risco e ao decl?nio da fun??o imune associada ao envelhecimento cronol?gico. Alguns estudos t?m demonstrado um aumento na incid?ncia de CECO em adultos jovens, sugerindo envolvimento de outros fatores na etiologia da doen?a, como uma altera??o do sistema imunol?gico. As c?lulas dendr?ticas (CDs) s?o c?lulas apresentadoras de ant?geno profissionais e estimuladoras eficazes para a expans?o clonal de linf?citos. Acredita-se que uma altera??o da fun??o das CDs em pacientes com c?ncer contribui para a falha da resposta antitumoral, levando a uma consequente progress?o da doen?a. O objetivo deste estudo foi avaliar, atrav?s da imunoistoqu?mica, se h? diferen?a na quantifica??o tissular dos subgrupos de CDs, associada ? idade, em esp?cimes de bi?psias de CECO. Para isso, foram selecionados casos de CECO de pacientes em tr?s diferentes faixas et?rias: G1 (< 40 anos de idade, n = 12), G2 (? 40 at? < 60 anos de idade, n = 15) e G3 (? 60 anos de idade, n = 14). Os marcadores utilizados foram S100, CD1a, CD207 (para CDs imaturas, imCDs), CD83 e CD208 (para CDs maduras, mCDs). As imagens foram capturadas dos campos com maior intensidade de marca??o nas l?minas histol?gicas e a quantifica??o celular foi realizada com o aux?lio do software Image J. A frequ?ncia e localiza??o das CDs foram avaliadas e analizadas estatisticamente nas regi?es intratumoral (intertumoral e ou estromal) e extratumoral. No geral, imCDs foram significativamente mais frequentes que mCDs em todos os grupos. ImCDs e mCDs mostraram preferencialmente localiza??o intratumoral e extratumoral respectivamente. Comparando G1 versus G2 / G3 foi observada um significativo menor n?mero de mCDs em G1. Na compara??o de G1 em rela??o a G2 ou G3 houve um n?mero significativamente menor de ambas, imCDs e mCDs. Nossos resultados mostram que existe uma menor quantidade de imCDs e mCDs em CECO afetando pacientes jovens em compara??o com idosos, sugerindo um comprometimento da resposta imune antitumoral em G1e permitindo a progress?o do tumor. / Disserta??o (Mestrado) ? Programa de P?s-Gradua??o em Odontologia, Universidade Federal dos Vales do Jequitinhonha e Mucuri, [2016]. / Oral squamous cell carcinoma (OSCC) is a malignant neoplasm that mainly affects elderly people. The onset of the disease is related with exposure to risk factors and the decline of immune function associated with chronological aging. Previous studies have demonstrated an increased incidence of CECO in young adults, suggesting involvement of other factors in the etiology of the disease, such as a change in the immune system. Dendritic cells (DCs) are professional antigen presenting cells and effective for stimulating clonal expansion of lymphocytes. It is believed that a change in the function of DC in cancer patients contributes to the failure of the anti-tumor response, leading to a consequent progression of the disease. The aim of this study was to evaluate, by immunohistochemistry, if there were differences in tissue quantification of DCs subsets, associated with age in specimens of OSCC biopsies. For this OSCC cases were selected at three different ages: G1 (< 40 years old, n = 12), G2 (? 40 to <60 years old, n = 15) and G3 (? 60 years old, n = 14). The markers used were S100, CD1a, CD207 (for immature DCs, imDCs), CD83 and CD208 (for mature DCs, mDCs). Images were captured from fields with higher intensity staining in histological sections and cell quantification was performed with Image J software help. The frequency and localization of immunostained DCs were analyzed in intratumoral (intranestal and/or extranestal) and extratumoral areas and statistically compared. Overall, imDCs than mDCs were significantly more frequent in all groups. ImDCs and mDCs showed preferential intratumoral and extratumoral localization, respectivel. Comparing G1 than G2/G3 showed a significant lesser number of mDCs. G1 versus G2 or G3 there is a significantly lower number of both imDCs and mDCs. Our results show a lower number of both imDCs and mDCs in OSCC affecting younger than elderly patients, suggesting impairment of an effective antitumor immune response in G1 and enabling tumor progression, showing a gradual establishment of the antitumor immune response mediated by DCs according to the age, but with defects in quality.

Cette thèse s’insère dans le cadre général de l’étude de la dynamique des populations. La population prise en compte étant constituée de cellules souches normales et/ou cancéreuses. Nous proposons et analysons trois modèles mathématiques décrivant la dynamique de cellules souches. Le premier modèle proposé est un modèle d’équations aux dérivées partielles structurées en âge que nous transformons, via la méthode des caractéristiques, en un système d'équations différentielles à retard pour lequel on étudie l'existence et la stabilité des points d'équilibres. On effectue, après, des simulations numériques permettant d'illustrer le comportement des états d'équilibres. Dans le deuxième modèle, on considère que la durée du cycle cellulaire dépend de la population totale de cellules quiescentes. La méthode des caractéristiques nous permet de réduire notre modèle structuré en âge à un système d'équations différentielles avec un retard dépendant de l'état pour lequel on effectue une analyse détaillée de la stabilité. Nous confirmons, ensuite, les résultas analytiquement obtenus par des simulations numériques. Pour le troisième et dernier modèle de cette thèse, on propose un système d'équations différentielles ordinaires décrivant la dynamique de cellules souches saines et cancéreuses et prenant en compte leurs interactions avec les réponses immunitaires. Ce modèle nous a permis de souligner l'ampleur de l'impact que peuvent avoir différentes infections sur la prolifération tumoral que ce soit par le biais de leurs fréquences, leurs durées ou la façon dont ils agissent sur le système immunitaire. / This thesis fits into the general framework of the study of population dynamics. The population particularly considered in this work is comprised of stem cells with both cases of healthy and cancerous cells being investigated. We propose and analyze three mathematical models describing stem cells dynamics. The first model is an age-structured partial differential model that we reduce to a delay differential system using the characteristics method. We investigate the existence and stability of the steady states of the reduced delay differential system. We, then, conduct some numerical simulations to illustrate the behavior of the steady states. In the second model, the duration of the cell cycle is considered to depend upon the total population of quiescent cells. The method of characteristics reduces the age-structured model to a system of differentialequations with a state-dependent delay. We perform a detailed stability analysis of the resulting delay differential system. We confirm the analytical results by numerical simulations. The third and final model, proposed in this thesis, is an ordinary differential equations model describing healthy and cancerous stem cells dynamics and their interactions with immune system responses. Through this model, we show that the frequency, the duration of infections and their action (positive or negative) on immune responses may impact significantly tumor proliferation.

Inflammaging is a hallmark of human aging. Defined as low-grade, chronic inflammation, it is characterized by heightened proinflammatory cytokines in the blood and tissues and predicts morbidity and mortality. Despite this, the etiology of inflammaging and its role in infection have remained elusive, an issue this thesis addressed. First, we provided a comprehensive overview of an intranasal Streptococcus pneumoniae colonization model (Chapter 2). We described in detail the colonization technique, and demonstrated how to isolate and phenotype recruited cells, quantify bacterial load and measure production of immune mediators in the nasopharynx. Since both myeloid cell recruitment and tumour necrosis factor (TNF) production were increased following S. pneumoniae colonization with age, we investigated whether TNF directly augmented monocyte frequency (Chapter 3). TNF increased CCR2 expression on monocytes in old mice, leading to their enhanced egress from the bone marrow, resulting in enrichment of this population in the circulation. Monocyte numbers directly influenced plasma IL-6 levels, and this negatively impacted anti-bacterial responses, as monocyte blockade improved pneumococcal clearance in old mice. Lastly, to better understand the fundamental source of inflammaging, we studied the impact of the host microbiome on its development. This work was rooted in Elie Metchnikoff’s early predictions that leakage of intestinal bacterial products could dysregulate macrophage function, resulting in inflammation that would progress aging (Chapter 4). We showed that old mice had increased intestinal permeability, aberrant expression of cellular junction genes and increased microbial translocation from the gut to the blood. Germ-free mice lived longer than their conventionally colonized counterparts, and were protected from the development of inflammaging and defective macrophage function. Together, these studies resolve a major disparity in the field by demonstrating that systemic TNF production is initiated by increased levels of circulating bacterial products, driving functional defects in myeloid cells, which ultimately impairs anti-bacterial immunity. / Thesis / Candidate in Philosophy

Immunosenescence is considered a possible factor in the development of age-related macular degeneration and choroidal neovascularization (CNV). However, age-related changes of myeloid cells (MCs), such as microglia and macrophages, in the healthy retina or during CNV formation are illdefined. In this study, Cx3cr1-positive MCs were isolated by fluorescence-activated cell sorting from six-week (young) and two-year-old (old) Cx3cr1GFP/+ mice, both during physiological aging and laser-induced CNV development. High-throughput RNA-sequencing was performed to define the age-dependent transcriptional differences in MCs during physiological aging and CNV development, complemented by immunohistochemical characterization and the quantification of MCs, as well as CNV size measurements. These analyses revealed that myeloid cells change their transcriptional profile during both aging and CNV development. In the steady state, senescent MCs demonstrated an upregulation of factors contributing to cell proliferation and chemotaxis, such as Cxcl13 and Cxcl14, as well as the downregulation of microglial signature genes. During CNV formation, aged myeloid cells revealed a significant upregulation of angiogenic factors such as Arg1 and Lrg1 concomitant with significantly enlarged CNV and an increased accumulation of MCs in aged mice in comparison to young mice. Future studies need to clarify whether this observation is an epiphenomenon or a causal relationship to determine the role of immunosenescence in CNV formation.

The oldest-old (people aged 80 or older) constituted 5 % of the population in Sweden in 2000, an increase from 1.5 % fifty years earlier. The immune system undergoes dramatic changes at high age, sometimes referred to as “immunosenescence”. However, the natures of these changes, and in particular, their clinical consequences are incompletely understood. In a previous longitudinal study, a set of immune parameters were identified and termed immune risk phenotype (IRP) because of an association with increased mortality. The IRP consists of changes in the T lymphocyte compartment, in particular an inverted CD4/CD8 ratio. The IRP was found to be associated with cytomegalovirus (CMV) infection, which through expansions of cytolytic anti-viral CD8 cell responses was ascribed a role in the development of IRP. The general aim of this thesis was to increase the knowledge of changes in the immune system and their clinical consequences in the oldest-old. The population-based random sample of the longitudinal NONA-Immune Study (n = 138, mean age 90 years at baseline) was used for all investigations. In paper I, the effects on sample size of various exclusion protocols for immune studies of the elderly was examined. The commonly used SENIEUR protocol, selecting individuals representing ‘normal ageing’, excluded 90 % of nonagenarians. Based on different protocol criteria, individuals were grouped into ‘very healthy’, ‘moderately healthy’ or ‘frail’. The prevalence of CMV was similar across the groups. Further, differentiated CD8 populations associated with CMV, i.e. those expressing CD56, CD57 and CD45RA while lacking expression of CD27 and CD28, were equally distributed across the groups of the oldest-old, but were, as expected, significantly increased in the elderly compared to a middle aged control group. The findings showed that lymphocyte subsets associated with IRP might serve as significant biomarkers of ageing independent of the overall health status, also supporting the notion that immunological studies of the oldest-old should be done in population-based non-selected populations. The IRP and the presence of low-grade inflammation, for example increase of   IL-6 in plasma, constitute major predictors of 2-year mortality in the oldest-old. In paper II, the CD4/CD8 ratio and IL-6 were found to predict 97 % of observed survival and 57 % of deaths over 2 years. The impact of IRP and IL-6 on 2-year survival was independent of age, sex and several diseases. The longitudinal design allowed temporal evaluations, suggesting a sequence of events starting with IRP and leading to inflammation in the decline state.                                      Four-year mortality in the oldest-old (paper III) was found to be mainly related to markers of inflammation and IRP. Individuals with both inverted CD4/CD8 ratio and high IL-6 level had significantly higher 4 year mortality (82 %) compared to individuals with CD4/CD8 ratio ³ 1 and low IL-6 level (29 %) at baseline. The presence of IRP and increased IL-6 level showed some associations with presence of diseases; in particular, IL6 was associated with the presence of cognitive impairment. However, despite being strong predictors of mortality, IRP and IL-6 could not be linked to any specific cause of death, probably due to the multi-factorial nature of these factors.                                                                                                                             The prevalence of antinuclear antibodies (ANA) in the oldest-old was higher compared to younger controls (paper IV). The difference across age was most pronounced in men, showing low levels at younger age, whereas the prevalence among the oldest-old men reached a similar level as in women. There was no association between the presence of ANA and IRP, CMV status or health status in the oldest-old.

L’atteinte de l’âge sur les populations lymphocytaires T conventionnelles CD4 et CD8 avec l’avancée en âge est relativement bien décrite en périphérie lymphoïde secondaire chez la souris, et dans le sang périphérique chez l’homme. Deux paramètres sont observés : réduction du nombre de ces cellules et altération du ratio naïve/mémoire. À l’inverse, l’évaluation des tissus lymphoïdes tertiaires et des tissus extra lymphoïdes dans les réponses immunes, reste à affiner. Notre étude au cours du vieillissement physiologique du compartiment T fut menée dans des tissus lymphoïdes et non lymphoïdes de souris C57BL/6 wild-type, âgées entre 2 et 6 mois, entre 10 et 14 mois et entre 22 et 26 mois. Nous avons démontré que la lymphopénie T classiquement décrite liée au vieillissement dans les organes lymphoïdes secondaires ne s’applique pas à tout l’organisme : les compartiments intestinaux étudiés présentent une accumulation de cellules TCRαβ+ CD4+ (TCD4) et CD8+ (TCD8). Nos résultats dévoilent un impact différentiel du vieillissement sur le nombre absolu des différents compartiments cellulaires TCRαβ+ dans les organes lymphoïdes et la muqueuse intestinale. Ces résultats suggèrent donc que la lymphopénie T décrite dans les organes lymphoïdes s’établissant au cours du vieillissement pourrait être essentiellement liée à une redistribution des lymphocytes. A l’inverse, la persistance des cellules T régulatrices dans les organes lymphoïdes secondaires pourrait être liée à une production locale dans la muqueuse intestinale. Il semble donc que l’équilibre TCD8/TCD4 peut être différemment affecté selon le site considéré et cette observation peut fournir une justification pour la plus grande susceptibilité aux infections observée avec l’âge. / Consequences of ageing on conventional CD4 and CD8 T lymphocytes populations is relatively well described in murine secondary lymphoid organs and in human peripheral blood: reduction the number of these cells and alteration of naïve/effector-memory ratio in favour of effector-memory cells. Conversely, evaluation in tertiary lymphoid tissues and non-lymphoid tissues remains to be refined. We conducted an exhaustive analysis of T cell compartments during physiological aging in lymphoid and non-lymphoid tissues isolated from wild-type C57BL/6 mice aged of 2 to 6 months, 10 to 14 months and 22 to 26 months. We demonstrated that T lymphopenia described classically associated with aging in the secondary lymphoid organs does not apply to the whole organism: intestinal compartments studied show an accumulation of TCRαβ+ CD4+ cells (TCD4) and CD8+ (TCD8). Our results reveal a differential impact of aging on the absolute number of different TCRαβ+ cellular compartments in lymphoid organs and intestinal mucosa. T cell lymphopenia in secondary lymphoid organs currently associated to ageing may essentially reflect T cell redistribution. TCD8/TCD4 balance may be affected differently depending on the site considered and this observation may provide a rationale for the greater susceptibility to infection observed with age.