Sildenafil in endotoxin-induced pulmonary hypertension: an experimental study

Abstract Background Sepsis and septic shock still represent great challenges in critical care medicine. Sildenafil has been largely used in the treatment of pulmonary arterial hypertension, but its effects in sepsis are unknown. The aim of this study was to investigate the hypothesis that sildenafil can attenuate endotoxin-induced pulmonary hypertension in a porcine model of endotoxemia. Methods Twenty pigs were randomly assigned to Control group (n = 10), which received saline solution; or to Sildenafil group (n = 10), which received sildenafil orally (100 mg). After 30 minutes, both groups were submitted to endotoxemia with intravenous bacterial lipopolysaccharide endotoxin (LPS) infusion (4 µg.kg-1.h-1) for 180 minutes. We evaluated hemodynamic and oxygenation functions, and also lung histology and plasma cytokine (TNFα, IL-1β, IL6, and IL10) and troponin I response. Results Significant hemodynamic alterations were observed after 30 minutes of LPS continuous infusion, mainly in pulmonary arterial pressure (from Baseline 19 ± 2 mmHg to LPS30 52 ± 4 mmHg, p< 0.05). There was also a significant decrease in PaO2/FiO2 (from Baseline 411 ± 29 to LPS180 334 ± 49, p< 0.05). Pulmonary arterial pressure was significantly lower in the Sildenafil group (35 ± 4 mmHg at LPS30, p< 0.05). The Sildenafil group also presented lower values of systemic arterial pressure. Sildenafil maintained oxygenation with higher PaO2/FiO2 and lower oxygen extraction rate than Control group but had no effect on intrapulmonary shunt. All cytokines and troponin increased after LPS infusion in both groups similarly. Conclusion Sildenafil attenuated endotoxin-induced pulmonary hypertension preserving the correct heart function without improving lung lesions or inflammation.

Disseminated intravascular coagulation phenotype is regulated by the TRPM7 channel during sepsis

BACKGROUND: Sepsis is an uncontrolled inflammatory response against a systemic infection that results in elevated mortality, mainly induced by bacterial products known as endotoxins, producing endotoxemia. Disseminated intravascular coagulation (DIC) is frequently observed in septic patients and is associated with organ failure and death. Sepsis activates endothelial cells (ECs), promoting a prothrombotic phenotype contributing to DIC. Ion channel mediated calcium permeability participates in coagulation. The transient reception potential melastatin 7 (TRPM7) non-selective divalent cation channel that also contains an α-kinase domain, which is permeable to divalent cations including Ca2+, regulates endotoxin-stimulated calcium permeability in ECs and is associated with increased mortality in septic patients. However, whether endothelial TRPM7 mediates endotoxemia-induced coagulation is not known. Therefore, our aim was to examine if TRPM7 mediates coagulation during endotoxemia. RESULTS: The results showed that TRPM7 regulated endotoxin-induced platelet and neutrophil adhesion to ECs, dependent on the TRPM7 ion channel activity and by the α-kinase function. Endotoxic animals showed that TRPM7 mediated neutrophil rolling on blood vessels and intravascular coagulation. TRPM7 mediated the increased expression of the adhesion proteins, von Willebrand factor (vWF), intercellular adhesion molecule 1 (ICAM-1), and P-selectin, which were also mediated by the TRPM7 α-kinase function. Notably, endotoxin-induced expression of vWF, ICAM-1 and P-selectin were required for endotoxin-induced platelet and neutrophil adhesion to ECs. Endotoxemic rats showed increased endothelial TRPM7 expression associated with a procoagulant phenotype, liver and kidney dysfunction, increased death events and an increased relative risk of death. Interestingly, circulating ECs (CECs) from septic shock patients (SSPs) showed increased TRPM7 expression associated with increased DIC scores and decreased survival times. Additionally, SSPs with a high expression of TRPM7 in CECs showed increased mortality and relative risk of death. Notably, CECs from SSPs showed significant results from the AUROC analyses for predicting mortality in SSPs that were better than the Acute Physiology and Chronic Health Evaluation II (APACHE II) and the Sequential Organ Failure Assessment (SOFA) scores. CONCLUSIONS: Our study demonstrates that sepsis-induced DIC is mediated by TRPM7 in ECs. TRPM7 ion channel activity and α-kinase function are required by DIC-mediated sepsis-induced organ dysfunction and its expression are associated with increased mortality during sepsis. TRPM7 appears as a new prognostic biomarker to predict mortality associated to DIC in SSPs, and as a novel target for drug development against DIC during infectious inflammatory diseases.

Efeito da ingestão de suco de uva sobre a expressão de microRNA no plasma e em células mononucleares em mulheres com sobrepeso / Effect of grape juice ingestion on microRNA expression in plasma and mononuclear cells in overweight women

Introdução: O padrão da dieta ocidental, caracterizado pela ingestão elevada de açúcares e lipídios, está relacionado à gênese da inflamação metabólica, a qual é caracterizada por um quadro de inflamação crônica, sistêmica e de baixa intensidade. A inflamação metabólica contribui para o desenvolvimento de doenças crônicas não transmissíveis (DCNT), as quais representam a principal causa de morte no mundo. Por outro lado, a ingestão de uma dieta saudável, rica em frutas e hortaliças, é relevante para a redução do risco das DCNT. Nesse contexto, destaca-se o suco de uva, bebida rica em compostos bioativos, que possui potencial efeito anti-inflamatório. Estudos recentes sugerem que nutrientes e compostos bioativos dos alimentos possam atenuar a inflamação por meio da modulação da expressão de microRNA, os quais representam novos biomarcadores inflamatórios, contribuindo para a identificação do risco de desenvolvimento de doenças cardiometabólicas. Objetivo: Investigar o efeito da ingestão de suco de uva integral sobre a expressão de microRNA no plasma e em células mononucleares do sangue periférico (PBMC) e sua relação com a resposta inflamatória. Métodos: Estudo intervencional, no qual, durante quatro semanas, as voluntárias ingeriram 500 mL diários de suco de uva integral, com coletas de sangue no momento basal, 2 e 4 semanas após o início da ingestão da bebida. A população do estudo foi composta por mulheres saudáveis (n = 20), com idade entre 18 e 40 anos e índice de massa corpórea classificado como sobrepeso (25,0 - 29,9 kg/m2). Foram avaliados o perfil lipídico, o hemograma e as concentrações plasmáticas de glicose, insulina, leptina, adiponectina, lipopolissacarídeos e seus ligantes plasmáticos LBP e sCD14 e biomarcadores inflamatórios (IL-10, IL-6, TNF-α, proteína C reativa, MCP-1, VCAM-1 e ICAM-1), bem como a concentração dos principais compostos bioativos presentes no suco de uva. Dois painéis de microRNA, sendo um plasmático e outro de PBMC, de 8 indivíduos foram avaliados, em todos os tempos, pelo ensaio de PCR em tempo real. Posteriormente, foram selecionados os microRNA relacionados à resposta inflamatória e à endotoxemia metabólica, os quais foram avaliados no restante das voluntárias. As proteínas-alvo dos microRNA em PBMC foram analisadas por Western Blotting. O consumo alimentar foi avaliado pela análise de três recordatórios de 24 horas coletados em todos os tempos experimentais. Resultados: A ingestão de suco de uva, após 4 semanas de intervenção, em PBMC, promoveu redução da expressão gênica do TNF-α, IL-6 e das subunidades p50 e p65, as quais compõem o fator de transcrição NF-kB. Em relação ao conteúdo de proteínas nestas células, houve apenas diminuição da fosforilação da TAK-1 em relação ao seu conteúdo total. Dentre os miRNA analisados, o miR-144-5p e o miR-144-3p tiveram a sua expressão aumentada no plasma e em PBMC, respectivamente. A concentração plasmática de sICAM-1 reduziu significativamente na semana 4 em relação a semana 2, enquanto, a concentração plasmática de gama glutamil transferase aumentou significativamente na última semana. As voluntárias apresentaram menor pressão arterial diastólica no final do estudo em relação ao momento basal. Conclusão: A ingestão do suco de uva, após 4 semanas de intervenção, influenciou a concentração de biomarcadores relacionados ao processo inflamatório mediado pela via do fator de transcrição NF-kB em mulheres com sobrepeso.

Introduction: The Western diet pattern, characterized by excessive ingestion of sugars and lipids, is related to the genesis of metabolic inflammation, which is characterized by a condition of a chronic, systemic, and low-grade inflammation. Metabolic inflammation contributes to the development of chronic non-communicable diseases (NCDs), the leading cause of death in the world. On the other hand, the adoption of a healthy diet rich in fruits and vegetables is relevant to reduce the risk of NCDs. Grape juice is a beverage rich in bioactive compounds which have a potential anti-inflammatory effect. Recent studies suggest that nutrients can modulate inflammation through microRNAs action, which have been highlighted because they are new inflammatory biomarkers, contributing to the identification of the risk of cardiometabolics diseases. Objective: Investigate the effect of the grape juice ingestion on microRNA expression in plasma and peripheral blood mononuclear cells (PBMC) and its relationship with the development of the inflammatory process. Methods: Interventional study, in which, for four weeks, the volunteers ingested 500 mL of whole grape juice daily, with blood drawn at baseline, 2 and 4 weeks after the intake. The study population consisted of healthy women (n = 20), aged between 18 and 40 years and body mass index classified as overweight (25.0 - 29.9 kg/m2). The lipid profile, blood count and plasma concentrations of glucose, insulin, leptin, adiponectin, lipopolysaccharides, and their plasma ligands LBP and sCD14 and inflammatory biomarkers (IL-10, IL-6, TNF-α, C-reactive protein, MCP-1, VCAM-1 and ICAM-1) were evaluated, as well as the concentration of the main bioactive compounds present in grape juice. Two microRNA panels, one plasma and the other PBMC, from 8 individuals were evaluated, at all times, by real-time PCR. Subsequently, microRNAs related to the inflammatory response and metabolic endotoxemia were selected, which were evaluated in the rest of the volunteers. MicroRNA target proteins in PBMC were analyzed by Western Blotting. Food consumption was assessed by analyzing three 24-hour recalls collected at all experimental times. Results: The intake of grape juice, after 4 weeks of intervention, in PBMC, promoted a reduction in gene expression of TNF-α, IL-6 and p50 and p65 subunits, which composes the NF-kB transcription factor. Regarding the protein content in these cells, there was only a decrease in TAK-1 phosphorylation in relation to its total content. Among the analyzed miRNAs, miR-144-5p and miR-144-3p had their expression increased in plasma and in PBMC, respectively. The plasma concentration of sICAM-1 significantly reduced in week 4 compared to week 2, while the plasma concentration of gamma glutamyl transferase increased significantly in the last week. The volunteers had lower diastolic blood pressure at the end of the study compared to baseline. Conclusion: The intake of grape juice after 4 weeks of intervention influenced the concentration of biomarkers related to the inflammatory process mediated by the transcription factor NF-kB pathway in overweight women.

Recuperando el "hilo perdido" en la madeja de la enterocolitis necrotizante del recién nacido / Recovering the "lost thread" in the skein of necrotizing enterocolitis of the newborn / Recuperando o "fio perdido" no novelo de enterocolite necrosante do recém-nascido

Propósito de la revisión: el objetivo de la revisión es delinear la fisiopatología de la Entero Colitis Necrotizante (ECN) clásica del recién nacido, proponer un listado de antecedentes perinatales que definan un grupo de riesgo y establecer parámetros simples y objetivos, que ayuden a establecer un diagnóstico clínico precoz. Recientes hallazgos: La mortalidad de la Entero Colitis Necrotizante clásica del recién nacido sigue siendo elevada. Aunque la enfermedad tiene varias presentaciones, es única con una única vía de instalación, la hipoperfusión intestinal como agente agresor inicial. Extracto: La falta de estrategias de prevención y el diagnóstico clínico muy tardío explican la mortalidad elevada de la ECN. Existe una gran confusión sobre el origen de la enferme-dad, dando a entender que existen diferentes tipos de ECN, aunque la enfermedad es única, tiene diferentes manifestaciones según las condiciones del recién nacido con una vía común de instalación, la hipoperfusión intestinal como agente agresor inicial. En este artículo se postula que reconocer a la hipoperfusión intestinal como agente agresor inicial, es "encontrar el hilo perdido", que permitirá desarrollar estrategias de prevención y tratamiento, al identificar los pacientes en riesgo de ECN y lograr el diagnóstico de manera precoz.

Purpose of the review: The objective of the review is to delineate the pathophysiology of the classic Entero Necrotizing Colitis (NEC) of the newborn, propose a list of perinatal ante-cedents that define a risk group and establish simple and objective parameters that help to establish an early clinical diagnosis. Recent findings: The mortality of the classic Necrotizing Entero Colitis of the newborn is still high. Although the disease has several presentations, it is unique with a single installation route, intestinal hypoperfusion as the initial offending agent. Excerpt: The lack of prevention strategies and very late clinical diagnosis explain the high mortality of NEC. There is great confusion about the origin of the disease, giving to under-stand that there are different types of NEC, although the disease is unique, it has different manifestations according to the conditions of the newborn with a common route of installation, intestinal hypoperfusion as an initial assailant agent. This article postulates that recog-nizing intestinal hypoperfusion as the initial offending agent is "finding the lost thread", that developing prevention and treatment strategies, by identifying patients at risk of NEC and achieving a diagnosis in a way early.

Objetivo da revisão: O objetivo da revisão é delinear a fisiopatologia da Entero Colite Necrosante (NEC) clássica do recém-nascido, propor uma lista de antecedentes perinatais que definem um grupo de risco e estabelecer parâmetros simples e objetivos que auxiliem no diagnóstico clínico precoce. Descobertas recentes: A mortalidade da Entero Colite Necrosante clássica do recém-nascido ainda é alta. Embora a doença tenha várias apresentações, é única com uma única via de instalação, a hipoperfusão intestinal como agente agressor inicial. Resumo: A falta de estratégias de prevenção e o diagnóstico clínico muito tardio explicam a alta mortalidade da NEC. Há grande confusão sobre a origem da doença, sugerindo que existem diferentes tipos de NEC, embora a doença seja única, ela tem diferentes manifestações dependendo das condições do recém-nascido com uma via comum de instalação, a hipoperfusão intestinal como agente. assaltante inicial. Este artigo postula que reconhecer a hipoperfusão intestinal como agente agressor inicial é "encontrar o fio condutor", o que permitirá o desenvolvimento de estratégias de prevenção e tratamento, por meio da identificação de pacientes em risco de NEC e do diagnóstico precoce.

Study on synergistic effect of Qingkailing Injection and Shengmai Injection on organ injury in endotoxemia rats / 中国中药杂志

As a dangerous disease with rapid progression, endotoxemia is easy to induce the damage to multiple organs. However, its specific and efficient treatment methods are still lacking at present. Both Qingkailing Injection(QKLI) and Shengmai Injection(SMI) have been proved effective in anti-inflammation, anti-endotoxin and organ protection. In this study, carrageenan and endotoxin were injected successively into rats to establish an endotoxemia model. Different doses of QKLI and SMI were administered to the endotoxemia rats by intraperitoneal injection separately or in combination. Then the count of white blood cells, the number of platelets, the content of cytokines, biochemical indexes, organ coefficient and pathological changes of main organs in the rats were detected. The results showed that the rats in the model group had obvious symptoms of endotoxemia, i.e., leucopenia, thrombocytopenia, increase in cytokines(IL-6 and TNF-α) and biochemical indexes of liver and kidney function as well as pathological damage to liver, kidney and lung. QKLI alone can alleviate the above symptoms of endotoxemia and the organ injury. SMI alone is less effective in improving disseminated intravascular coagulation(DIC) and cytokine secretion complicated with endotoxemia, but capable of reducing the inflammation degree of the lung, liver and kidney. The combination of QKLI and SMI remarkably increased the number of platelets in the peripheral blood, improved the liver and kidney function and reduced inflammatory factors, with lung, liver, kidney and other organ structures protected well. Moreover, the improvement effect of the combination of QKLI and SMI was stronger than those of the two injections alone at fixed doses, indicative of a synergistic effect.

Study of thermo-regulation as a worsening marker of experimental sepsis in an animal model / Estudo da termorregulação como marcador de agravamento do quadro de sepse experimental em modelo animal / Estudio de la termorregulación como marcador asociado al empeoramiento del cuadro de la sepsis experimental en un modelo con animales

Objective: to analyze variations in body temperature and in plasma nitrate and lactate concentrations in rats submitted to the experimental sepsis model. Method: a total of 40 rats divided equally into five groups. The induction of endotoxemia was performed with intravenous administration of lipopolysaccharide, 0.5 mg/Kg, 1.5 mg/Kg, 3.0 mg/Kg, and 10 mg/Kg, respectively. The control group received 0.5 mL of saline solution. The experiment lasted six hours, with evaluations performed at 0 (baseline data), 2nd, 4th, and 6thhours. Results: The animals that received doses up to 3.0 mg/kg showed a significant increase in body temperature compared to the group with 10 mg/kg, which showed a decrease in these values. The increase in plasma nitrate and lactate concentrations in the groups with lipopolysaccharide was significantly higher than in the group that received the saline solution and was correlated with the increase in body temperature. Conclusion: the variations in body temperature observed in this study showed the dose-dependent effect of lipopolysaccharide and were correlated with the increase in the concentrations of nitrate and plasma lactate biomarkers. The implications of this study are the importance of monitoring body temperature, together with the assessment of these pathophysiological markers, which suggest worsening in the prognosis of sepsis.

Objetivo: analisar as variações na temperatura corporal e nas concentrações de nitrato e lactato plasmáticos em ratos submetidos ao modelo de sepse experimental. Método: foram utilizados 40 ratos divididos igualmente em cinco grupos. A indução da endotoxemia foi realizada com administração endovenosa de lipopolissacarídeo, respectivamente 0,5 mg/Kg, 1,5 mg/Kg, 3,0 mg/Kg e 10 mg/Kg. O grupo controle recebeu 0,5 mL de solução salina. O experimento teve duração de seis horas, com avaliações realizadas na 0 (dados basais), 2a, 4a e 6a hora. Resultados: os animais que receberam doses de até 3,0 mg/Kg apresentaram aumento significativo na temperatura corporal em relação ao grupo com 10 mg/Kg, que apresentou diminuição nesses valores. O aumento nas concentrações de nitrato e lactato plasmáticos nos grupos com lipopolissacarídeo foi significativamente superior ao grupo que recebeu salina e esteve correlacionado com o aumento na temperatura corporal. Conclusão: as variações na temperatura corporal observadas neste estudo mostraram efeito dose dependentes de lipopolissacarídeo e estiveram correlacionadas com o aumento nas concentrações dos biomarcadores nitrato e lactato plasmáticos. O estudo traz como implicações, a importância no monitoramento da temperatura corporal, em conjunto com a avaliação destes marcadores fisiopatológicos, os quais sugerem agravamento no prognóstico da sepse.

Objetivo: analizar las variaciones de la temperatura corporal y de las concentraciones de nitrato y lactato en plasma en ratones sometidos a un modelo de sepsis experimental. Método: se utilizaron 40 ratones divididos en cinco grupos iguales. La inducción de la endotoxemia se realizó mediante administración intravenosa de 0,5 mg/Kg, 1,5 mg/Kg, 3,0 mg/Kg y 10 mg/Kg de lipopolisacárido, respectivamente. El grupo de control recibió 0,5 mL de solución salina. El experimento duró seis horas, con evaluaciones realizadas a la hora 0 (datos de referencia) y a la 2a, 4a y 6ahora. Resultados: los animales que recibieron dosis de hasta 3,0 mg/kg presentaron un aumento significativo de la temperatura corporal, en comparación con el grupo al que se le administró 10 mg/kg, que presentó una disminución de dichos valores. En los grupos a los que se les administró lipopolisacárido, el aumento en las concentraciones de nitrato y lactato en plasma fue significativamente mayor que en el grupo al que se le administró la solución salina y estuvo correlacionado con el aumento de la temperatura corporal. Conclusión: las variaciones de la temperatura corporal observadas en este estudio mostraron que los efectos dependieron de la dosis de lipopolisacárido, y estuvieron correlacionadas con el aumento en la concentración de biomarcadores, como el nitrato y lactato en plasma. El estudio reveló la importancia del control de la temperatura corporal, junto con la evaluación de estos marcadores fisiopatológicos, que sugieren un empeoramiento en el pronóstico de la sepsis.

Protective effect of dexmedetomidine against organ dysfunction in a two-hit model of hemorrhage/resuscitation and endotoxemia in rats

Dexmedetomidine (DEX), a selective agonist of α2-adrenergic receptors, has anti-inflammation properties and potential beneficial effects against trauma, shock, or infection. Therefore, this study aimed to investigate whether DEX might protect against multiple-organ dysfunction in a two-hit model of hemorrhage/resuscitation (HS) and subsequent endotoxemia. Eighty Wistar rats were randomized into four groups: NS (normal saline), HS/L (HS plus lipopolysaccharide), HS/L+D (HS/L plus dexmedetomidine), and HS/L+D+Y (HS/L+D plus yohimbine). Six hours after resuscitation, blood gas (PaO2) and serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urine nitrogen (BUN), creatinine (Cr), TNF-α, IL-β, IL-6, IL-8, IL-10, and nitric oxide (NO) were measured. The histopathology was assayed by staining. Malondialdehyde (MDA) and superoxide dismutase (SOD) levels and heme oxygenase-1 (HO-1) were assayed. The PaO2 levels in HS/L rats were lower whereas the ALT, AST, BUN, Cr, TNF-α, IL-β, IL-6, IL-8, IL-10, and NO levels were higher compared to the control group. The HS/L+D increased PaO2 and further increased IL-10 and decreased ALT, AST, BUN, Cr, TNF-α, IL-β, IL-6, IL-8, and NO levels of the HS/L groups. In addition, the MDA in the HS/L groups increased whereas SOD activity decreased compared to the control group. Moreover, the HO-1 expression levels were increased by DEX administration in lung, liver, and kidney tissues. Lungs, livers, and kidneys of the HS/L group displayed significant damage, but such damage was attenuated in the HS/L+D group. All of the above-mentioned effects of DEX were partly reversed by yohimbine. DEX reduced multiple organ injury caused by HS/L in rats, which may be mediated, at least in part, by α2-adrenergic receptors.

Bowel Wall Thickness, Elasticity, Intestinal Permeability, and Inflammatory Cytokines in Patients with Cirrhosis and Ascites / 대한내과학회지

BACKGROUND/AIMS: Previous studies have reported that endotoxemia is associated with pathogenesis and complications in cirrhosis. Endotoxin stimulates the secretion of inflammatory cytokines, which contributes to the development of complications. In addition, endotoxin easily invades the gut barrier system because of the increased intestinal permeability due to portal hypertensive enteropathy. In this report, we explored changes in cytokine levels and intestinal permeability and measured the thickness and elasticity of the bowel wall using ultrasonography in cirrhotic patients.METHODS: We enrolled 40 patients with cirrhosis classified as Child-Pugh B or C and 20 healthy volunteers. Abdominal ultrasonography examinations were used to evaluate bowel wall parameters in the ascending colon and terminal ileum. Intestinal permeability was measured using dual sugar absorption tests with lactulose and mannitol. Levels of tumor necrosis factor (TNF)-α and IL-10 were determined from blood samples. We compared these outcomes between cirrhotic patients and healthy controls and between Child-Pugh B and C patients. In addition, we explored the correlation between cytokine levels, intestinal permeability ratio, and bowel wall parameters in cirrhotic patients.RESULTS: In cirrhotic patients, the ascending colon wall elasticity decreased (20.4 vs. 10.9 kPa, p = 0.048) and the terminal ileum wall thickness increased (4.2 vs. 1.9 mm, p < 0.001). The intestinal permeability ratio and levels of the cytokines TNF-α and IL-10 increased (0.219 vs. 0.017, p < 0.001; 22.47 vs. 13.48 pg/mL, p < 0.001; and 14.91 vs. 8.57 pg/mL, p = 0.019, respectively) in cirrhotic patients. However, there were no significant differences between Child-Pugh classes and no significant correlations between bowel wall parameters and intestinal permeability or cytokine levels.CONCLUSIONS: Ultrasonography revealed bowel wall thickening and decreases in elasticity; in addition, intestinal permeability and cytokine levels increased in cirrhotic patients compared with healthy controls.

Mild encephalopathy with a reversible splenial lesion in a girl with acute pyelonephritis / 소아과

We report the case of a 12-year-old girl who had mild encephalopathy with a reversible splenial lesion (MERS) associated with acutepyelonephritis caused by Escherichia coli. The patient was admitted with a high fever, and she was diagnosed with acute pyelonephritis based on pyuria and the results of urine culture, which detected cefotaxime-sensitive E. coli. Although intravenous cefotaxime and tobramycin were administered, her fever persisted and her C-reactive protein level increased to 307 mg/L. On day 3 of admission, she demonstrated abnormal neuropsychiatric symptoms, such as delirium, ataxia, and word salad. Magnetic resonance imaging (MRI) of the brain performed on day 4 showed marked hyperintensities in the bilateral corpus callosum and deep white matter on diffusion-weighted images, with corresponding diffusion restriction on apparent diffusion coefficient mapping. No abnormalities or pathogens were detected in the cerebrospinal fluid; however, lipopolysaccharides (LPS, endotoxin) were detected in plasma (41.6 pg/mL), associated with acute neurological deterioration. Her clinical condition gradually improved, and no neurological abnormalities were observed on day 6. Follow-up brain MRI performed 2 weeks later showed near-disappearance of the previously noted hyperintense lesions. In this patient, we first proved endotoxemia in a setting of MERS. The release of LPS following antibiotic administration might be related to the development of MERS in this patient. The possibility of MERS should be considered in patients who present with acute pyelonephritis and demonstrate delirious behavior.

A calpain inhibitor protects against fractalkine production in lipopolysaccharide-treated endothelial cells

BACKGROUND: Fractalkine (CX3CL1) is a chemokine with a unique CX3C motif and is produced by endothelial cells stimulated with lipopolysaccharide (LPS), tumor necrosis factor (TNF)-α, interleukin (IL)-1, and interferon-γ. There have been several reports that the caspase/calpain system is activated in endotoxemia, which leads to cellular apoptosis and acute inflammatory processes. We aimed to determine the role of the caspase/calpain system in cell viability and regulation of fractalkine production in LPS-treated endothelial cells. METHODS: Human umbilical vein endothelial cells (HUVECs) were stimulated with 0.01–100 μg/mL of LPS to determine cell viability. The changes of CX3CL1 expression were compared in control, LPS (1 μg/mL)-, IL-1α (1 μg/mL)-, and IL-1β (1 μg/mL)-treated HUVECs. Cell viability and CX3CL1 production were compared with 50 μM of inhibitors of caspase-1, caspase-3, caspase-9, and calpain in LPS-treated HUVECs. RESULTS: Cell viability was significantly decreased from 1 to 100 μg/mL of LPS. Cell viability was significantly restored with inhibitors of caspase-1, caspase-3, caspase-9, and calpain in LPS-treated HUVECs. The expression of CX3CL1 was highest in IL-1β-treated HUVECs. CX3CL1 production was highly inhibited with a calpain inhibitor and significantly decreased with the individual inhibitors of caspase-1, caspase-3, and caspase-9. CONCLUSION: The caspase/calpain system is an important modulator of cell viability and CX3CL1 production in LPS-treated endothelial cells.

The Curves Exercise Suppresses Endotoxemia in Korean Women with Obesity

Obesity and metabolic syndrome is a worldwide pandemic and associated with high cardiovascular risk. Metabolic endotoxemia (ME) is thought to be an underlying molecular mechanism. It triggers toll-like receptor 4-mediated inflammatory adipokines and causes a chronic low grade inflammatory status, which results in cardiovascular risk increase. Exercise is the best nonpharmacological treatment to improve prognosis. In this study, we examined the circulating endotoxin level in Korean obese women and investigated effects of exercise on it. Women over body mass index (BMI) 25 kg/m2 participated in a resistance training exercise, Curves. At baseline and after 12 weeks exercise, tests including blood samples were taken. In Korean obese women, the fasting endotoxin was 1.45 ± 0.11 EU/mL. Ingestion of a high calorie meal led to a peak level after 2 hours (postprandial 2 hours [PP2]) and a significant rise over the 4 hours (postprandial 4 hours [PP4]) in it (1.78 ± 0.15 and 1.75 ± 0.14 EU/mL for PP2 and PP4, P < 0.05 vs. fasting). After exercise, BMI and hip circumference were reduced significantly. The total cholesterol (TC) at fasting, PP2 and PP4 were decreased significantly. All levels of circulating endotoxin at fasting, PP2 and PP4 showed reduction. But, the peak change was only significant (baseline vs. 12 weeks for PP2; 1.78 ± 0.15 vs. 1.48 ± 0.06 EU/mL, P < 0.05). We report the circulating endotoxin level in Korean obese women for the first time. Also, we establish that energy intake leads to endotoxemia and exercise suppresses the peak endotoxemia after meal. It suggests an impact for a better prognosis in obese women who follow regular exercise.

Laminar lesions in horses with systemic oxidative stress, committed by experimentally induced or naturally occurring gastrointestinal disorders / Lesões laminares em cavalos com estresse oxidativo sistêmico, acometidos por doenças gastrintestinais experimentalmente induzidas ou de ocorrência natural

Laminitis in horses can be associated with lesions in multiple organs secondary to sepsis. Twenty-one horses suffering from gastrointestinal disorders were used in the experiment; 7 horses with experimentally induced endotoxemia and intestinal ischaemia, and 14 horses suffering from naturally occurring colic syndrome. Tissue samples of lungs, liver, heart, brain, cerebellum and hoof laminar tissue were collected for histopathological and oxidative stress evaluation using nitrotyrosine and superoxide dismutase (SOD2) immunostaining. The horses were divided into two groups: the non-oxidative lesions group (NOLG), with 7 horses showing weak immunostaining in lungs, liver and kidney, and the oxidative lesions group (OLG), with 14 horses showing immunostaining indicating systemic oxidative stress in multiple organs. The horses from OLG showed increase of laminar lesions and SOD2 immunostaining in multiple organs when compared to the horses from the NOLG. No differences were found ln regard to laminar immunostaining by nitrotyrosine and SOD2 between experimental groups. It was concluded that systemic oxidative stress can be associated with the development of laminar lesions, and that the laminar tissue does not respond to oxidative stress with increase of SOD as occurs in other organs.(AU)

A laminite em equinos pode estar associada à lesão em múltiplos órgãos secundária a sepse. Foram utilizados 21 cavalos com afecções gastrintestinais, sendo sete com endotoxemia e isquemia intestinal induzidos experimentalmente, e 14 cavalos com síndrome cólica de origem natural. Amostras teciduais de pulmão, rim, fígado, coração, cérebro e cerebelo e de tecido laminar do casco foram coletadas para avaliação de lesão histopatológica e estresse oxidativo, pela imunomarcação de nitrotirosina e superóxido dismutase (SOD2). Os animais foram divididos em dois grupos: grupo sem lesão oxidativa (NOLG), com sete cavalos com fraca imunomarcação em pulmão, fígado e rim, e grupo lesão oxidativa (OLG), contendo 14 cavalos com imunomarcação indicando estresse oxidativo em múltiplos órgãos. Os cavalos do grupo OLG apresentaram aumento de lesões laminares e imunomarcação para SOD2 em múltiplos órgãos, quando comparados ao NOLG. Não houve diferença sobre a imunomarcação laminar para nitrotirosina e SOD2 entre os grupos experimentais. Conclui-se que o estresse oxidativo sistêmico está associado ao desenvolvimento de lesões laminares, e que o tecido laminar não responde ao estresse oxidativo com aumento de SOD como ocorre nos outros órgãos.(AU)

Protective Effects of S-Adenosylmethionine and Its Combinations With Taurine and/or Betaine Against Lipopolysaccharide or Polyinosinic-polycytidylic Acid-induced Acute Hepatotoxicity

BACKGROUND: Several mechanisms for the pathogenesis of many liver diseases are related with oxidative stress, endotoxins, and infections by many microorganisms. These can lead to chronic hepatitis, cirrhosis, and even liver cancer. The aim of this study was to evaluate the effects of S-adenosylmethionine (SAMe) and its combinations with taurine and/or betaine against hepatotoxicites induced by lipopolysaccharide (LPS) or polyinosinic-polycytidylic acid (polyI:C). METHODS: RAW 264.7 macrophage cells and seven-week-old male C57BL/6 mice were pretreated with SAMe (SAM or AdoMet), taurine, and/or betaine. In order to mimic hepatic injury like endotoxemia or viral infection, cells and mice were treated with LPS or polyI:C. Concentrations of glutathione (GSH), mRNA expressions of GSH synthesizing enzymes, and inflammatory markers were measured by biochemical assays and quantitative real-time PCR. RESULTS: In RAW 264.7 cells and mice, pretreatment of SAMe alone or SAMe with taurine and/or betaine attenuated the decrease in GSH levels and mRNA expressions of GSH synthesizing enzymes. In addition, pretreatment of SAMe with taurine and/or betaine prevented the excessive increase in inflammatory mediators produced by LPS or polyI:C treatment. CONCLUSIONS: Treatment with SAMe in combination with taurine and betaine, would have anti-oxidant functions in addition to anti-inflammatory action against bacterial and/or viral inflammation.

High-fat Diet Accelerates Intestinal Tumorigenesis Through Disrupting Intestinal Cell Membrane Integrity

BACKGROUND: Excess energy supply induces chronic low-grade inflammation in association with oxidative stress in various tissues including intestinal epithelium. The objective of this study was to investigate the effect of high-fat diet (HFD) on intestinal cell membrane integrity and intestinal tumorigenesis in ApcMin/+ mice. METHODS: Mice were fed with either normal diet (ND) or HFD for 12 weeks. The number of intestinal tumors were counted and biomarkers of endotoxemia, oxidative stress, and inflammation were determined. Changes in intestinal integrity was measured by fluorescein isothiocyanate (FITC)-dextran penetration and membrane gap junction protein expression. RESULTS: HFD group had significantly higher number of tumors compared to ND group (P < 0.05). Blood total antioxidant capacity was lower in HFD group, while colonic 8-hydroxy-2'-deoxyguanosine level, a marker of oxidative damage, was higher in HFD group compared to that of ND group (P < 0.05). The penetration of FITC-dextran was substantially increased in HFD group (P < 0.05) while the expressions of membrane gap junction proteins including zonula occludens-1, claudin-1, and occludin were lower in HFD group (P < 0.05) compared to those in ND group. Serum concentration of lipopolysaccharide (LPS) receptor (CD14) and colonic toll-like receptor 4 (a LPS receptor) mRNA expression were significantly higher in HFD group than in ND group (P < 0.05), suggesting that significant endotoxemia may occur in HFD group due to the increased membrane permeability. Serum interleukin-6 concentration and myeloperoxidase activity were also higher in HFD group compared to those of ND group (P < 0.05). CONCLUSIONS: HFD increases oxidative stress disrupting intestinal gap junction proteins, thereby accelerating membrane permeability endotoxemia, inflammation, and intestinal tumorigenesis.

Activating transcription factor-3 induction is involved in the anti-inflammatory action of berberine in RAW264.7 murine macrophages

Berberine is an isoquinoline alkaloid found in Rhizoma coptidis, and elicits anti-inflammatory effects through diverse mechanisms. Based on previous reports that activating transcription factor-3 (ATF-3) acts as a negative regulator of LPS signaling, the authors investigated the possible involvement of ATF-3 in the anti-inflammatory effects of berberine. It was found berberine concentration-dependently induced the expressions of ATF-3 at the mRNA and protein levels and concomitantly suppressed the LPS-induced productions of proinflammatory cytokines (TNF-α, IL-6, and IL-1β). In addition, ATF-3 knockdown abolished the inhibitory effects of berberine on LPS-induced proinflammatory cytokine production, and prevented the berberine-induced suppression of MAPK phosphorylation, but had little effect on AMPK phosphorylation. On the other hand, the effects of berberine, that is, ATF-3 induction, proinflammatory cytokine inhibition, and MAPK inactivation, were prevented by AMPK knockdown, suggesting ATF-3 induction occurs downstream of AMPK activation. The in vivo administration of berberine to mice with LPS-induced endotoxemia increased ATF-3 expression and AMPK phosphorylation in spleen and lung tissues, and concomitantly reduced the plasma and tissue levels of proinflammatory cytokines. These results suggest berberine has an anti-inflammatory effect on macrophages and that this effect is attributable, at least in part, to pathways involving AMPK activation and ATF-3 induction.

Microbiota e barreira intestinal: implicações para obesidade / Microbiota and intestinal barrier: implications for obesity

A epidemia da obesidade é considerada um importante problema de saúde pública na sociedade ocidental, pois ela se relaciona a comorbidades como síndrome metabólica, diabetes mellitus e hipertensão. A microbiota intestinal pode contribuir para o desenvolvimento da obesidade através do aumento da extração energética dos componentes da dieta, da lipogênese, da permeabilidade intestinal e da endotoxemia, mediada especialmente pelos lipopolissacarídeos. Estudos têm demonstrado diferenças na composição da microbiota intestinal entre indivíduos obesos e magros. Ao que parece, o aumento na proporção de Firmicutes em relação a Bacteroidetes parece estar presente na obesidade, podendo ser alterado à medida que ocorre perda de peso. Assim, o objetivo deste estudo foi revisar a literatura acerca dos mecanismos que relacionam a microbiota e a barreira intestinal ao desenvolvimento ou agravamento da obesidade (AU)

The epidemic of obesity is considered an important public health problem in the Western society and is related to comorbidities such as metabolic syndrome, diabetes mellitus, and hypertension. The intestinal microbiota may contribute to the development of obesity by increasing energy extraction from the dietary components, lipogenesis, intestinal permeability, and endotoxemia, especially mediated by lipopolysaccharides. Studies have demonstrated differences in composition of the intestinal microbiota between obese and lean individuals. Apparently, the increase in the proportion of Firmicutes in relation to Bacteroidetes seems to be present in obesity and can be changed during weight loss. The aim of this study was to review the mechanisms that relate microbiota and intestinal barrier to the development or worsening of obesity (AU)

Novas perspectivas para o papel de amilóide sérica A (SAA) na obesidade e resistência à insulina / New insights into the role of serum amyloid A (SAA) on obesity and insulin resistance

Chronic low-grade endotoxemia is an important player in obesity and insulin resistance associated to a high-fat diet (HFD). On the other hand, although it is known that intense endotoxemia and infection reduce appetite and induce intense catabolism, leading to weight loss during the acute inflammatory phase, the late effects of an intense endotoxemia were previously unexplored. Here we report that, besides the concurrent effects, multiple and intense endotoxemia causes long lasting biochemical alterations in the adipose tissue that intensify the harmful effects of a HFD. Mice submitted to multiple and severe endotoxemia had increased the adipose tissue expression of TLR-4, CD14 and SAA3, remaining altered after one week in recovery. When associated to a HFD, mice previously submitted to acute endotoxemia showed a more severe weight gain and impaired insulin sensitivity. Adopting the HFD as an obesogenic stimulus, we evaluated the participation of the protein serum amyloid A (SAA) in obesity development. Using a SAA-targeted antisense oligonucleotide, we observed that the depletion of SAA prevented metabolic alterations, endotoxin elevation, weight gain and insulin resistance in a diet-induced obesity protocol. Inadequate sleep is another important factor to be considered in the obesity epidemic. We found that sleep restriction (SR) causes biochemical and morphological alterations in mice adipose tissue. The levels of serum resistin and the adipose tissue mRNA expression of resistin, TNF-α and IL-6 were increased after SR. When associated to a HFD, mice previously submitted to SR gained more weight with increased macrophage infiltration in the epididymal adipose tissue, and insulin resistance. SAA is also part of the initial biochemical alterations caused by SR. It was observed that the expression of SAA in liver and adipose tissue is upregulated, with return to baseline when sleep is restored. Furthermore, 48 hours of total sleep restriction in healthy human volunteers also caused a serum elevation in SAA concentrations. Considering that SAA induces cell proliferation, we suggest that situations with an increase in SAA production and the consecutive preadipocyte proliferation would prime the adipose tissue to further adipocyte differentiation and hypertrophy. Furthermore, we suggest that SAA alter LPS signaling, possibly inhibiting its clearance. The mechanism associating inflammation and obesity is complex and encompass a diversity of factors; the inflammatory protein SAA may be one of them. In conclusion, our data describes the relationship between SAA, acute inflammation, sleep restriction and obesity

Endotoxemia crônica de baixo grau tem um importante papel na obesidade e resistência à insulina associada a uma ração hiperlipídica. Por outro lado, embora se saiba que a endotoxemia intensa e infecção reduzam o apetite e induzam a um intenso catabolismo, conduzindo a perda de peso durante a fase aguda da inflamação, os efeitos tardios da endotoxemia intensa nunca foram explorados. Aqui mostramos que, além dos efeitos correntes, a endotoxemia aguda provoca alterações bioquímicas prolongadas no tecido adiposo que intensificam os efeitos deletérios de uma ração hiperlipídica. Camundongos submetidos à endotoxemia aguda apresentaram aumento na expressão de TLR-4, CD14 e SAA3 no tecido adiposo, permanecendo alteradas após uma semana em recuperação. Quando associado a uma ração hiperlipídica, os camundongos previamente submetidos à endotoxemia aguda mostraram um ganho de peso mais pronunciado e uma maior resistência à insulina. Adotando a ração hiperlipídica como um estímulo obesogênico, foi avaliada a participação da proteína amilóide sérica A (SAA) no desenvolvimento da obesidade. Usando um oligonucleotídeo antisense anti-SAA, observamos que a depleção da SAA previne as alterações metabólicas, elevação de endotoxina, ganho de peso e resistência à insulina associadas a ração rica em gordura. O sono inadequado é outro fator importante a ser considerado na epidemia de obesidade. Descobrimos que a restrição do sono (SR) provoca alterações bioquímicas e morfológicas no tecido adiposo de camundongos. A concentração de resistina no soro e a expressão de mRNA no tecido adiposo de resistina, TNF-α e IL- 6 foram aumentadas após SR. Quando associado a uma ração hiperlipídica, os camundongos submetidos previamente à SR ganharam mais massa com aumento da infiltração de macrófagos no tecido adiposo epididimal, e resistência à insulina. SAA também faz parte das alterações bioquímicas iniciais provocadas pelo SR. Observou-se que a expressão de SAA no fígado e tecido adiposo é regulada positivamente, com retorno ao basal quando o sono é restaurado. Além disso, 48 horas de restrição de sono total em voluntários humanos saudáveis também causou uma elevação nas concentrações séricas de SAA. Considerando que SAA induz proliferação, sugerimos que situações onde ocorra aumento na produção de SAA e a consecutiva proliferação celular, o tecido adiposo se tornaria predisposto a futura diferenciação e hipertrofia. Além disso, sugerimos que SAA altera a sinalização de LPS, possivelmente inibindo sua depuração. O mecanismo de associação entre a inflamação e a obesidade é complexo e inclui uma diversidade de fatores; a proteína inflamatória SAA pode ser um deles. Em conclusão, nossos dados descrevem a relação entre SAA, inflamação aguda, restrição do sono e obesidade

Differential expression of miRNAs in myocardial tissues of rats with lipopolysaccharide- induced endotoxemia / 南方医科大学学报

<p><b>OBJECTIVE</b>To analyze the differential miRNA expression profile in the myocardium of rats with lipopolysaccharide (LPS)-induced endotoxemia and explore the role of miRNA in endotoxin-induced myocardial injury.</p><p><b>METHODS</b>Twenty male SD rats received intraperitoneal injection of 10 mg/kg LPS (n=10) or an equivalent amount of saline solution (n=10). At 24 h after LPS injection, the rats were sacrificed to detect myocardial expressions of TLR4, TNF-α and IL-1β using real-time PCR and for observing myocardial ultrastructures under transmission electron microscopy. The differentially expressed miRNA in the myocardium were detected using a miRNA array, and the common differentially expressed miRNAs were selected for verifying their actual expressions using real-time PCR.</p><p><b>RESULTS</b>TLR4, TNF-α and IL-1β were over-activated in the myocardium of LPS-treated rats, in which mitochondria swelling, structural damaged and cytoplasmic vacuoles were observed. In LPS-challenged rats, miR-194-3p, miR-344a-3p, miR-465-3p, miR-501-5p, miR-3596c, miR-185-3p, and miR-877 were found up-regulated significantly, whereas miR-208b-3p, miR-547-3p, miR-141-3p, miR-28-5p, and miR-3585-5p down-regulated in the myocardium.</p><p><b>CONCLUSION</b>Significant differential expression of the miRNAs occurs in the myocardium of LPS-treated rats, suggesting their involvement in endotoxin-induced myocardial injury.</p>

Probiotics as Complementary Treatment for Metabolic Disorders

Over the past decade, growing evidence has established the gut microbiota as one of the most important determinants of metabolic disorders such as obesity and type 2 diabetes. Indeed, obesogenic diet can drastically alter bacterial populations (i.e., dysbiosis) leading to activation of pro-inflammatory mechanisms and metabolic endotoxemia, therefore promoting insulin resistance and cardiometabolic disorders. To counteract these deleterious effects, probiotic strains have been developed with the aim of reshaping the microbiome to improve gut health. In this review, we focus on benefits of widely used probiotics describing their potential mechanisms of action, especially their ability to decrease metabolic endotoxemia by restoring the disrupted intestinal mucosal barrier. We also discuss the perspective of using new bacterial strains such as butyrate-producing bacteria and the mucolytic Akkermansia muciniphila, as well as the use of prebiotics to enhance the functionality of probiotics. Finally, this review introduces the notion of genetically engineered bacterial strains specifically developed to deliver anti-inflammatory molecules to the gut.