Alterações metabólicas são muito frequentes em doentes graves. Sepse grave e choque séptico são condições clínicas muito prevalentes em unidades de terapia intensiva (UTI). A mortalidade da sepse grave e, especialmente do choque séptico, persiste alta, apesar das terapêuticas desenvolvidas nas últimas décadas. Controle rigoroso da glicemia parece ser uma terapia adjuvante muito importante, especialmente em doentes cirúrgicos graves. Ainda há controvérsias sobre o controle glicêmico rigoroso em doentes clínicos. Além da hiperglicemia, alguns estudos procuraram associar distúrbios no metabolismo de lipoproteínas e pior prognóstico para doentes graves. Também foi descrita a associação de hiperglicemia e quantidades mais baixas de lipoproteínas, sugerindo, possivelmente, o controle glicêmico rigoroso como fator importante para correção dos distúrbios do metabolismo de lipoproteínas. Neste estudo, dosamos LDL, HDL, triglicerídeos, colesterol total, ácidos graxos livres e Ox-LDL em 63 pacientes com diagnóstico de sepse grave ou choque séptico, divididos em dois grupos, sendo um grupo mantendo controle glicêmico rigoroso e outro grupo mantendo um controle glicêmico mais liberal, com internação em unidade de terapia intensiva (UTI) nas primeiras 72 horas de internação. Independentemente do grupo alocado, as concentrações séricas de LDL, HDL, colesterol total estiveram abaixo dos valores considerados normais. De outro modo, as concentrações séricas de ácidos graxos livres, triglicérides e Ox-LDL estiveram acima dos valores considerados normais. Ao longo das 72 horas houve manutenção das concentrações séricas de HDL e de colesterol total e das concentrações séricas elevados de ox-LDL e triglicérides. Houve um aumento progressivo das concentrações séricas de LDL e diminuição das concentrações séricas de ácidos graxos livres mais pronunciada nos doentes submetidos a controle glicêmico rigoroso. Ao longo do período de estudo, os pacientes sobreviventes apresentaram menores necessidades de insulina exógena, porém com concentrações séricas glicêmicas similares. As dosagens de Ox-LDL, LDL, HDL e PCR permaneceram similares entre os sobreviventes e os não sobreviventes / Metabolic disturbances are very frequent among critical care patients. Severe sepsis and septic shock are clinical conditions responsible for a great number of patients admitted to ICU. Severe sepsis and septic shock mortality rates remain high instead of new approaches developed in last decades. Intensive glycemic control appeared to be an important adjuvant therapy, especially among surgical intensive care patients. There are still some controversies about the benefits of intensive glycemic control among clinical intensive care patients. Beyond hyperglycemia, some studies have tried to associate lipid metabolism disturbances to worse prognosis. There are also descriptions of association between hyperglycemia and lower lipoproteins levels, suggesting the possible positive effects of intensive glycemic control and better control of lipid disturbances. In this study, we collected sequential serum LDL, HDL, triglycerides, total cholesterol, free fatty acids and Ox-LDL for 63 patients diagnosed as severe sepsis or septic shock admitted to ICU, in the first 72 hours after beginning of the symptoms. Patients were randomly allocated into two different groups, one for intensive glycemic control and the other maintaining more liberal glycemic levels. Results: Serum levels of LDL, HDL, and total cholesterol were below levels considered normal in both groups. Contrary, serum levels of free fatty acids, triglycerides and Ox-LDL were above normal levels in both groups. Along initial 72 hours we noticed a clear increase in LDL serum levels and decrease in free fatty acids serum levels more pronounced in the intensive glycemic control group. Survivors needed less dosages of exogenous insulin, despite of similar glycemic levels. Serum levels of Ox-LDL, LDL, HDL and CRP were similiar for survivors and non-survivors

Sepsis é uma disfunção de órgãos causada por uma resposta desregulada do hospedeiro em decorrência de uma infecção e que eventualmente leva a morte. A identificação de moléculas que minimizem este processo pode fornecer alvos terapêuticos para prevenir a falência de órgãos durante a sepse. O supressor de sinalização de citocinas 1 (SOCS1) é conhecido por regular negativamente a sinalização de receptores de citocinas e de receptores do tipo Toll (TLRs). No entanto, os alvos celulares e mecanismos moleculares envolvidos nas ações de SOCS1 durante a sepse são desconhecidos. Para determinar o papel de SOCS1 durante a sepse polimicrobiana, camundongos C57BL/6 foram tratados com um peptídeo inibidor do domínio KIR (kinase inhibitor region) do SOCS1 (iKIR) e submetidos à CLP (ligação e perfusão do ceco). O tratamento com iKIR aumentou a mortalidade, a carga bacteriana e a produção de citocinas inflamatórias induzida pela CLP. Além disso, observou-se que animais deficientes de SOCS1 nas células mielóides (SOCS1?myel) também tiveram aumento na carga bacteriana e na produção de citocinas proinflamatórias, quando comparados com camundongos SOCS1fl. O aumento na susceptibilidade a sepse foi acompanhado pelo aumento da via glicolítica nas células peritoneias e no pulmão desses animais. Assim, foi observado aumento da produção de ácido láctico e da expressão de enzimas glicolíticas como hexoquinase-1 (Hk1), lactato desidrogenase A (Ldha) e o transportador de glicose 1 (Glut-1) em camundongos sépticos tratados com iKIR ou SOCS1?myel. A expressão desses genes da via glicolítica foi dependente da via de ativação STAT3/HIF-1?. O tratamento com 2-deoxiglicose (inibidor da via glicolítica) diminuiu a susceptibilidade à sepse em camundongos tratados com iKIR. Estes resultados indicam um papel até agora desconhecido de SOCS1, como um regulador de reprogramação metabólica que reduz a resposta inflamatória exacerbada e o dano de órgãos durante a sepse. / Sepsis is a life-threatening organ dysfunction caused by a dysregulated host response to infection. Identification of pleiotropic molecular brakes might provide therapeutic targets to prevent organ failure during sepsis. Suppressor of cytokine signaling 1 (SOCS1) is known to negatively regulate signaling by cytokine and Tolllike receptors (TLRs). However, the cellular targets and molecular mechanisms involved in SOCS1 actions during sepsis are unknown. To address this in a cecal ligation puncture (CLP) model of sepsis, we treated C57BL/6 mice with an antagonist peptide (iKIR) that blocks the kinase inhibitory region (KIR) domain of SOCS1 and prevents its actions. iKIR treatment increased mortality, bacterial burden and inflammatory cytokine production induced after CLP. We also found that myeloid cell-specific SOCS1 deletion (SOCS1?myel) rendered mice more susceptible to sepsis, shown by higher bacterial loads and inflammatory cytokines than SOCS1fl littermate control mice. O aumento na susceptibilidade a sepse foi acompanhado pelo aumento da via glicolítica nas células peritoneias e pulmão desses animais. These effects were accompanied by increase of glycolysis function in peritoneal cells and lung of SOCS1?myel. Thus, it was observed increased expression of the glycolytic enzymes, hexoquinase-1 (Hk1), lactate dehydrogenase A (Ldha), and glucose transporter 1 (Glut-1) in iKIR-treated or SOCS1?myel septic mice. These events were dependent on the activation of STAT3/HIF-1? pathway. Blocking glycolysis with 2-deoxyglucose ameliorated the increased susceptibility to sepsis in iKIR-treated CLP mice. Together, we unveiled a heretofore unknown role of SOCS1 as a regulator of metabolic reprograming that reduces overwhelming inflammatory response and organ damage during sepsis.

Introdução: Apesar dos recentes estudos, a sepse ainda é uma das mais importantes causas de hospitalização e morte nas Unidades de Terapia Intensiva (UTI). A equipe de enfermagem tem papel primordial no reconhecimento dos sinais e sintomas iniciais e acredita-se que a aferição dos sinais vitais de forma correta e completa, possa ser uma ferramenta simples, de baixo custo e não invasiva, para avaliar a predisposição a um desfecho de alta ou óbito nos pacientes sépticos em UTI. Objetivo: Avaliar a relação dos sinais vitais aferidos pela equipe enfermagem no momento do diagnóstico de sepse, com o desfecho clínico e com os scores MEWS e qSOFA. Métodos: Trata-se de um estudo com delineamento observacional, analítico que utilizou dados de fontes secundárias e o tratamento dos dados com abordagem metodológica quantitativa. Resultados: A média de idade dos pacientes foi de 56,4 anos e a maioria do sexo masculino, 53,4%. As doenças cardiovasculares ocuparam 43,1% das comorbidades existentes. O tempo médio de internação em UTI foi 8,0 dias e de internação hospitalar total de 29,6 dias e os pacientes sobreviventes permaneceram mais tempo internados tanto hospitalar quanto na UTI. A frequência cardíaca variou entre 38 e 162 bpm e foi observado que os pacientes que foram a óbito apresentavam-se mais taquicárdicos do que os sobreviventes. O desfecho de óbito ocupou mais de 70% dos casos analisados e após a aplicação dos escores qSOFA e MEWS não foi observado associação estatisticamente significativa entre eles e o desfecho (p=0,215). Conclusão: A aferição da frequência cardíaca no momento do diagnóstico de sepse pode ser utilizado como ferramenta efetiva, rápida, não invasiva e de baixo custo, para auxiliar no diagnóstico e predição de maior risco de óbito nos pacientes sépticos em UTI / Introduction: Despite recent studies, sepsis is still one of the most important causes of hospitalization and death in Intensive Care Units (ICUs). The nursing team has a primordial role in the recognition of the initial signs and symptoms and it is believed that the correct and complete measurement of the vital signs can be a simple, low-cost and non-invasive tool to evaluate the predisposition to an outcome discharge or death in septic patients in ICU. Objective: To evaluate the relationship of the vital signs measured by the nursing team at the moment of the diagnosis of sepsis, with the clinical outcome and the MEWS and qSOFA scores. Methods: This is a study with an observational, analytical design that used data from secondary sources and the treatment of the data with a quantitative methodological approach. Results: The mean age of the patients was 56.4 years and the majority of the males, 53.4%. Cardiovascular diseases accounted for 43.1% of the existing comorbidities. The mean ICU length of hospital stay was 8.0 days and total hospital stay was 29.6 days, and the surviving patients remained longer hospitalized both in the hospital and in the ICU. The heart rate varied between 38 and 162 bpm and it was observed that patients who died had more tachycardia than the survivors. The outcome of death was more than 70% of the cases analyzed and after the application of the qSOFA and MEWS scores, no statistically significant association between them and the outcome (p = 0.215) was observed. Conclusion: Heart rate measurement at the time of diagnosis of sepsis can be used as an effective, fast, non-invasive and low cost tool to assist in the diagnosis and prediction of a higher risk of death in septic patients in ICU

Dissertação de mestrado em Nutrição Clínica apresentada à Faculdade de Ciências da Nutrição e Alimentação da Universidade do Porto / Resumo da tese: A sépsis, considerada uma situação clínica complexa e um grave problema de saúde pública mundial, é uma causa comum de admissão no Serviço de Medicina Intensiva (SMI) e a principal causa de morte dos doentes críticos. O ângulo de fase (AF), determinado pela análise da Bioimpedância eléctrica (BIE) consiste numa medida directa da estabilidade das células e reflecte a contribuição de fluídos e membranas celulares do corpo humano, sendo interpretado como indicador de integridade da membrana e preditor de massa celular corporal. A aplicação do AF tem-se revelado de grande eficácia na aferição dos compartimentos corporais em diversas situações clínicas, nomeadamente em neoplasias, pré e pós-operatórios, traumatismos, doenças hepáticas, insuficiência renal, SIDA e sépsis. Nestas condições, tem sido sugerido como indicador de prognóstico e factor preditivo de sobrevivência. Alguns estudos clínicos demonstram que, baixos AF estão associados ao agravamento da doença e um pior prognóstico clínico, com consequente aumento da mortalidade em doentes críticos. Objectivos: i) Aferir a capacidade de prognóstico do AF, obtido por análise da BIE, em doentes críticos com sépsis, enquando indicador fiável e precoce de mortalidade; ii) Analisar a relação entre o AF e os índices clínicos (SAPS ll e SOFA) em doentes sépticos críticos, associando estes indicadores com a gravidade da sépsis; iii) Relacionar os valores do AF dos doentes internados no SMI, com o grau de falência orgânica, o tempo de internamento e a evolução clínica dos mesmos; iv) Avaliar a associação entre a AF e parâmetros bioquímicos, tradicionalmente utilizados na avaliação de risco nutricional, como a albumina plasmática e a proteína C-reactiva (PCR); v) Averiguar a sensibilidade e utilidade do AF para monitorizar alterações no estado nutricional, avaliando assim a efectividade da terapia nutricional (...). / Thesis abstract: Introduction: Sepsis is a complex clinical situation, a serious public health problem, a common cause of admission to the Unit of Intensive Care Unit (ICU) and the leading cause of death in critically ill patients. Phase angle (PA) determined by the Bioelectrical Impedance Analysis (BIA), is a direct measure of cell stability and reflects the human body contribution of fluid and cellular membranes, thus being interpreted as a membrane integrity indicator and body cell mass predictor. The PA application has been demonstrated large efficiency in body compartments gauging in several clinical settings, such as cancer, pre and post-operative trauma, liver disease, kidney failure, AIDS and sepsis. In these conditions, has been suggested as a prognostic indicator and predictor of survival. Some clínical studies show that low PA is associated with worsening disease and a worse clinical outcome with consequent increase in morbidity and mortality in critically ill patients. Objectives i) Assess the ability of phase angle prediction, determined by the BIA, in critical patients with sepsis, while reliable and early indicator of mortality; ii) Analyze the relationship between the phase angle and the clinical indicators (SAPS ll and SOFA) in critically patients, linking them with the severity of sepsis; iii) List the values of the PA of patients in the ICU, the degree of organ failure, length of stay and clinical outcome of same; iv) Evaluate the association between PA and biochemical parameters traditionally used in the assessment of nutritional risk, such as albumin and plasma C-reactive protein (CRP); v) Establishing the sensitivity and usefulness of the PA to monitor changes in nutritional status, thus evaluating the effectiveness of nutritional therapy(...).

A doença pneumocócica invasiva é um grande desafio global. Bacteremia está fortemente relacionada às condições clínicas de imunodepressão. Pacientes com câncer raramente são alvo de estudos clínicos. Neste estudo, nosso objetivo primário foi avaliar os fatores prognósticos em pacientes com câncer com bacteremia pneumocócica. Os objetivos secundários foram descrever os episódios de bacteremia e as características fenotípicas das cepas isoladas. O desenho foi uma coorte retrospectiva, que incluiu pacientes com câncer com mais de 18 anos de idade que tiveram bacteremia pneumocócica, de janeiro de 2009 a junho de 2015, no Instituto do Câncer do Estado de São Paulo. Os critérios de infecção foram aqueles delineados pelo NHSN/CDC. Os desfechos avaliados foram mortalidade em 48h e em 10 dias, e sobrevida dos pacientes. As variáveis independentes analisadas foram: idade, sexo, etnia, ECOG, Karnofsky escore, SOFA, diagnóstico do câncer, presença de metástases, quimioterapia, radioterapia, neutropenia, uso prévio de antibióticos, internação hospitalar prévia, internação prévia em UTI, comorbidades (DPOC sistêmica, Hipertensão Arterial, Diabetes Mellitus e Insuficiência Cardíaca), tabagismo, vacinação antipneumocócica prévia, sítio de infecção, infecção polimicrobiana, suscetibilidade antimicrobiana, sorotipo e tratamento. A associação das características com os desfechos foi verificada por meio dos testes qui-quadrado, teste de razão de verossimilhança ou teste exato de Fisher. A análise multivariada foi realizada por regressão logística stepwise, incluindo todas as variáveis com P < 0,10. A sobrevida foi estimada de acordo com cada característica avaliada pela função de Kaplan-Meier, seguida de testes log-rank para comparar a sobrevida entre as categorias das variáveis qualitativas. As Razões de Risco (HR) foram calculadas com os respectivos intervalos de confiança de 95% usando regressão de Cox bivariada. Todas as variáveis que apresentaram nível de significância de 0,10 (p < 0,1) foram testadas no modelo múltiplo, e as variáveis do modelo final foram selecionadas pelo método stepwise com critério de seleção Backward com entrada e saída de 5%. Os testes foram realizados com nível de significância de 5%. Foram detectadas 165 bacteremias pneumocócicas no período do estudo em 161 pacientes oncológicos; destas, 4 episódios foram excluídos por se tratarem de recidivas. Assim, 161 episódios foram incluídos para análise descritiva e estatística. A casuística foi composta em sua maioria por homens (62,7%), com uma média de idade de 61,3 anos. A maioria da casuística foi composta por indivíduos com tumores sólidos (121/75,2%). Entre os tumores sólidos, os mais prevalentes foram os do trato gastrointestinal, seguido dos tumores de cabeça e pescoço e trato respiratório. Entre as neoplasias hematológicas (40/24,8%), as mais prevalentes foram mieloma múltiplo (20/50,0%), seguidos de linfomas não Hodgkin (14/35,0%). A mortalidade em 48h e em 10 dias foi, respectivamente, de 20,5% e 33,5%. Entre 120 cepas testadas, 61 (50,8%) corresponderam aos encontrados na vacina VCV13 e 89 (74,2%) na VPV23. As variáveis associadas a mortalidade em 48h, na análise multivariada, foram: SOFA (OR 1,37, p < 0,001) bacteremia polimicrobiana (OR 3,78, p0,018). Na análise multivariada para mortalidade em 10 dias, os fatores associados foram: febre 48h antes da bacteremia (OR 0,35, p0,026), neutropenia vigente (OR 4,01, p0,011), ECOG ¾ (OR 5,83, p0,001), SOFA (OR 1,41, P < 0,001 - um aumento unitário do SOFA resultou em um aumento de 59% na chance de mortalidade) e uso de quinolonas (OR 0,08, p0,023). Na sobrevida global, as variáveis relacionadas detectadas foram: presença de metástases (HR 1,97, p < 0,001), KARNOFSKY < 90 (HR 1,56, p0,021), SOFA (HR 1,27, p < 0,001), uso de anticorpos no último mês (HR 3,25, p0,026) e bacteremia polimicrobiana (HR 1,69, p0,046). Em conclusão, a bacteremia pneumocócica apresentou alta mortalidade em pacientes oncológicos, com prognóstico relacionado a fatores intrínsecos do hospedeiro e aos episódios de infecção, como o SOFA presente como preditor de mortalidade em 48h, 10 dias e piora da sobrevida global. A presença de bacteremia polimicrobiana associou-se a mortalidade em 48h e piorou a sobrevida. Presença de febre e uso de quinolonas foram fatores de proteção para a mortalidade em 10 dias; a presença de febre provavelmente encurtou o tempo entre o início da sepse e a demanda por serviços de saúde, com antibioticoterapia mais precoce. Não foi possível estimar o efeito protetor da vacinação antipneumocócica devido ao pequeno número de pacientes vacinados / Invasive pneumococcal disease is a major global challenge. Bacteremia is strongly related to the clinical conditions of immunodepression. Patients with cancer are rarely the target of clinical trials. In this study, our primary objective was to evaluate the prognostic factors in patients with pneumococcal bacteremia. The secondary objectives were to describe the episodes of bacteremia and the phenotypic characteristics of the isolated strains. The design was a retrospective cohort that included cancer patients over 18 years of age who had pneumococcal bacteremia from January 2009 to June 2015 at the \"Instituto do Câncer do Estado de São Paulo\". The criteria for infection were those outlined by the NHSN / CDC. The outcomes evaluated were mortality in 48h and 10 days and overall survival. The independent variables analyzed were: age, sex, ethnicity, ECOG, Karnofsky score, SOFA, diagnosis of cancer, presence of metastasis, chemotherapy, radiotherapy, neutropenia, previous use of antibiotics, previous hospital admission, previous ICU admission, comorbidities (systemic COPD, Hypertension, Diabetes mellitus and Congestive Heart Failure), smoking, previous antipneumococcal vaccination, site of infection, polymicrobial infection, antimicrobial susceptibility, serotype and treatment. The association of the characteristics with the outcomes was verified using chi-square tests, likelihood ratio test or Fisher\'s exact test. Multivariate analysis was performed by stepwise logistic regression, including all variables with P < 0.10. Survival was estimated according to each characteristic assessed by the Kaplan-Meier function, followed by log-rank tests to compare survival between categories of qualitative variables. The Hazard Ratios (HR) were calculated with the respective 95% confidence intervals using bivariate Cox regression. All variables that presented a significance level of 0.10 (p < 0.1) were tested in the multiple model, and the variables of the final model were selected by stepwise method with Backward selection criterion with input and output of 5%. The tests were performed with a significance level of 5%. We detected 165 pneumococcal bacteremias during the study period in 161 cancer patients; of these, 4 episodes were excluded because they were recurrences. Thus, 161 episodes were included for descriptive and statistical analysis. Our study consisted mostly of men (62.7%), with a mean age of 61.3 years. The majority of cases were individuals with solid tumors (121 / 75.2%). Among the solid tumors, the most prevalent tumors were those of the gastrointestinal tract, followed by tumors of the head and neck and respiratory tract. Among the hematological malignancies (40/24.8%), the most prevalent were multiple myeloma (20 / 50.0%), followed by non-Hodgkin\'s lymphomas (14 / 35.0%). Mortality in 48h and 10 days was, respectively, 20.5% and 33.5%. Among 120 strains tested, 61 (50.8%) corresponded to those found in the VCV13 vaccine and 89 (74.2%) in the VPV23. The variables associated with 48-h mortality in the multivariate analysis were: SOFA (OR 1.37, p < 0.001) and polymicrobial bacteremia (OR 3.78, p 0.018). In the multivariate analysis for 10-day mortality, the associated factors were: fever 48h before bacteremia (OR 0.35, p 0.026), current neutropenia (OR 4.01, p 0.011), ECOG ¾ (OR 5.83 , p 0.001), SOFA (OR 1.41, P < 0.001 - a unit increase of SOFA resulted in a 59% increase in chance of mortality) and use of quinolones (OR 0.08, p0.023). In the overall survival, the related variables detected were: presence of metastases (HR 1.97, p < 0.001), KARNOFSKY < 90 (HR 1.56, p0.021), SOFA (HR 1.27, p < 0.001), use of antibodies in the last month (HR 3.25, p0.026) and polymicrobial bacteremia (HR 1.69, p0.046). In conclusion, pneumococcal bacteremia presented high mortality in cancer patients, with a prognosis related to intrinsic host factors and infection episodes, such as SOFA present as a predictor of mortality in 48h, 10 days and worsening of overall survival. The presence of polymicrobial bacteremia was associated with 48-h mortality and worsened survival. Presence of fever and use of quinolones were protective factors for mortality in 10 days; the presence of fever probably shortened the time between the onset of sepsis and the demand for health services, with earlier antibiotic therapy. It was not possible to estimate the protective effect of anti-pneumococcal vaccination due to the small number of patients vaccinated

In under a century, melioidosis, the infection due to Burkholderia pseudomallei, has emerged from Whitmore’s series of glanders-like infections amongst the morphia addicts in Burma to a major cause of mortality in northeastern Thailand and northern Australia. Also endemic in other parts of south-east Asia, melioidosis may have varied presentations ranging from severe, overwhelming infection to chronic, low grade disease. Observational evidence had suggested that granulocyte colony stimulating factor (G-CSF), a naturally occurring substance produced by the body in response to infection, may have been useful in reducing the high mortality associated with the more severe forms of this infection. Other observations linked the occurrence of this disease to various environmental factors, such as contamination of drinking water and the annual rainfall. This thesis explores and attempts to quantify these associations. There are three parts to this thesis. In the first part, I reviewed the epidemiology and management of patients with melioidosis. The use of G-CSF and meropenem was associated with a fall in mortality, although other factors may have at least partially contributed to this effect. In the second part, I progressed towards a clinical trial of G-CSF. There was no other evidence supporting the use of G-CSF in severe sepsis and ethical issues precluded a trial in Darwin. There was not evidence from laboratory models of G-CSF action in melioidosis to support the use of G-CSF in patients, although there remained some doubt regarding the applicability of such models to human disease. I examined clinical methods to identify patients at high risk of death from melioidosis. A simple scoring system based on clinical and laboratory parameters was developed and externally validated. However, clinical definitions of severe sepsis appeared to be better predictors of mortality. A clinical trial based on clinical definitions was commenced in Thailand. In the final part, I explored the question of whether different strains or B. pseudomallei or different environmental conditions caused different patterns of infection. There was no evidence that strain types of this bacterium determine the pattern or severity of disease, but weather conditions appeared to influence the distribution of disease in northern Australia.

Doctor of Philosophy / Department of Industrial & Manufacturing Systems Engineering / David H. Ben-Arieh / Chih-Hang Wu / Patients who suffer from sepsis or septic shock are of great concern in the healthcare system. Recent data indicate that more than 900,000 severe sepsis or septic shock cases developed in the United States with mortality rates between 20% and 80%. In the United States alone, almost $17 billion is spent each year for the treatment of patients with sepsis. Clinical trials of treatments for sepsis have been extensively studied in the last 30 years, but there is no general agreement of the effectiveness of the proposed treatments for sepsis. Therefore, it is necessary to find accurate and effective tools that can help physicians predict the progression of disease in a patient-specific way, and then provide physicians recommendation on the treatment of sepsis to lower risk for patients dying from sepsis. The goal of this research is to develop a risk assessment tool and a risk management tool for sepsis. In order to achieve this goal, two system dynamic mathematical models (SDMMs) are initially developed to predict dynamic patterns of sepsis progression in innate immunity and adaptive immunity. The two SDMMs are able to identify key indicators and key processes of inflammatory responses to an infection, and a sepsis progression. Second, an integrated-mathematical-multi-agent-based model (IMMABM) is developed to capture the stochastic nature embedded in the development of inflammatory responses to a sepsis. Unlike existing agent-based models, this agent-based model is enhanced by incorporating developed SDMMs and extensive experimental data. With the risk assessment tools, a Markov decision process (MDP) is proposed, as a risk management tool, to apply to clinical decision-makings on sepsis. With extensive computational studies, the major contributions of this research are to firstly develop risk assessment tools to identify the risk of sepsis development during the immune system responding to an infection, and secondly propose a decision-making framework to manage the risk of infected individuals dying from sepsis. The methodology and modeling framework used in this dissertation can be expanded to other disease situations and treatment applications, and have a broad impact to the research area related to computational modeling, biology, medical decision-making, and industrial engineering.

Η σήψη αποτελεί μια από τις σημαντικότερες αιτίες νοσηλείας και θνησιμότητας στον ανεπτυγμένο κόσμο, όπου σχεδόν τα δύο - τρίτα του πληθυσμού υποφέρουν από παχυσαρκία. Σαν αποτέλεσμα, η συνύπαρξη των δύο αυτών καταστάσεων έχει γίνει όλο και συχνότερη στην κλινική πράξη και ένας συνεχώς αυξανόμενος αριθμός κλινικών μελετών προσπαθεί να προσεγγίσει την πιθανή επίδραση της παχυσαρκίας στην νοσηρότητα και θνησιμότητα των ασθενών με σήψη, με έως τώρα αντιφατικά αποτελέσματα. Σκοπός της παρούσας μελέτης είναι να διερευνήσει τον τρόπο με τον οποίο η παχυσαρκία επηρεάζει την ανοσιακή απάντηση των σηπτικών ασθενών, εκτιμώντας τον αριθμό και την κατάσταση ενεργοποίησης των μακροφάγων του λιπώδους ιστού, τα επίπεδα του TNFα στον ορό και στον λιπώδη ιστό και δείκτες οξειδωτικού stress στο πλάσμα. Ασθενείς και Μέθοδοι: Μελετήθηκαν 106 ασθενείς, οι οποίοι χωρίστηκαν σε τέσσερις ομάδες (Ελέγχου n=26, Παχυσαρκίας n=27, Σήψης n=27, Σήψης & Παχυσαρκίας n=26). Ο αριθμός των μακροφάγων στο υποδόριο και ενδοκοιλιακό λίπος και οι υπότυποί τους (M1 και M2) αναγνωρίστηκαν με ανοσοϊστοχημική τεχνική υπό μικροσκόπηση. Τα επίπεδα του TNFα mRNA στο υποδόριο και ενδοκοιλιακό λίπος μετρήθηκαν με real-time reverse transcription-PCR. Στον ορό τα επίπεδα του TNFα μετρήθηκαν με sandwich enzyme-linked immunosorbent assay (ELISA). Το οξειδωτικό stress στο πλάσμα εκτιμήθηκε χρησιμοποιώντας επιλεγμένους βιοδείκτες [TBARS (thiobarbituric acid-reactive substances), Πρωτεϊνικά Καρβονύλια, TAC (total antioxidant capacity)]. Αποτελέσματα: Παρατηρήθηκε ότι η σήψη αυξάνει τον ολικό αριθμό και τον Μ2 υπότυπο των μακροφάγων στο ενδοκοιλιακό λίπος, ενώ η παχυσαρκία δεν φάνηκε να επηρεάζει τη συγκέντρωση των μακροφάγων στο λίπος. Η παχυσαρκία βρέθηκε ότι αυξάνει τα επίπεδα του TNFα mRNA (P<0.05) στο ενδοκοιλιακό λίπος καθώς επίσης και τα επίπεδα των TBARS (P<0.001) και Πρωτεϊνικών Καρβονυλίων (P<0.001) στο πλάσμα των σηπτικών ασθενών. Τα επίπεδα της TAC στο πλάσμα βρέθηκε ότι μειώνονται και τα επίπεδα TNFα στον ορό ότι αυξάνονται με τη σήψη, ενώ δεν επηρεάζονταν από την παχυσαρκία. Συμπεράσματα: Η παχυσαρκία σχετίζεται με αυξημένη παραγωγή TNFα στον λιπώδη ιστό και αύξηση του οξειδωτικού stress, προάγοντας την προ-φλεγμονώδη απάντηση στους σηπτικούς ασθενείς. / Sepsis is one of the most important causes of mortality in the developed world, where almost two thirds of the population suffer from obesity. Therefore, the coexistence of both conditions has become frequent in clinical practice and a growing number of clinical studies attempts to examine the potential effect of obesity on sepsis with controversial results up to now. The present study investigates how obesity influences the immune response of septic patients, by assessing the number and activation state of adipose tissue macrophages, serum and adipose tissue tumor necrosis factor-alpha (TNFα) levels and plasma oxidative stress markers. Subjects/methods: The study included 106 patients, divided into four groups (control n = 26, obesity n = 27, sepsis n = 27 and sepsis and obesity n = 26). The number of macrophages in subcutaneous and visceral adipose tissue (SAT and VAT) and their subtypes (M1 and M2) were defined with immunohistochemical staining techniques under light microscopy. TNFα mRNA levels were determined in SAT and VAT using real-time reverse transcription-PCR. Serum levels of TNFα were determined with sandwich enzyme-linked immunosorbent assay. Plasma oxidative stress was evaluated using selective biomarkers [thiobarbituric acid-reactive substances (TBARS), protein carbonyls and total antioxidant capacity (TAC)]. Results: Sepsis increased the total number of macrophages and their M2 subtype in VAT, whereas obesity did not seem to affect the concentration of macrophages in fat. Obesity increased TNFα mRNA levels (P < 0.05) in VAT as well as the plasma TBARS (P < 0.001) and protein carbonyls (P < 0.001) in septic patients. The plasma TAC levels were decreased and the serum TNFα levels were increased in sepsis although they were not influenced by obesity. Conclusions: Obesity is associated with elevated TNFα adipose tissue production and increased oxidative stress biomarkers, promoting the proinflammatory response in septic patients.

<p>Nitric oxide (NO) is an endogenous gaseous signaling molecule produced by three NO synthase isoforms (NOS1, 2, 3) and important in host defense. The induction of NOS2 during bacterial sepsis is critical for pathogen clearance but its sustained activation has long been associated with increased mortality secondary to multiple organ dysfunction syndrome (MODS). High levels of NO produced by NOS2 incite intrinsic cellular dysfunction, in part by damaging macromolecules through nitration and/or nitrosylation. These include mitochondrial DNA (mtDNA) and enzymes of key mitochondrial pathways required for maintenance of normal O2 utilization and energy homeostasis. However, animal studies and clinical trials inhibiting NOS2 have demonstrated pronounced organ dysfunction and increased mortality in response to live bacterial infections, confirming that NOS2 confers pro-survival benefits. Of particular interest here, the constitutive NOS1 and NOS3 have been linked to the up-regulation of nuclear genes involved in mitochondrial biogenesis but no comparable role has been described for NOS2. <italic> Therefore, I hypothesized that NOS2 is indispensible for host protection but must be tightly regulated to ensure NO levels are high enough to activate mitochondrial and other pro-survival genes, but below the threshold for cellular damage.</italic></p><p>This hypothesis was explored with two major Aims. The <italic>first Aim</italic> was to define the role of NOS2 in the activation of mitochondrial biogenesis in the heart of <italic>E. coli</italic>-treated mice. The <italic>second</italic> was to investigate the ability of NOS2 to be transcriptionally regulated by an enzyme previously shown to induce mitochondrial biogenesis, heme oxygenase-1 (HO-1). This hypothesis was tested using an <italic>in vivo</italic> model of sublethal heat-killed <italic>E. coli</italic> (<italic>HkEC</italic>) peritonitis in C57B/L6 (Wt), NOS2-/-, and TLR4-/- mice. Additionally, <italic>in vitro</italic> systems of mouse AML-12 or Hepa 1-6 cells pretreated with HO-1 activators or <italic>Hmox1</italic> shRNA prior to inflammatory challenge with lipopolysaccharide (LPS) +/- tumor necrosis factor-&alpha; (TNF-&alpha;). For the first Aim, Wt, NOS2-/-, and TLR4-/- mice were treated with (<italic>HkEC</italic> and cardiac tissue analyzed for mitochondrial function, expression of nuclear and mitochondrial proteins needed for mitochondrial biogenesis, and histological expression of NOS2 and TLR4 relative to changes in mitochondrial mass. For the second Aim, Wt mice were pretreated with hemin or carbon monoxide (CO) to activate HO-1 prior to <italic>HkEC</italic>-peritonitis. Liver tissue in these animals was evaluated at four hours for HO-1 induction, <italic>Nos2</italic> mRNA expression, cytokine profiles, and nuclear factor (NF)-&kappa;B activation. Liver cell lines were pretreated with hemin, CO-releasing molecule (CORM), or bilirubin one hour before LPS exposure and the <italic>Nos2</italic> transcriptional response evaluated at two and 24 hours. The MTT assay was used to confirm that <italic>in vitro</italic> treatments were not lethal. </p><p>These studies demonstrated that <italic>HkEC</italic> induced mtDNA damage in the heart that was repaired in Wt mice but not in NOS2-deficient mice. In KO mice, sustained mtDNA damage was associated with the reduced expression of nuclear (NRF-1, PGC-1&alpha;) and mitochondrial (Tfam, Pol-&gamma;) proteins needed for mitochondrial biogenesis. The findings thus supported that NOS2 is required for mitochondrial biogenesis in the heart during Gram-negative challenge. Evaluation of the relationship between HO-1 and NOS2 in murine liver was more complex; HO-1 activation in <italic>HkEC</italic>-treated Wt mice attenuated 4-hour <italic>Nos2</italic> gene transcription. In liver cell lines, hemin, CORM, and bilirubin were unable to suppress <italic>Nos2</italic> expression at the time of maximal induction (2 hours). <italic>Nos2</italic> was, however, suppressed by 24 hours, suggesting that the regulatory impact of HO-1 induction was not engaged early enough to reduce <italic>Nos2</italic> transcription at 2 hours. It is concluded that NOS2 induction in bacterial sepsis optimizes the expression of the mitochondrial biogenesis transcriptional program, which subsequently can also be regulated by HO-1/CO in murine liver. This provides a potential new mechanism by which immune suppression and mitochondrial repair can occur in tandem during the acute inflammatory response.</p> / Dissertation

Hydroxyethyl starches (HES) are the most routinely used synthetic colloids during fluid resuscitation and have reported effects on coagulation. The overall goal of the investigation in this thesis was to evaluate the effects of tetrastarch administration on hemodynamic, laboratory, and hemostatic variables in healthy dogs and dogs with systemic inflammation. The objectives were to compare hemodynamic and laboratory variables in dogs receiving an isotonic crystalloid (0.9% NaCl) or tetrastarch during health and after induction of systemic inflammation; to compare the hemostatic effects of an isotonic crystalloid (0.9% NaCl) and synthetic colloid (tetrastarch) in healthy dogs and dogs with induced systemic inflammation; to compare two different protocols for TEG® activation and to determine the correlation between TEG® variables and traditional coagulation test results. Sixteen adult purpose-bred Beagles were randomized into one of two groups receiving fluid resuscitation with either 40 mL/kg IV isotonic crystalloid (0.9% NaCl) or synthetic colloid (tetrastarch) after administration of lipopolysaccharide (LPS; 5 μg/kg, IV) or an equal volume of placebo (0.9% NaCl, IV). Blood samples, for analysis, were collected at 0, 1, 2, 4, and 24 hours from the time of fluid resuscitation. After a 14-day washout period, the study was repeated such that dogs received the opposite treatment (LPS or placebo) and the same resuscitation fluid. Resuscitation with equal volumes of 0.9% NaCl and tetrastarch caused similar changes in hemodynamic and laboratory variables in dogs with LPS-induced systemic inflammation; however, larger increases in HR and blood pressure were seen within the first 2 hours following tetrastarch administration compared to 0.9% NaCl. Tetrastarch administration increased COP in all dogs, despite a decrease in TS. Tetrastarch bolus administration to dogs with LPS-induced systemic inflammation also resulted in a transient hypocoagulability characterized by a prolonged PTT, decreased clot formation speed and clot strength, and acquired type 1 von Willebrand disease. Considering the limited additional benefit of tetrastarch administration on hemodynamic variables demonstrated, as well as the transient adverse hemostatic effects of tetrastarch administration, the increased cost associated with the use of tetrastarch likely negates its use as a first line treatment during fluid resuscitation in dogs. / Pet Trust Fund