Sepse e choque séptico são causas frequentes de morte nas unidades de terapia intensiva, sendo responsável pelo alto custo global de internação, segundo DATA-SUS. Sabendo-se que o paciente diabético apresenta um maior risco de infecção e que 22% dos indivíduos com sepse apresentam diabetes, é de extrema importância o melhor entendimento da sua fisiopatologia no paciente diabético para que medidas intervencionistas sejam desenvolvidas, preservando vidas. O projeto buscou avaliar a sepse experimentalmente através do modelo de ligadura e perfuração do ceco (CLP - 2 perfurações) em ratos Wistar machos sadios e tornados diabéticos através da injeção endovenosa de aloxana (42 mg/kg, i.v., 10 dias). Além disso, buscamos avaliar se o tratamento com insulina alteraria as variáveis escolhidas. O trabalho avaliou parâmetros hematológicos e bioquímicos como uréia, creatinina, alanina aminotransferase (ALT), aspartato aminotransferase (AST) e fosfatase alcalina (FAL). Após 6 horas da realização da CLP foram coletados lavados broncoalveolares (LBA) e peritoneal (LPe) com o objetivo de estudar o perfil de citocinas através da dosagem das interleucinas (IL)-1β, IL-6, IL-10, fator de necrose tumoral (TNF)-α e cytokine-induced neutrophill chemoattractant (CINC)-1 e CINC-2, celularidade total e específica. Rim, pulmão e fígado foram coletados para análises morfológicas e da atividade da mieloperoxidase (MPO). Após 6 horas de CLP observamos que a hematimetria, hemoglobina, plaquetometria e celularidade do LBA não sofreram alterações nem após o tratamento com insulina. A sepse provocou diminuição da leucometria total nos animais diabéticos e controles, aumento da celularidade total do LPe com um predomínio de polimorfonucleares, que foi semelhante em ambos os grupos; aumento da concentração no LPe de IL-1β, IL-6, CINC-1, CINC-2 e IL-10; as concentrações do TNF-α permaneceram idênticas em ambos os grupos. Em relação aos marcadores hepáticos, foram observados que os animais diabéticos possuem os maiores valores de ALT, AST e FAL em relação ao grupo controle e já apresentavam uma disfunção morfológica de hepatócitos, porém, sem infiltrado neutrofílico. O tratamento com insulina reduziu os valores dessas enzimas em níveis próximos aos do controle. Dos marcadores renais, apenas a uréia sofreu aumento significativo nos animais diabéticos em relação aos controles, sendo exacerbada pela CLP. Alterações glomerulares, como capilares dilatados e redução do espaço de Bowman foram observadas nos animais com sepse, os quais não sofreram influência após o tratamento com insulina. Estes dados sugerem que a insulina teve um efeito hepato-protetor e que o padrão causado pela sepse nos animais controles e diabéticos não foi modificado pelo tratamento com insulina. / Sepsis and septic shock are common cause of death in intensive care units (ICU) and according to Brazilian DATA-SUS is one of the major causes of high cost hospitalization. Since diabetic patients have a higher risk of infection and represent approximately 22% of all the septic individuals, it is extremely important to understand the pathophysiology of sepsis in diabetic patients in order to develop interventional measure. The objective of this study is to evaluate the experimental sepsis model by cecal ligation and puncture (CLP 2 punctures) in Wistar healthy males rendered diabetic by intravenous injection of alloxan (42 mg/kg, i.v., 10 days). The study analyzed hematological and biochemical parameters such as urea, creatinine, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP). Furthermore, we studied how insulin treatment could modulate these parameters. After 6 hours of CLP bronchoalveolar (BAL) and peritoneal lavages (PeL) were collected and a cytokine profile was accessed: interleukin (IL)-1β, IL-6, IL-10, tumor necrosis factor (TNF)-α, cytokine-induced neutrophill chemoattractant (CINC)-1, CINC-2. Samples from kidney, lung and liver were collected for morphological analysis and for the measurement of myeloperoxidase activity (MPO). After 6 hours of CLP, red blood cell count, hemoglobin, platelet count, cytokine profile and cellularity of the BAL did not change. In addition, insulin treatment did not change these parameters. CLP caused a decrease in total leukocyte count in both groups diabetic and control rats. Moreover, this model induced a rise in total PeL cellularity with a predominance of polymorfonuclear cells, which was similar in both groups control and diabetic rats. In the PeL, proinflammatory cytokines such as IL-1β, IL-6, CINC-1, CINC-2 and the antinflammatory cytokine IL-10 were enhanced. The TNF-α concentrations remained similar in both groups. Hepatic markers inferring in hepatocyte dysfunction were higher in the diabetes-induced animals what was in part restored after insulin administration. Regarding renal markers, only urea levels were enhanced in diabetic rats and worsened after CLP induction. Morphological changes, such as capillary dilatation and reduced Bowman space was already observed in the kidney of animals with sepsis, insulin treatment did not corrected the values. These data suggest that insulin had a hepato-protective effect but further changes were not observed after insulin treatment.

A sepse é responsável por uma alta taxa de internação hospitalar e morbimortalidade. Devido à gravidade do quadro clínico e às limitações dos métodos tradicionais para identificação e isolamento bacteriano, é recomendado o início empírico de antimicrobiano(s) de largo espectro. O desenvolvimento da biologia molecular, particularmente da reação em cadeia da polimerase (PCR), tornou possível o diagnóstico rápido de agentes infecciosos. Entretanto, devido à diversidade dos possíveis agentes etiológicos na sepse, a utilização da PCR com primers específicos para cada agente se torna pouco prática. Com a PCR de largo espectro é possível que em uma só reação se identifique qualquer bactéria, possibilitando um tratamento precoce e direcionado. Desta forma, este trabalho teve como objetivo avaliar o papel da PCR de largo espectro no diagnóstico etiológico de pacientes com sepse e a comparação desta técnica com os métodos tradicionais de cultura. Foram incluídos 74 pacientes com diagnóstico de sepse atendidos na Unidade de Emergência do Hospital das Clínicas da Faculdade de Medicina de Ribeirão Preto USP e 14 voluntários sadios. Foi realizada a extração do DNA do soro, plasma e buffy-coat dos pacientes, seguida da realização da PCR de largo espectro com dois diferentes pares de primers, e sequenciamento das amostras positivas. Dos 74 pacientes, 39 (53%) eram homens; a média de idade foi 55 ± 19 anos; 37 (50%) tiveram sepse grave e 37 (50%) choque séptico; e a mortalidade foi 51%. A maioria das infecções primárias teve origem respiratória (66%), seguida de infecções gênito-urinárias (20%). A hemocultura foi positiva em 22 (30%) pacientes, e sua positividade foi significativamente maior em pacientes mais velhos (p< 0,05) e com valores mais altos de proteína C reativa (CRP) (p< 0,05). A PCR de largo espectro foi positiva em 44 (59%) pacientes considerando os dois pares de primers, sendo sua positividade significativamente maior que a da hemocultura (p< 0,001). Para o par Bak11W/Bak2 ela foi positiva em 25 (34%) pacientes, e para o par Taf/Tar, em 29 (39%) pacientes. Em relação às frações do sangue, amostras de 24 pacientes foram positivas na fração soro; 22 na fração plasma; e 18 na fração buffy-coat. Nenhuma característica clínica ou demográfica dos pacientes influenciou a positividade da PCR de largo espectro. A PCR de largo espectro foi negativa em todas as frações do sangue dos voluntários sadios. A sensibilidade, especificidade, valor preditivo positivo e valor preditivo negativo da PCR foram 59%, 100%, 100% e 32%, respectivamente. Em 40 (54%) pacientes os resultados da PCR e hemocultura foram concordantes. O coeficiente de concordância kappa obtido foi 0,147 (p = 0,131). Em relação ao sequenciamento, em 21 amostras de 16 pacientes foi possível identificar um agente etiológico. As bactérias mais detectadas foram Escherichia coli (3), Enterococcus sp. (2), Staphylococcus sp. (2) e Ralstonia sp. (2). Em apenas dois pacientes as amostras tiveram a mesma espécie bacteriana detectada na hemocultura e PCR de largo espectro (E. coli e Streptococcus pneumoniae). Em resumo, a PCR de largo espectro foi mais sensível do que a hemocultura na identificação bacteriana em pacientes com sepse atendidos em um Hospital de Emergência / Sepsis is responsible for a high rate of hospitalization and mortality. Due to the severity of clinical presentation and limitation of traditional methods for bacterial identification and isolation, it is recommended to initiate empirically broad-spectrum antimicrobial treatment. The development of molecular biology, particularly the polymerase chain reaction (PCR), enabled to realize a rapid diagnosis of infectious agents. However, due to the diversity of possible etiologic agents in sepsis, the use of PCR with specific primers for each agent becomes impractical. With the broad-range PCR, it is possible, in a single reaction, to identify any bacteria, allowing an early and directed treatment. Thus, this study aimed to evaluate the role of broad-range PCR in the etiologic diagnosis of patients with sepsis and compare this technique with traditional methods of culture. Seventy-four patients with sepsis admitted to the Emergency Unit of Clinical Hospital of Ribeirão Preto Medical School USP and 14 controls were included in the study. DNA from serum, plasma and buffy-coat were extracted from all patients and controls. Broad-range PCR was performed in all samples, followed by DNA sequencing of the amplicons. Out of 74 patients, 39 (53%) were male, mean age was 55 ± 19 years old; 37 (50%) patients had severe sepsis and 37 (50%) septic shock; the mortality rate was 51%. Most of primary infections were from respiratory tract (66%), followed by urinary tract infection (20%). Blood culture was positive in 22 (30%) patients, and its positivity was greater in older patients (p< 0,05) and patients with higher levels of C reactive protein (CRP) (p< 0,05). Broad-range PCR was positive in 44 (59%) patients, when considering both pairs of primers, and was significantly increased compared to blood culture positivity (p< 0,001). Broad-range PCR using Bak11W/Bak2 primers was positive in 25 (34%) patients, and using Taf/Tar primers, in 29 (39%) patients. Related to blood fractions, samples from 24 patients were positive in serum; 22 in plasma fraction; and 18 in buffy-coat. None of clinical and demographic characteristics influenced the broad-range PCR positivity. The sensitivity, specificity, positive predicted value and negative predictive value, when considered healthy persons as negative control, were 59%, 100%, 100% and 32%, respectively. In 40 (54%) patients, blood culture and PCR results were concordant. The concordance coefficient kappa obtained was 0,147 (p = 0,131). Regarding to etiologic agents, in 21 samples, from 16 patients, a bacteria was identified by sequencing. The most common bacteria were Escherichia coli (3), Enterococcus sp. (2), Staphylococcus sp. (2) and Ralstonia sp. (2). In only two patients, the same bacterial species were identified in both, blood culture and broad-range PCR (E. coli e Streptococcus pneumoniae). In conclusion, broad-range PCR was more sensitive than blood culture for bacterial identification in septic patients admitted to an Emergency Unit

O sistema nervoso central (SNC) é uma das primeiras regiões a ser acometida durante a sepse e choque séptico, o que contribui para o aumento da taxa de morbidade e mortalidade. Pacientes em choque séptico apresentam disfunção neuronal aguda, tais como o delírio, desorientação e coma. Em longo prazo, o dano cognitivo pode ocorrer ocasionando o comprometimento do aprendizado e formação de memória. Estudos demonstram que durante a resposta inflamatória sistêmica exacerbada, mediadores inflamatórios presentes na circulação sistêmica, são capazes de chegar ao SNC e ocasionar a ativação de células gliais, conduzindo a um estado de neuroinflamação. Nesse processo, algumas estruturas do SNC, tais como o hipocampo são mais vulneráveis à ação de espécies reativas de oxigênio (ERO), e a mediadores inflamatórios produzidos de forma excessiva durante a sepse. Neste contexto, a investigação de novas estratégias terapêuticas que sejam capazes de atenuar a resposta inflamatória exacerbada se faz necessário. Assim, o presente projeto teve como objetivo investigar prováveis propriedades antioxidante e anti-inflamatória do Hidrogênio molecular (H2), bem como sua possível ação neuroprotetora em ratos submetidos à sepse polimicrobiana, induzida por ligadura e perfuração cecal (CLP). Para isso o projeto foi dividido em dois protocolos experimentais. No primeiro protocolo ratos Wistar submetidos à cirurgia de CLP ou Sham, foram submetidos ao tratamento com inalação do H2 a 2%, por um período de 1h durante 10 dias consecutivos, e logo após foram submetidos a testes comportamentais para avaliação da memória de habituação, discriminativa e aversiva. No segundo protocolo os animais foram tratados com inalação do H2 por um período de 3h, e 24h após ao término da cirurgia de CLP/Sham foram decapitados para coleta do sangue e cérebro. A partir dos resultados dos testes comportamentais observamos que o tratamento com inalação do H2 durante a sepse experimental preveniu a perda de memória e o dano cognitivo, bem como foi capaz de diminuir os níveis de citocinas pró-inflamatórias de fase aguda tais como IL-1?, IL-6 e TNF? no córtex pré-frontal e hipocampo. A estratégia também foi capaz de diminuir os níveis de TBARS no plasma. Observamos um aumento da concentração da enzima catalase nos animais tratados com H2. Em conjunto os resultados indicam que o H2 foi capaz de inibir a resposta inflamatória e prevenir o dano cognitivo, agindo como uma substância neuroprotetora em ratos submetidos ao choque séptico experimental / The central nervous system is one of the first regions to be affected during Sepsis and septic shock, which contributes to the increased rate of morbidity and mortality. Patients with severe sepsis may present acute neuronal dysfunction such as delirium, disorientation, and unconscious. In the long term, cognitive damage can occur causing the commitment of learning and memory formation. Studies show that during the exacerbated systemic inflammatory response, inflammatory mediators present in the systemic circulation, are able to reach the CNS and cause the activation of glial cells, leading to a state of neuroinflammation. In this process, some CNS structures such as the hippocampus are more vulnerable to the action of reactive oxygen species (ROS), and to inflammatory mediators produced excessively during sepsis. In this context, the investigation of new therapeutic strategies that are capable of attenuating the exacerbated inflammatory response is necessary. Thus, the present project aimed to investigate the probable antioxidant and anti-inflammatory properties of molecular hydrogen (H2), as well as its possible neuroprotective action in rats submitted to polymicrobial sepsis induced by ligature and cecal puncture (CLP). In order to check this hypothesis, the project was divided into two experimental protocols. In the first protocol Wistar rats submitted to CLP or Sham surgery were submitted to 2% H2 inhalation treatment for a period of 1h for 10 consecutive days, and soon after they underwent behavioral tests to evaluate habituation memory, discriminative and aversive. In the second protocol the animals were treated with H2 inhalation for a period of 3h and 24h, and at the end of the treatment, they were decapitated for blood and brain collection. Plasma was already used for nitrate dosage, lipid peroxidation, antioxidant enzymes and inflammatory cytokines. From the results of the behavioral tests, we observed that treatment with H2 inhalation during the experimental sepsis prevented memory loss and cognitive damage, and was able to decrease the levels of acute-phase inflammatory cytokines such as IL-1?, IL -6 and TNF? in the prefrontal cortex and hippocampus. The therapeutic strategy was also able to decrease plasma TBARS levels. We also observed an increase in the concentration of the enzyme catalase in H2-treated animals. Together the results indicate that molecular hydrogen was able to inhibit the inflammatory response and prevent cognitive damage, acting as a neuroprotective substance, in rats submitted to experimental septic shock

Traumatic brain injury (TBI) is a major cause of death among trauma patients and is associated with a high rate of mortality due to complications such as bacterial pneumonia, sepsis, and subsequent coagulopathies. While severe TBI is positively associated with the development of pneumonia, mild traumatic brain injury (mTBI) results in a paradoxical decrease in mortality following bacterial pneumonia via an unidentified mechanism. New evidence suggests that mTBI stimulates vagus nerve signaling resulting in an anti-inflammatory state that is mediated by neurotransmitters (NT) such as acetylcholine (ACh) and substance P (Sub P). This anti-inflammatory state induced by mTBI has been correlated with an increased resistance to pneumonia (PNA) in mice and has been shown to be mediated in part by increased bacterial clearance in the lungs via enhanced neutrophil recruitment. However, it has not been investigated whether this reduced mortality is due to alterations in the coagulation system and if they have any effect on either the severity or occurrence of disseminated intravascular coagulation (DIC), a common sequelae of pneumonia-induced sepsis. Our study investigates whether administration of mTBI prior to pneumonia challenge in mice decreases mortality by ameliorating DIC. We assess and define DIC in our mouse models by changes in plasma coagulation parameters including fibrinogen, D-dimer, and plasminogen activator inhibitor-1 (PAI-1) Our study found that mTBI administration prior to pneumonia significantly decreased mortality in mice gavaged with high concentrations of Pseudomonas aeruginosa (Psd.). We also found that mTBI administration prior to pneumonia rescued fibrinogen levels and increased D-dimer levels in plasma, suggesting a compensated fibrinolytic state and amelioration of DIC. Circulating neutrophil and absolute leukocyte counts were also increased in mTBI/pneumonia models compared to those with pneumonia alone, supporting previous evidence implicating mTBI as the origin of enhanced bacterial clearance in lungs. Taken together, these data suggest that the increase in survival seen in patients with mTBI is in part due to alterations in coagulation.

Sepsis is defined as potentially fatal systemic inflammation, caused by an infection. It is the leading cause of ICU mortality and the 10th leading cause of death in the United States. Several models exist to mimic this disorder, and have demonstrated differential mortality rates between the models as well as the individual animals. Previous studies have shown that elevated levels of plasma mitochondrial DNA (mtDNA) correlated with mortality in septic patients, and cell-free mitochondrial DNA can elicit toll-like receptor mediated immune responses similar to LPS-mediated septicemia. However, the role of mtDNA in the pathophysiology sepsis is still unknown. The focus of this study was to create sepsis in a mouse model using the murine Cecal Ligation and Puncture (CLP) model, and measure plasma mtDNA levels. After CLP was performed on experimental mice, blood plasma was collected 24 hours later. Elevated amounts of circulating mtDNA were detectable in the plasma using real time PCR and cytochrome B2 as a marker of mitochondria. These data were correlated with plasma IL-6 levels, which were used to predict mortality within 5 days of CLP to stratify mice into two populations of those predicted to live or die following the procedure. We also aimed to investigate the effect of mtDNA and mitochondrial debris on naïve mouse macrophages in an in vitro study of the regulation of inflammatory cytokines interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α), and interleukin-1 beta (IL-1β). In order to observe the effects of mtDNA on murine macrophages, mitochondria was purified from mouse liver and used to stimulate these cells alongside positive control, LPS. Stimulation with mtDNA and mitochondrial debris resulted in increased levels of TNF-α mRNA in lysed cells as well as their surrounding media as compared to control cells, as well as increased transcript half life as measured over four hours post stimulation with transcription inhibitor actinomycin D. The increases in mRNA half-life elicited by mtDNA were comparable to those observed after LPS addition. Stimulation also caused increased binding of TNF-α mRNA to the RNA binding protein, AUF1, as measured by immunoprecipitation of RNA-protein complexes and assayed for TNF-α binding by PCR. These results demonstrate that mitochondrial damage-associated molecular patterns regulate TNF-α mRNA expression at the post-transcriptional level through AUF1, an mRNA destabilizing factor. This is a novel mechanism that likely contributes to sepsis pathophysiology, and demonstrates the involvement of the mitochondrial fission and fusion balance and its regulation in the sepsis innate immune response.

Sepse é um importante problema de saúde pública, uma vez que seu tratamento gera altos custos a um sistema de saúde já sobrecarregado. É uma síndrome de alta mortalidade e morbidade que afeta, em geral, pacientes jovens com plena capacidade produtiva. A identificação e o tratamento precoce desta síndrome reduzem a morbimortalidade, assim como o custo. A proteína C reativa (PCR) e a procalcitonina (PCT) são bem estudadas como ferramentas para diagnóstico de infecção bacteriana em imunocompetentes, mas seu uso como ferramenta diagnóstica ainda não está estabelecido em pacientes imunossuprimidos. Portanto, a proposta deste estudo é avaliar a acurácia diagnóstica destes biomarcadores, em pacientes críticos imunossuprimidos (vírus da imunodeficiência adquirida  HIV positivos, portadores de tuberculose (TBC), cirróticos e transplantados). Como o uso da proteína C ainda não está estabelecido, a primeira questão de pesquisa investigou seu potencial diagnóstico quando comparado ao teste padrão (cultural). O segundo artigo comparou a PCR com a PCT. Para isto foram realizados dois artigos de revisão sistemática com metanálise. O primeiro artigo comparou a acurácia em determinar infecção bacteriana em imunossuprimidos da PCR ao teste padrão-ouro (as culturas). A primeira revisão incluiu 1.418 pacientes e demonstrou uma boa acurácia da PCR como biomarcador no diagnóstico de infecção bacteriana, apresentando sensibilidade de 69% e especificidade de 76% com uma área sob a curva (AUC) de 0,77. Os resultados encontrados são similares aos da literatura para imunocompetentes,(3) sensibilidade de 75%, especificidade de 67% e Área Sob a Curva Receiver Operating Characteristic (AUROC) de 0,92. Quando a PCT foi comparada com a PCR, ambos os biomarcadores mostraram acurácia moderada na utilização como ferrramenta de diagnóstico de infecção bacteriana, com um diagnóstico da razão de chances (DOR) de 7,24 (95% CI (2,83-14,60) para PCT e de 5,56 (95% CI (5,21-10,30) para PCR. A PCT e a PCR apresentaram sensibilidade de 69% e 68% e uma especificidade de 75% e 71%, respectivamente. Ambas mostraram resultados semelhantes, podendo ser utilizadas no diagnóstico de sepse em imunossupressos. / Sepsis is a major public health problem, since its treatment generates high costs, a health system already overburdened. A high mortality and morbidity syndrome affects, in general, young patients with full production capacity. The identification and early treatment of this syndrome reduce morbidity and mortality as well as the cost. C-reactive protein (CRP) and procalcitonin (PCT) are well studied as tools for diagnosis of bacterial infection in immunocompetent patients, but its use as a diagnostic tool is not yet established in immunocompromised patients. Therefore, the purpose of this study is to evaluate the diagnostic accuracy of these biomarkers in immunosuppresses critical patients (human immunodeficiency virus, cirrhotic and transplant). As the use of the c protein is not yet established, the first research question investigated their diagnostic potential when compared to the pattern (cultural). The second article compared to CRP and PCT. For this, there were two articles of a systematic review and meta-analysis. The first article compared the accuracy in determining bacterial infection in immunosuppresses of CRP to the gold standard (cultures). Our first review included 1,418 patients and showed good accuracy of CRP as a biomarker for the diagnosis of bacterial infection presenting a sensibility of 69% and 76% specificity with an area under the curve (AUC) 0.77. The results are similar to those found in the literature for immunocompetent,(3) sensitivity 75%, specificity of 67% and Area Under the Receiver Operating Characteristic Curve (AUROC): 0.92. When the PCT was compared with PCR, both biomarkers showed a moderately accurate for use as tool diagnostic bacterial infection with a Odds ratio diagnostic (DOR) 7.24 (95% CI (2.83-14.60) and PCT to 5:56 (95% CI (5.21-10.30) for CRP. the PCT and CRP had a sensitivity of 69% and 68% and a specificity of 75% and 71%, respectively. Both showed similar results may be used in the diagnosis of sepsis in immunosuppression.

Presepsin (PSEP) is released during infectious diseases and can be detected in the blood. PSEP has shown promising results as sepsis marker. We examined the diagnostic and prognostic validity of PSEP in patients suspicious of sepsis on admission in the emergency department (ED). Methods One hundred twenty three patients with signs of SIRS and/or sepsis and 123 healthy individuals were enrolled. PSEP was determined on admission, after 8, 24 and 72 h. Results Mean PSEP concentrations of the control group and the patient group were 130 and 1945 pg/ml. PSEP differed between SIRS, sepsis, severe sepsis and septic shock and showed strong association with 30-day mortality ranging from 10.3% in the 1st to 32.1% in the 4th quartile. The ROC curve analyses revealed an AUC value of 0.743. Combined assessment of PSEP and MEDS score increased the AUC up to 0.878 demonstrating the close relationship with outcome. Based on the PSEP values in the different severity degrees, decision thresholds for risk stratification were established. The course of PSEP during the first 72 h was associated with effectiveness of treatment and outcome. Conclusions PSEP allowed outcome prediction already on admission to a similar degree as the clinical scores MEDS and APACHE II. Combination of PSEP with MEDS score improved the discriminatory power for outcome prediction. / Our study has been supported by Mitsubishi Chemical Europe through providing the PSEP reagents free of charge. Dr. Carpio has received speaker honoraria from Mitsubishi Chemical Europe. DIAneering – Diagnostics Engineering & Research consulted to Axis Shield Diagnostics, Mitsubishi Chemical Europe, Radiometer, Roche Diagnostics, Shanghai Kehua Bio-engineering. No potential conflict of interest to this paper was reported / Peer review

Bakgrund: Sepsis, svår sepsis och septisk chock är ett accelererande tillstånd där tid till antibiotika och vätskebehandling har stor betydelse för denna utsatta patientgrupp. Trots att nästan hälften av sepsispatienterna anländer till sjukhus med ambulans, så gör diffusa symtom att enbart en liten del identifierats eller mottagit någon form av behandling. Syfte: Att belysa svaga länkar i den tidiga vårdkedjan för patienter som i efterhand diagnostiserats med positiv blododling/sepsis. Metod: Kvantitativ registerstudie. Samtliga patienter (n=696) i Sahlgrenska Universitets sjukhus upptagningsområde som testades med positiv blododling (Sepsis) under perioden: 1 februari till 30 april 2012 analyserades. I studien inkluderades enbart de patienter (n=308) som sökt via ett akutintag dit de antingen tog sig själva eller anlände med ambulans. Resultat: Jämfört med de patienter som själva tog sig till akutintaget var ambulanspatienterna oftare äldre män med andnings- och medvetandepåverkan. Färre av ambulanspatienterna var vid liv efter 12 månader. Majoriteten av alla patienter hade förändrad temperatur (&gt;90 %) men enbart ett fåtal (&lt;5 %) hade ett systoliskt blodtryck under 90 mmHg. Vi fann ingen statistisk säkerställd skillnad i tid till antibiotika (&lt;4 tim) mellan grupperna. Ambulanspersonalen diagnostiserade 39 % av patienterna som infektion (32,5 %) och sepsis (6,5 %) men ansåg att 68 % behövde omedelbar vård (Retts: orange/röd). Tolv procent fick intravenös infusion, 7 % fick febernedsättande och 4 % behandlades med smärtlindrande medicin. Tid till antibiotika minskade från tre- till en timme i de fall där ambulanspersonal misstänkte sepsis. Konklusion: Då sepsis inte diagnostiserades prehospitalt tog det tre gånger så lång tid till att antibiotikabehandling påbörjades. Detta trotts att majoriteten (68 %) av ambulanspatienterna av ambulanssjuksköterskan bedömdes behöva omedelbar vård (REETS röd/orange). Enligt gällande prehospitala behandlingsriktlinjer är vätskebehandling enbart indicerat vid svår sepsis (då systoliskt blodtryck &lt;90 mmHg). Nya mer generösa behandlingsriktlinjer för patienter med misstänkta infektioner/sepsis bör övervägas då rekommendationer intrahospitalt förekommer både gällande intravenös infusion samt febernedsättande läkemedel. / Program: Specialistsjuksköterskeutbildning med inriktning mot ambulanssjukvård

Os leucotrienos (LTs) e as citocinas são mediadores inflamatórios produzidos durante a sepse e que podem apresentar uma tarefa moduladora na secreção de arginina-vasopressina (AVP). Na fase aguda da sepse as concentrações plasmáticas de AVP encontram-se aumentadas e na fase tardia em níveis basais, apesar da hipotensão progressiva. Nosso objetivo foi analisar o efeito da injeção intraperitoneal de MK-886, (inibidor da síntese de leucotrienos), sobre a produção de LTs e de citocinas no sítio infeccioso e plasma de animais sépticos. Foram usados ratos Wistar que receberam injeções intraperitoneais de MK-886 (4mg/kg) ou veículo (DMSO 5%) 1h antes da indução de sepse experimental por ligadura e perfuração cecal (CLP) ou da operação fictícia (OF). Os animais foram decapitados 0, 4 e 24h após CLP ou OF. O sangue foi coletado para a determinação do sódio sérico, hematócrito, citocinas, LTB4. O lavado peritoneal foi coletado para dosagem de citocinas. O hipotálamo foi removido e incubado para a análise de cis-LTs. O sódio sérico diminuiu, o hematócrito aumentou e os cis-leucotrienos hipotalâmicos não apresentaram alterações significantes após 4 horas de CLP. A sepse provocou um aumento nas concentrações de TNF-a, no sítio infeccioso, mas não foi detectada no plasma. As citocinas IL-1b e a IL-6 aumentaram em 4 horas tanto no sítio infeccioso como no plasma. A análise dos resultados de LTB4 plasmático mostrou uma diminuição das concentrações 24 horas após o CLP. A administração do MK-886 não modificou o sódio, o hematócrito e aparentemente nem os cis-leucotrienos hipotalâmicos. No sítio infeccioso a droga causou um aumento nas concentrações de TNF-a 4 e 24 horas enquanto as de IL-1b não se modificaram e as de IL-6 aumentaram apenas 4 horas após o CLP. Entretanto, no plasma, a droga diminuiu acentuadamente as concentrações tanto de IL-1b como de IL-6. A análise do LTB4 plasmático mostrou efeito da droga em 4 horas, mas não em 24 horas da sepse. Os resultados deste estudo sugerem que os LTS regulam de maneira diferente a produção de citocinas no plasma e no foco infeccioso e podem explicar os efeitos destes mediadores sobre a secreção de AVP durante a sepse. / Leukotrienes (LTs) and cytokines are inflammatory mediators produced during sepsis and may have a task in modulating the secretion of vasopressin (VP). In the acute phase of sepsis, the plasma AVP concentrations are increased and in the late stage at basal levels, despite the progressive hypotension. Our goal was to analyze the effect of intraperitoneal injection of MK-886 (leukotriene synthesis inhibitor) on the production of LTs and cytokines in the infectious site and plasma of septic animals. We used Wistar rats that received intraperitoneal injection of MK-886 (4.0mg/kg) or vehicle (DMSO 5%) 1 h before induction of experimental sepsis by cecal ligation and puncture (CLP) or sham operation (OF).The animals were decapitated at 0, 4 and 24 h after surgeries and blood was collected for determination of serum sodium, hematocrit, cytokines and LTB4. The peritoneal fluid was collected for cytokine assay. The hypothalamus was removed and incubated for the analysis of cis-LTs. Serum sodium decreased, hematocrit increased and hypothalamic cis-leukotrienes did not show significant changes after 4 hours of CLP. Sepsis caused increase in the TNF-a concentrations in the site of infection, but was not detected in plasma. The cytokines IL-1b and IL-6 increased by 4 hours both in the plasma as in the site of infection. The results of LTB4 showed a decrease in plasma concentrations 24 hours after CLP. The administration of MK-886 did not alter sodium, hematocrit, and apparently neither the cis-hypothalamic leukotrienes. At the site of infection the drug caused an increase in the concentrations of TNF-a 4 and 24 hours while IL-1b remained unchanged and IL-6 increased only 4 hours after CLP. However, in the plasma the drug markedly diminished the concentrations of IL-1b and IL-6. The analysis of LTB4 in plasma showed effects of the drug in 4 hours but not 24 hours of sepsis. The results of this study suggest that LTs differentially regulate the cytokine production in the plasma and in the site of infection, and may explain the effects of these mediators in AVP secretion during sepsis.

Orientador: Antonio Nader Filho / Banca: Ruben-Pablo Schocken-Iturrino / Banca: José Jurandir Fagliari / Banca: Luiz Francisco Zafalon / Banca: Nilson Roberti Benites / Resumo: Com o objetivo de avaliar a eficácia do ácido lático frente ao iodo na anti-sepsia após a ordenha na prevenção da mastite bovina, foram estudados 668 quartos mamários de vacas holandesas em dois rebanhos leiteiros, sendo um experimental e outro comercial. Para tanto adotou-se o delineamento experimental tipo split-herd baseado na redução da ocorrência natural de novos casos de infecções intramamárias (recomendado pelo protocolo do National Mastistis Council). Os resultados obtidos evidenciaram que o ácido lático apresentou uma eficácia de 57,4% na redução do número de novos casos de IIM causados por Corynebacterium spp., Staphylococcus spp. e Streptococcus spp. entre as vacas estudadas no rebanho experimental. No rebanho comercial a eficácia foi de 42,9% entre os casos de IIM determinados por Staphylococcus spp., porém não mostrou eficácia entre os casos de IIM determinados por Corynebacterium spp. e Streptococcus spp.. Tanto o ácido lático quanto o iodo não interferiram nas taxas de cura espontânea, persistência de IIM e na ocorrência de casos clínicos de mastite entre as vacas estudadas nos rebanhos experimental e comercial. A utilização do ácido lático na anti-sepsia dos tetos de vacas após a ordenha, mostrou-se eficaz na redução do número de novos casos de IIM, desde que acompanhada por outras medidas de controle. / Abstract: With the objective of evaluate the efficacy of a lactic acid against an iodine postmilking teat dip in preventing bovine mastitis, 668 mammary quarters from Holsteins cows in two dairy herds, one experimental and other commercial, were studied. The National Mastitis Council recommended protocol for determining efficacy of a postmilking barrier teat dip based on reduction of naturally occurring new intramammary infections was used, in a split-herd experimental design. Results demonstrated that lactic acid showed an efficacy of 57.4% in reducing new intramammary infections caused by Corynebacterium spp., Staphylococcus spp. and Streptococcus spp., among the experimental herd cows. In the commercial herd, the efficacy was 42.9%, but only in reducing the number of new cases of intramammary infections caused by Staphylococcus spp., and not for new cases of intramammary infections caused by Corynebacterium spp., and Streptococcus spp. Both, lactic acid and iodine didn't interfere in spontaneous cure, intramammary infections persistence and clinical mastitis rates in experimental and commercial herds. The use of a lactic acid teat dip antisepsis after each milking was effective in reducing new cases of IMI, as part of a mastitis control program. / Doutor