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Background: Pretreatment with low doses of LPS (lipopolysaccharide, bacterial endotoxin) reduces the pro-inflammatory response to a subsequent higher LPS dose, a phenomenon known as endotoxin tolerance. Moreover, hydrogen sulfide (H2S), an endogenous gaseous mediator (gasotransmitter) can exert anti-inflammatory effects. Here we investigated the potential role of H2S in the development of LPS tolerance. THP1 differentiated macrophages were pretreated with the H2S donor NaHS (1 mM) or the H2S biosynthesis inhibitor aminooxyacetic acid (AOAA, 1 mM)., Methods: To induce tolerance, cells were treated with a low concentration of LPS (0.5 μg/ml) for 4 or 24 h, and then treated with a high concentration of LPS (1 μg/ml) for 4 h or 24 h. In in vivo studies, male wild-type and CSE(-/-) mice were randomized to the following groups: Control (vehicle); Endotoxemic saline for 3 days before the induction of endotoxemia with 10 mg/kg LPS) mg/kg; Tolerant (LPS at 1 mg/kg for 3 days, followed LPS at 10 mg/kg). Animals were sacrificed after 4 or 12 h; plasma IL-6 and TNF-α levels were measured. Changes in histone H3 and H4 acetylation were analyzed by Western blotting., Results: LPS tolerance decreased pro-inflammatory cytokine production. AOAA did not affect the effect of tolerance on reducing cytokine production. Treatment of the cells with the H2S donor reduced cytokine production. Induction of the tolerance increased the acetylation of H3; AOAA reduced histone acetylation. H2S donation increased histone acetylation. Tolerance did not affect the responses to H2S with respect to histone acetylation., Conclusions: In conclusion, both LPS tolerance and H2S donation decrease LPS-induced cytokine production in vitro and modulate histone acetylation. However, endogenous, CSE-derived H2S does not appear to play a significant role in the development of LPS tolerance.

Aims: Changes in the gene expression are one of the molecular events involved in the Systemic of Inflammatory Response Syndrome during sepsis. The preconditioning with low doses of lipopolysaccharide (LPS) reduces the expression of pro-inflammatory genes leading to less tissue damage and better outcome. This hyporesponsive state called tolerance is associated to alterations in chromatin structure and nitric oxide (NO) production. In the current study, we demonstrated that tolerance induced by LPS was found to be NO-dependent and related to epigenetic changes., Main Methods: THP-1 cells were cultivated in RPMI medium (Control), submitted to tolerance (500ng/mL of LPS 24h before challenge with 1000ng/mL of LPS during 24h Tolerant group) and challenge (1000ng/mL of LPS during 24h Directly challenged group). The analyses performed were: cytokines production, histone acetyl transferases/histone deacetylases (HAT/HDAC) activity, nitrosylation of HDAC-2 and -3, expression of acetylated histones H3 and H4. HDAC and Nitric Oxide Synthases (NOS) activities were inhibited with 30mM trichostatin (TSA) and 100μM LNAME, respectively., Key Findings: Administration of low doses of LPS repressed the production of IL-6 and IL-10, however this effect was abolished with the inhibition of NOS activity and by TSA in the case of IL-10. Tolerance modulates the activity of HAT and, consequently, the acetylation of histones H3 and H4. Inhibition of NO decreases acetylation of Histones. The HDACs 2 and 3 were nitrosylated after the tolerance induction., Significance: The tolerance to LPS regulates the cytokine production by modulating chromatin structure and this event is NO dependent., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Creatine (Cr) is a dietary supplement that presents beneficial effects in experimental models of heart and brain ischemia and reperfusion (I/R) injury. It can improve adenosine 5'-triphosphate generation and reduce cell damage. This study evaluated the effects of Cr supplementation in a model of lung I/R., Methods: Forty male Wistar rats were divided into 4 groups: sham operated, Cr+sham, I/R, and Cr+I/R. We investigated the effects of 5 days of Cr supplementation (0.5 g/kg/day by gavage) before left pulmonary artery ischemia (90 minutes) and reperfusion (120 minutes) on pulmonary and systemic response., Results: Cr inhibited the I/R-induced increase in exhaled nitric oxide (p < 0.05), total cells (p < 0.01), and neutrophils (p < 0.001) in bronchoalveolar lavage fluid and in the systemic circulation (p < 0.001). The levels of interleukin-1β (p < 0.05), tissue damping, and tissue elastance (p < 0.05) were also minimized. Cr also inhibited pulmonary edema formation (total proteins in bronchoalveolar lavage fluid, p < 0.001; histologic edema index, p < 0.001) and neutrophils accumulation in lung tissue (p < 0.001). As possible mechanisms underlying Cr effects, we observed a reduced expression of caspase 3 (p < 0.05), reduced expression of Toll-like receptor (TLR) 4, and increased expression of TLR7 in lung tissue (p < 0.001)., Conclusions: Cr supplementation presents pulmonary and systemic protective effects in acute lung injury induced by I/R in rats. These beneficial effects seem to be related to the anti-inflammatory and anti-oxidant properties of Cr and modulation of TLRs., (Copyright © 2016 International Society for Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.)

Sepsis is a systemic inflammatory response to infection eliciting high mortality rate which is a serious health problem. Despite numerous studies seeking for therapeutic alternatives, the mechanisms involved in this disease remain elusive. In this study we evaluated the influence of cholesteryl ester transfer protein (CETP), a glycoprotein that promotes the transfer of lipids between lipoproteins, on the inflammatory response in mice. Human CETP transgenic mice were compared to control mice (wild type, WT) after polymicrobial sepsis induced by cecal ligation and puncture (CLP), aiming at investigating their survival rate and inflammatory profiles. Macrophages from the peritoneal cavity were stimulated with LPS in the presence or absence of recombinant CETP for phenotypic and functional studies. In comparison to WT mice, CETP mice showed higher survival rate, lower IL-6 plasma concentration, and decreased liver toll-like receptor 4 (TLR4) and acyloxyacyl hydrolase (AOAH) protein. Moreover, macrophages from WT mice to which recombinant human CETP was added decreased LPS uptake, TLR4 expression, NF-κB activation and IL-6 secretion. This raises the possibility for new therapeutic tools in sepsis while suggesting that lowering CETP by pharmacological inhibitors should be inconvenient in the context of sepsis and infectious diseases.

Acute respiratory distress syndrome (ARDS) is the most severe lung inflammatory manifestation and has no effective therapy nowadays. Sepsis is one of the main illnesses among ARDS causes. The use of fluid resuscitation is an important treatment for sepsis, but positive fluid balance may induce pulmonary injury. As an alternative, fluid resuscitation with hypertonic saline ((HS) NaCl 7.5%) has been described as a promising therapeutical agent in sepsis-induced ARDS by the diminished amount of fluid necessary. Thus, we evaluated the effect of hypertonic saline in the treatment of LPS-induced ARDS. We found that hypertonic saline (NaCl 7.5%) treatment in rat model of LPS-induced ARDS avoided pulmonary function worsening and inhibited type I collagen deposition. In addition, hypertonic saline prevented pulmonary injury by decreasing metalloproteinase 9 (MMP-9) activity in tissue. Focal adhesion kinase (FAK) activation was reduced in HS group as well as neutrophil infiltration, NOS2 expression and NO content. Our study shows that fluid resuscitation with hypertonic saline decreases the progression of LPS-induced ARDS due to inhibition of pulmonary remodeling that is observed when regular saline is used.

Acute lung injury (ALI) is a common complication associated with septic shock that directly influences the prognosis of sepsis patients. Currently, one of the main supportive treatment modalities for septic shock is fluid resuscitation. The use of hypertonic saline (HS: 7.5% NaCl) for fluid resuscitation has been described as a promising therapy in experimental models of sepsis-induced ALI, but it has failed to produce similar results in clinical practice. Thus, we compared experimental timing versus clinical timing effectiveness (i.e., early vs. late fluid resuscitation) after the inflammatory scenario was established in a rat model of bacterial lipopolysaccharide-induced ALI. We found that late fluid resuscitation with hypertonic saline (NaCl 7.5%) did not reduce the mortality rates of animals compared with the mortality late associated with early treatment. Late fluid resuscitation with both hypertonic and normal saline increased pulmonary inflammation, decreased pulmonary function, and induced pulmonary injury by elevating metalloproteinase-2 and metalloproteinase-9 activity and collagen deposition in the animals, unlike early treatment. The animals with lipopolysaccharide-induced ALI that received late resuscitation with any kind of fluids demonstrated aggravated pulmonary injury and respiratory function. Moreover, we showed that the therapeutic window for a beneficial effect of fluid resuscitation with hypertonic saline is very narrow.

Introduction: the inflammatory response caused by sepsis leads to metabolic changes, which may result in significant lean mass loss in septic patient. Because of this, when digestive tract is functional, nutritional therapy (NT) must be initiated within 48 hours of intensive treatment to reduce protein loss., Objective: to evaluate enteral nutritional therapy (ENT) in adult septic patients with exclusive ENT for ≥ 72 hours and length of stay ≥7 days in Intensive Care Unit and its relationship with clinical prognosis., Methods: we prospectively analyzed the adequacy of enteral nutrition administered, factors associated with non-conformity, gastrointestinal tolerance and outcome. Statistical tests of chi-square and Student's t as well as Mann-Whitney and Spearman and Pearson correlations (p < 0.05) were used. A multiple logistic regression model has been done by using the stepwise method to evaluate the association between predictors of clinical outcome., Results: 53 patients, 67.9% male and 52.8% elderly were enrolled in this study. The average time for starting ENT was 30 (23.5) hours, and 88.7% of patients achieved nutritional goal in 48 hours. The mean volume delivered in relation to prescribed was 78.9%. When the sample was stratified according to administered/ prescribed calories, patients who received < 80% had a higher mortality rate (p = 0.001) and the caloric intake ≥ 80% was the determining factor in patients' clinical prognosis (p = 0.021)., Conclusion: septic patients received early enteral nutrition. The nutritional goal and the mean volume delivered in relation to the prescribed volume meet the intensive care guidelines. The nutritional support was associated with clinical outcome, and caloric intake ≥ 80% determining the clinical prognosis. The pauses caused by reflux were significant in the group of patients who died and it may be related to disease severity., (Copyright AULA MEDICA EDICIONES 2014. Published by AULA MEDICA. All rights reserved.)

Purpose: To quantify the physical activity in daily life (PADL), muscle strength, and exercise capacity in the short and medium term in survivors of severe sepsis and septic shock., Methods: Prospective cohort study with a follow-up from hospital admission to 3 months after hospital discharge. Seventy-two patients admitted to the ICU for severe sepsis or septic shock and a control group of healthy sedentary subjects (n = 50) were enrolled. All patients had their PADL quantified by an accelerometer during their hospital stay and 3 months after. Exercise capacity and handgrip and quadriceps muscle strength were also evaluated., Results: During hospitalization, patients spent the majority of their time inactive in a lying or sitting position (90 ± 34% of daily time). Physical inactivity was partially reduced 3 months after (58 ± 20% of daily time). However, the time patients spent walking was only 63% of the time reported for healthy subjects. Patients also showed lower movement intensity when compared with controls (2.1 ± 0.3 vs 2.5 ± 0.4 m/s(2)). At hospital discharge, muscle strength and exercise capacity were approximately 54% of the predicted value, and these parameters showed significant increase in patients 3 months after (70% of predicted value). Multivariable analysis demonstrated that the use of systemic corticosteroids and hospitalization time negatively influenced quadriceps strength and exercise capacity at the time of hospital discharge., Conclusion: Our results suggest that survivors of sepsis admitted to the ICU have a substantial reduction in physical activity, exercise capacity, and muscle strength compared to healthy subjects that persist even 3 months after hospital discharge.

Hydrogen sulfide (H2S) is an endogenous gaseous biological mediator, which regulates, among others, the oxidative balance of cells under normal physiological conditions, as well as in various diseases. Several previous studies have reported that H2S attenuates inflammatory mediator production. In this study, we investigated the role of H2S in chromatin modulation in an in vitro model of lipopolysaccharide (LPS)-induced inflammation and evaluated its effects on inflammatory cytokine production. Tamm-Horsfall protein 1 (THP-1) differentiated macrophages were pre-treated with sodium hydrosulfide (NaHS) (an H2S donor) at 0.01, 0.1, 0.5 or 1 mM for 30 min. To stimulate cytokine production, the cells were challenged with bacterial LPS (1 µg/ml) for 1, 4, 8 or 24 h. Histone H3 acetylation was analyzed by chromatin immunoprecipitation (ChIP), cytokine production was measured by ELISA and histone deacetylase (HDAC) activity was analyzed using a standard biochemical assay. H2S inhibited the production of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in a concentration-dependent manner; it was most effective at the two highest concentrations used. This effect was associated with a decrease in histone H3 acetylation at the IL-6 and TNF-α promoters in the cells exposed to H2S or H2S + LPS. The findings of the present study suggest that H2S suppresses histone acetylation, which, in turn, inhibits chromatin openness, leading to a decrease in the gene transcription of various pro-inflammatory cytokines. Therefore, this mechanism may contribute to the previously demonstrated anti-inflammatory effects of H2S and various H2S donors.

Objective: To investigate the role of coronary driving pressure (CDP) in myocardial microcirculatory blood flow during sepsis. We hypothesised that in septic shock there is an impaired autoregulation of microcirculation, and blood flow is totally dependent on CDP. We analysed the effect of lipopolysaccharide (LPS)-induced shock on myocardial microcirculation, separating subendocardial and epicardial areas. We then studied the effect of CDP increases using noradrenaline (NOR) or metaraminol (Aramine [ARA]) on myocardial microcirculation and function, and we analysed the effect of volume infusion on CDP and myocardial function., Design and Setting: Endotoxaemia was induced in male Wistar rats by an intraperitoneal injection of LPS 10 mg/kg. Animals were divided into a control (CT) group, an LPS-injected group, and an LPS-injected group treated with saline fluid, NOR or ARA., Main Outcome Measures: Ninety minutes later, a haemodynamic evaluation was performed. NOR or ARA were used to manage the mean arterial pressure (MAP) and CDP, and we inserted a catheter into the left ventricle to measure cardiac parameters. To measure blood flow in the myocardium and other organs, microspheres were introduced into the left ventricle using an infusion pump., Results: After LPS treatment, left ventricular (LV) systolic function (dP/dt max) and diastolic function (dP/dt min) decreased by 34% and 15%, respectively, and load-independent indices (LV contractility in ejection phase and dP/dt max÷end-diastolic volume) were reduced. The CDP was also reduced (by 58%) in the endotoxaemic rats. Myocardial blood flow was reduced (by 80%) in animals with an MAP≤65 mmHg. NOR increased the CDP (LPS, 38 mmHg [SEM, 2 mmHg]; LPS+NOR, 59 mmHg [SEM, 3 mmHg]) and microcirculatory perfusion (LPS, 2 mL/min/g tissue [SEM, 0.6 mL/min/g]; LPS+NOR, 6.2 mL/min/g [SEM, 0.8 mL/min/g]). ARA was also effective in improve microcirculation but saline volume infusion was ineffective in improving CDP or myocardial function. CDP showed a significant correlation with subendocardial blood flow., Conclusions: Myocardial blood flow in the LV subendocardium and the right ventricle decreases in endotoxaemic rats. Increasing CDP improves myocardial blood flow and function. Thus, in endotoxaemia, microcirculatory blood flow is pressure dependent, suggesting that it may be beneficial to treat patients with sepsis using a higher CDP.

Objectives: Muscle injury due to crushing (muscle compression injury) is associated with systemic manifestations known as crush syndrome. A systemic inflammatory reaction may also be triggered by isolated muscle injury. The aim of this study was to investigate the plasma levels of interleukins (IL) 1, 6 and 10 and tumor necrosis factor alpha (TNF-α), which are markers for possible systemic inflammatory reactions, after isolated muscle injury resulting from lower-limb compression in rats., Methods: Male Wistar rats were subjected to 1 h of compression of their lower limbs by means of a rubber band. The plasma levels of IL 1, 6 and 10 and TNF-α were measured 1, 2 and 4 h after the rats were released from compression., Results: The plasma levels of IL 10 decreased in relation to those of the other groups, with a statistically significant difference (p < 0.05). The method used did not detect the presence of IL 1, IL 6 or TNF-α., Conclusion: Our results demonstrated that the changes in plasma levels of IL 10 that were found may have been a sign of the presence of circulating interleukins in this model of lower-limb compression in rats.

Sepsis impairs the autoregulation of myocardial microcirculatory blood flow, but whether this impairment is correlated with myocardial remodeling is unknown. This study investigated the role of coronary driving pressure (CDP) as a determinant of microcirculatory blood flow and myocardial fibrosis in endotoxemia and sepsis. The study is composed of two parts: a prospective experimental study and an observational clinical study. The experimental study was performed on male Wistar rats weighing 300 to 320 g. Endotoxemia was induced in rats by lipopolysaccharide (LPS) injection (10 mg·kg intraperitoneally). Hemodynamic evaluation was performed 1.5 to 24 h after LPS injection by measuring the mean arterial pressure, CDP, left ventricular end-diastolic pressure, dP/dtmax, and dP/dtmin. Microspheres were also infused into the left ventricle to measure myocardial blood flow, and myocardial tissue was histologically assessed to analyze collagen deposition. The CDP, mean arterial pressure, and myocardial blood flow were reduced by 55%, 30%, and 70%, respectively, in rats 1.5 h after LPS injection compared with phosphate buffer saline injection (P < 0.05). The CDP was significantly correlated with subendocardial blood flow (r = 0.73) and fibrosis (r = 0.8). Left ventricular function was significantly impaired in the LPS-treated rats, as demonstrated by dP/dtmax (6,155 ± 455 vs. 3,746 ± 406 mmHg·s, baseline vs. LPS; P < 0.05) and dP/dtmin (-5,858 ± 236 vs. -3,516 ± 436 mmHg·s, baseline vs. LPS; P < 0.05). The clinical study was performed on 28 patients with septic shock analyzed for CDP. The CDP data and histological slices were collected from septic patients. In addition, the clinical data demonstrated fibrosis and 45% CDP reduction in nonsurvivors compared with survivors. In conclusion, the left ventricular subendocardial blood flow was positively correlated with CDP, and higher CDP was negatively correlated with myocardial collagen deposition. Thus, early reductions in myocardial blood flow and CDP facilitate late myocardial fibrosis in rats and likely in humans.

Maple syrup urine disease (MSUD) is an inborn error of metabolism caused by a severe deficiency in the activity of the branched-chain α-keto acid dehydrogenase complex, leading to accumulation of the branched-chain amino acids (BCAA) leucine, isoleucine, and valine. Infections have a significant role in precipitating acute metabolic decompensation in patients with MSUD; however, the mechanisms underlying the neurotoxicity in this disorder are poorly understood. In this study, we subjected rats to the coadministration of lipopolysaccharide (LPS), which is a major component of gram-negative bacteria cell walls, and high concentrations of BCAA (H-BCAA) to determine their effects on the permeability of the blood-brain barrier (BBB) and on the levels of matrix metalloproteinases (MMP-2 and MMP-9). Our results demonstrated that the coadministration of H-BCAA and LPS causes breakdown of the BBB and increases the levels of MMP-2 and MMP-9 in the hippocampus of these rats. On the other hand, examination of the cerebral cortex of the 10- and 30-day-old rats revealed a significant difference in Evan's Blue content after coadministration of H-BCAA and LPS, as MMP-9 levels only increased in the cerebral cortex of the 10-day-old rats. In conclusion, these results suggest that the inflammatory process associated with high levels of BCAA causes BBB breakdown. Thus, we suggest that BBB breakdown is relevant to the perpetuation of brain inflammation and may be related to the brain dysfunction observed in MSUD patients.

The objective of this randomized animal study and laboratory investigation was to investigate whether lipopolysaccharide tolerance redirects neutrophil migration between organs. Male BALB/c mice received subcutaneous injections of lipopolysaccharide (1 mg/kg) for 5 days, followed by cecal ligation and puncture (CLP). Cytokines and adhesion molecules were measured after tolerance and CLP challenge. Increased numbers of neutrophils were observed in the peritoneal cavity of tolerant mice, which was associated with increased levels of adhesion molecules and chemokines. In contrast, nontolerant mice accumulated higher numbers of neutrophils in the lungs compared with those in the peritoneal cavity. Neutrophil function accessed by hydrogen peroxide production from neutrophils recovered from peritoneal cavity showed that tolerance increased the capacity to produce hydrogen peroxide. Mortality was reduced in tolerant animals. This study demonstrated that tolerance reduces leukocyte accumulation in the lung after CLP by redirecting neutrophils to the site of infection.

Aim: To investigate pro-atherosclerotic markers (endothelial dysfunction and inflammation) in patients one year after liver transplantation., Methods: Forty-four consecutive liver transplant (LT) outpatients who were admitted between August 2009 and July 2010, were followed-up by for 1 year, exhibited no evidences of infection or rejection, all of them underwent tacrolimus-based immunosuppressive regimens were consecutively enrolled. Inflammatory cytokines (TNFα, IFNγ, IL-8, and IL-10), endothelial biomarkers (sVCAM-1, sICAM-1, MPO, adiponectin, PAI-1, SAP, SAA, E-selectin, and MMP-9), high sensitive C-reactive protein, and Framingham risk score (FRS) were assessed. The anthropometric data, aminotransferases, metabolic syndrome features, glucose and lipid profiles, and insulin resistance data were also collected. The LT recipients were compared to 22 biopsy-proven non-alcoholic steatohepatitis (NASH) patients and 20 healthy controls (non-obese, non-diabetics, and non-dyslipidemic)., Results: The LT recipients had significantly younger ages and lower body mass indices, aminotransferases, fasting glucose and insulin levels, glucose homeostasis model and metabolic syndrome features than the NASH patients. Classic cardiovascular risk markers, such as Hs-CRP and FRS [2.0 (1.0-8.75)], were lower in the LT patients compared to those observed in the NASH patients (P = 0.009). In contrast, the LT recipients and NASH patients had similar inflammatory and endothelial serum markers compared to the controls (pg/mL): lower IL-10 levels (32.3 and 32.3 vs 62.5, respectively, P = 0.019) and higher IFNγ (626.1 and 411.9 vs 67.9, respectively, P < 0.001), E-selectin (48.5 and 90.03 vs 35.7, respectively, P < 0.001), sVCAM-1 (1820.6 and 1692.4 vs 1167.2, respectively, P < 0.001), and sICAM-1 (230.3 and 259.7 vs 152.9, respectively, P = 0.015) levels., Conclusion: Non-obese LT recipients have similar pro-atherosclerotic serum profiles after a short 1-year follow-up period compared to NASH patients, suggesting a high risk of atherosclerosis in this population.

Endotoxin tolerance is defined as a reduced capacity of the host to respond to LPS activation following a first exposure to this stimulus. It affects all leukocytes and regarding macrophages, most studies focus on the reduced ability of these cells to secrete pro-inflammatory cytokines. Therefore, we evaluated other macrophages functions (fungicidal capacity, reactive oxygen species production and antigen presentation) in cells from tolerant mice. We have performed a tolerance model in our laboratory that does not stimulate directly the place from where the cells will be removed (peritoneal cavity). Mouse received subcutaneous injections of LPS in the scruff for 5 days and we analyze the capacity of peritoneal macrophages to phagocyte using three different receptors: Fc, C3b and mannose receptors. We found a reduction in the phagocytosis of erythrocytes and Candida albicans related to the Fc and mannose receptors. These differences can be due to a macrophage reprogramming, as demonstrated by altered expression of cytokines and chemokines. Despite this reduction in phagocytosis capacity, macrophages from tolerant animals exhibited enhanced hydrogen peroxide production and expression of antigen presentation molecules, suggesting that their ability to combat an infection is improved. In summary, our data indicates that LPS tolerance drives macrophages from a predominant release of proinflammatory mediators that amplify inflammation and host damage toward a better killing and antigen presentation state., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Objective: We investigated the effect of two different saline solutions on the mechanisms of injury after intestinal ischemia: oxidative stress and inflammatory responses., Methods: Wistar rats underwent transient superior mesenteric artery occlusion and were studied for 6 hours after reperfusion. After randomization, the animals were divided into four groups: Sham; Hypertonic Saline, in which they received infusion of 4 mL/kg body weight of 7.5% hypertonic saline; Saline, in which they received infusion of 33 mL/kg body weight of 0.9% saline; and Non Treatment. The infusion was performed immediately prior to the reperfusion. The plasma concentrations of interleukin 6 and interleukin 10 were measured. Tissue samples (lung, liver, and intestine) were collected for malondialdehyde, myeloperoxidase, and interleukin measurements., Results: The animals that received infusions (Hypertonic Saline and Saline) showed lower levels of tissue malondialdehyde, myeloperoxidase, interleukin 6, and interleukin 10 compared with the Non Treatment group. The plasma concentrations of interleukin 6 and interleukin 10 were higher in the animals treated with 7.5% hypertonic saline compared with Saline and Non Treatment groups., Conclusion: In this model of transient intestinal ischemia, the adequate maintenance of intravascular volume decreased oxidative stress and the synthesis of inflammatory markers. Both 7.5% Hypertonic Saline and Saline attenuated the deleterious effects observed after intestinal ischemia.

Objective: To determine the factors that influence the adequacy of enteral nutritional therapy in an intensive care unit., Methods: This prospective observational study was conducted in an intensive care unit between 2010 and 2012. Patients >18 years of age underwent exclusive enteral nutritional therapy for ≥72 hours. The energy and protein requirements were calculated according to the ICU protocols. The data regarding enteral nutrition, the causes of non-compliance, and the biochemical test results were collected daily., Results: Ninety-three patients admitted to the intensive care unit were evaluated. Among these patients, 82% underwent early enteral nutritional therapy, and 80% reached the nutritional goal in <36 hours. In addition, 81.6%±15.4% of the enteral nutrition volume was infused, with an adequacy of 82.2%±16.0% for calories, 82.2%±15.9% for proteins, and a mean energy balance of -289.9±277.1 kcal/day. A negative correlation of C-reactive protein with the volume infused and the energy and protein balance was observed. In contrast, a positive correlation was found between C-reactive protein and the time required to reach nutritional goals. Extubation was the main cause for interrupting the enteral nutritional therapy (29.9% of the interruption hours), and the patients >60 years of age exhibited a lower percentage of recovery of the oral route compared with the younger patients (p=0.014)., Conclusion: Early enteral nutritional therapy and the adequacy for both energy and protein of the nutritional volume infused were in accordance with the established guidelines. Possible inadequacies of energy and protein balance appeared to be associated with an acute inflammatory response, which was characterized by elevated C-reactive protein levels. The main cause of interruption of the enteral nutritional therapy was the time spent in extubation.

The effects of hypertonic saline solution (HSS) have been shown in several animal models of ischemia and shock. Literature has shown potential benefits of HSS modulating inflammatory response after sepsis in an animal model. We studied the HSS effects in sepsis through cecal ligation and puncture (CLP) in Balb-C mice. Groups studied: 1- CLP without treatment (CLP-C); 2- CLP treated with normal saline solution NaCl 0.9% - 34 ml/Kg (CLP-S); 3- CLP treated with HSS NaCl 7.5% - 4 ml/Kg (CLP-H); and 4- group (Basal) without no CLP or treatment. Volume infusion was always applied 30 min after CLP. Lung and peritoneal lavage were harvested after 6h and 24h of CLP to analyze cytokines amount, oxide nitric, lipid peroxidation and neutrophil infiltration. Neutrophil infiltration, ICAM-1, CXCR-2, and CXCL-1 in lung were reduced by HSS (CLP-H) compared to CLP-C or CLP-S. Neutrophil in peritoneal lavage was increased in 24h with HSS (CLP-H) compared to CLP and CLP-S. Peritoneal CXCR-2 was increased in CLP-C and CLP-S but presented a lower increase with HSS (CLP-H) after 6 hours. GRK-2 presented difference among the groups at 24 h, showing a profile similar to neutrophil infiltration. Pro-inflammatory cytokines (TNF-α and IL-6) were reduced by HSS treatment; CLP-S increased TNF-α. IL-10 was increased in lung tissue by the HSS treatment. The oxidative stress (TBARS and nitric oxide biochemistry markers) was reduced with HSS. Animal survival was 33.3% in CLP-C group, 46.6% in CLP-S group and 60% in the CLP-H group after the sixth day. The HSS protects the animal against sepsis. Our results suggest that the volume replacement modulate pro and anti-inflammatory mediators of an inflammatory response, but HSS presented a more effective and potent effect.

Inflammation is powerful response to destroy invading organisms, and an exaggerated response can lead to death of the host. Macrophages secrete mediators that activated circulating neutrophils leading to its migration into infectious site. Recently, it has been shown that lymphocytes have an action modulating the early phase of inflammatory response. In this article, we analyze the role of B1 in the inflammatory response of different origins and finally focus attention on sepsis. B lymphocyte deficiency has been linked to acute infection presumably owing to the lack of an adaptive immune response to effectively clear pathogens. Individuals with X-linked agammaglobulinemia (XLA) present B1 lymphocyte deficiency caused by mutations in the Bruton tyrosine kinase (Btk). Some data show that B1 cells might contribute to susceptibility in experimental paracoccidioidomycosis. On the other hand, B1 cells are shown to be detrimental in other mouse models of microbial infection, such as experimental Chagas disease, leishmaniasis, and Staphylococcus aureus-induced arthritis. B1 cell plays a protective role in the host of the effects of endotoxemia. In a murine model of endotoxemia by lipopolysaccharide, B1 cell participates in both interleukin 10 and immunoglobulin M secretion with a consequent reduction in mortality.

The role of hepatitis C virus (HCV) in the pathogenesis of atherosclerosis and cardiovascular events is unclear. The aim of this study was to evaluate the direct effect of HCV on cardiovascular risk and correlate it with pro and anti-inflammatory cytokines in patients with HCV. HCV monoinfected patients, genotype 1, naive, non-obese (BMI<30) and non-diabetics were included and compared to controls (blood donors). Patients with prior diagnosis of cardiovascular diseases, hypertension, chronic renal failure, cancer and chronic use of lipid-lowering drugs or immunosuppressants were excluded. Age, BMI, systolic blood pressure (SBP) and diastolic (DBP), fasting glucose and lipid levels were determined. Serum cytokines (IL-6, IL-10 and TNF-α) and Framingham score were also evaluated. 62 HCV patients, 34 (54.8%) were males and none of them was smoking. The Framingham scores (median and 25th and 75th percentiles) were 12% (6.5-14%), showing an intermediate cardiovascular risk in patients with HCV. There was significant direct correlation between Framingham and total cholesterol (p=0.043) and DBP (p=0.007). HDL-C (p=0.002) was inversely correlated with the Framingham score. HCV patients had higher levels of proinflammatory cytokines (IL-6 and TNF-α) compared to controls (p<0.0001) and the relation of proinflammatory/anti-inflammatory TNF-α/IL10 and IL-6/IL10 were higher in HCV patients (p<0.01). The Framingham score was directly correlated to IL-6 and TNF-α, but differences were not statistically significant. Patients with HCV monoinfected, nonobese, naïve and non diabetic have an intermediate cardiovascular risk, as measured by the Framingham score and high levels of proinflammatory cytokines (IL-6 and TNF)., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Muscle crush injury is associated with systemic manifestations known as crush syndrome. A systemic inflammatory response syndrome may be triggered by isolated crush injury. Using myeloperoxidase (MPO) activity and plasma fatty acid composition, we investigated the inflammatory response in distant organs after isolated limb compression in rats. Male Wistar rats were submitted to 1h of hind limb compression by a latex ribbon. Myeloperoxidase activity was measured in muscle, liver, and lung at progressive times (1, 2 or 4h) after bandage release. Plasma fatty acid composition was evaluated as an indirect measure of oxidative stress. The liver and hind limb muscles showed a transient increase in MPO activity. Pulmonary MPO activity, otherwise, increased progressively throughout the study and reached statistically significant values at 4h when compared to all other groups (p<0.05). Plasma levels of unsaturated fatty acids decreased gradually after decompression (p<0.05 compared to controls after 4h). Blunt traumatic muscle compression was associated with rapid and transient muscle and liver inflammatory cell infiltration but otherwise, polymorphonuclear cells showed progressive aggregation in lungs. The plasmatic unsaturated index decreased throughout the 4h after muscle release. We demonstrated that limb compression was associated with oxidative stress and distant inflammatory responses. Progressive inflammatory cell infiltration in lungs could be related with the delayed systemic adverse responses found after crush injury., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Objective: Volume replacement in septic patients improves hemodynamic stability. This effect can reduce the inflammatory response. The objective of this study was to evaluate the effect of 7.5% hypertonic saline solution versus 0.9% normal saline solution for volume replacement during an inflammatory response in endotoxemic rats., Methods: We measured cytokines (serum and gut), nitrite, and lipid peroxidation (TBARS) as indicators of oxidative stress in the gut. Rats were divided into four groups: control group (C) that did not receive lipopolysaccharide; lipopolysaccharide injection without treatment (LPS); lipopolysaccharide injection with saline treatment (LPS +S); and lipopolysaccharide injection with hypertonic saline treatment (LPS +H). Serum and intestine were collected. Measurements were taken at 1.5, 8, and 24 h after lipopolysaccharide administration., Results: Of the four groups, the LPS +H group had the highest survival rate. Hypertonic saline solution treatment led to lower levels of IL-6, IL-10, nitric oxide, and thiobarbituric acid reactive substances compared to 0.9% normal saline. In addition, hypertonic saline treatment resulted in a lower mortality compared to 0.9% normal saline treatment in endotoxemic rats. Volume replacement reduced levels of inflammatory mediators in the plasma and gut., Conclusion: Hypertonic saline treatment reduced mortality and lowered levels of inflammatory mediators in endotoxemic rats. Hypertonic saline also has the advantage of requiring less volume replacement.

Objective: Liver failure can occur as a consequence of the systemic inflammation after acute pancreatitis. We assessed the effect of volume repositioning with hypertonic saline solution or normal saline on hepatic cytokine production and the expression of heat-shock proteins and apoptotic proteins after acute pancreatitis., Methods: Wistar rats were divided in four groups: C - control animals that were not subjected to insult or treatment; NT - animals that were subjected to acute pancreatitis and received no treatment; normal saline - animals that were subjected to acute pancreatitis and received normal saline (NaCl 0.9%); and HS - animals that were subjected to acute pancreatitis and received hypertonic saline solution (NaCl 7.5%). Acute pancreatitis was induced by retrograde transduodenal infusion of 2.5% sodium taurocholate into the pancreatic duct. At 4, 12 and 24 h following acute pancreatitis induction, TNF-alpha, IL-1-beta, IL-6 and IL-10, caspase-2 and -7, Apaf-1, AIF and HSP60 and 90 were analyzed in the liver., Results: Casp2 decreased in the normal saline and hypertonic saline groups (p<0.05 versus. C) at 12 h. Apaf-1, AIF and HSP90 remained unchanged. At 4 h, Casp7 increased in the NT group (p<0.01 versus C), although it remained at the baseline levels in the reperfused groups. HSP60 increased in all of the groups at 4 h (p< 0.001 vs. C). However, the hypertonic saline group showed lower expression of HSP60 than the normal saline group (p<0.05). Hypertonic saline solution maintained the production of cytokines at normal levels. Volume reperfusion with normal or hypertonic saline significantly modulated the expression of Casp7., Conclusion: Volume replacement with hypertonic or normal saline was effective in reducing caspase 7. However, only hypertonic solution was capable of regulating cytokine production and HSP60 expression at all time points.

In sepsis, toll-like receptor (TLR)-4 modulates the migration of neutrophils to infectious foci, favoring bacteremia and mortality. In experimental sepsis, organ dysfunction and cytokines released by activated macrophages can be reduced by gastrin-releasing peptide (GRP) receptor (GRPR) antagonist RC-3095. Here we report a link between GRPR and TLR-4 in experimental models and in sepsis patients. RAW 264.7 culture cells were exposed to lipopolysaccharide (LPS) or tumor necrosis factor (TNF)-α and RC-3095 (10 ng/mL). Male Wistar rats were subjected to cecal ligation and puncture (CLP), and RC-3095 was administered (3 mg/kg, subcutaneously); after 6 h, we removed the blood, bronchoalveolar lavage, peritoneal lavage and lung. Human patients with a clinical diagnosis of sepsis received a continuous infusion with RC-3095 (3 mg/kg, intravenous) over a period of 12 h, and plasma was collected before and after RC-3095 administration and, in a different set of patients with systemic inflammatory response syndrome (SIRS) or sepsis, GRP plasma levels were determined. RC-3095 inhibited TLR-4, extracellular-signal-related kinase (ERK)-1/2, Jun NH(2)-terminal kinase (JNK) and Akt and decreased activation of activator protein 1 (AP-1), nuclear factor (NF)-κB and interleukin (IL)-6 in macrophages stimulated by LPS. It also decreased IL-6 release from macrophages stimulated by TNF-α. RC-3095 treatment in CLP rats decreased lung TLR-4, reduced the migration of cells to the lung and reduced systemic cytokines and bacterial dissemination. Patients with sepsis and systemic inflammatory response syndrome have elevated plasma levels of GRP, which associates with clinical outcome in the sepsis patients. These findings highlight the role of GRPR signaling in sepsis outcome and the beneficial action of GRPR antagonists in controlling the inflammatory response in sepsis through a mechanism involving at least inhibition of TLR-4 signaling.

The development of atherosclerosis and the inflammatory response were investigated in LDLr-KO mice on three high-fat diets (40% energy as fat) for 16 weeks: trans (TRANS), saturated (SAFA) or ω-6 polyunsaturated (PUFA) fats. The following parameters were measured: plasma lipids, aortic root total cholesterol (TC), lesion area (Oil Red-O), ABCA1 content and macrophage infiltration (immunohistochemistry), collagen content (Picrosirius-red) and co-localization of ABCA1 and macrophage (confocal microscopy) besides the plasma inflammatory markers (IL-6, TNF-α) and the macrophage inflammatory response to lipopolysaccharide from Escherichia coli (LPS). As expected, plasma TC and TG concentrations were lower on the PUFA diet than on TRANS or SAFA diets. Aortic intima macrophage infiltration, ABCA1 content, and lesion area on PUFA group were lower compared to TRANS and SAFA groups. Macrophages and ABCA1 markers did not co-localize in the atherosclerotic plaque, suggesting that different cell types were responsible for the ABCA1 expression in plaques. Compared to PUFA, TRANS and SAFA presented higher collagen content and necrotic cores in atherosclerotic plaques. In the artery wall, TC was lower on PUFA compared to TRANS group; free cholesterol was lower on PUFA compared to TRANS and SAFA; cholesteryl ester concentration did not vary amongst the groups. Plasma TNF-α concentration on PUFA and TRANS-fed mice was higher compared to SAFA. No difference was observed in IL-6 concentration amongst groups. Regarding the macrophage inflammatory response to LPS, TRANS and PUFA presented higher culture medium concentrations of IL-6 and TNF-α as compared to SAFA. The PUFA group showed the lowest amount of the anti-inflammatory marker IL-10 compared to TRANS and SAFA groups. In conclusion, PUFA intake prevented atherogenesis, even in a pro-inflammatory condition., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Despite significant advances in the care of critically ill patients, acute lung injury continues to be a complex problem with high mortality. The present study was designed to characterize early lipopolysaccharide (LPS)-induced pulmonary injury and small interfering RNA targeting focal adhesion kinase (FAK) as a possible therapeutic tool in the septic lung remodeling process. Male Wistar rats were assigned into endotoxemic group and control group. Total collagen deposition was performed 8, 16, and 24 h after LPS injection. Focal adhesion kinase expression, interstitial and vascular collagen deposition, and pulmonary mechanics were analyzed at 24 h. Intravenous injection of small interfering RNA targeting FAK was used to silence expression of the kinase in pulmonary tissue. Focal adhesion kinase, total collagen deposition, and pulmonary mechanics showed increased in LPS group. Types I, III, and V collagen showed increase in pulmonary parenchyma, but only type V increased in vessels 24 h after LPS injection. Focal adhesion kinase silencing prevented lung remodeling in pulmonary parenchyma at 24 h. In conclusion, LPS induced a precocious and important lung remodeling. There was fibrotic response in the lung characterized by increased amount in total and specific-type collagen. These data may explain the frequent clinical presentation during sepsis of reduced lung compliance, oxygen diffusion, and pulmonary hypertension. The fact that FAK silencing was protective against lung collagen deposition underscores the therapeutic potential of FAK targeting by small interfering RNA.

In the past two years we observed several changes in the diagnostic and therapeutic approach of patients with acute heart failure (acute HF), which led us to the need of performing a summary update of the II Brazilian Guidelines on Acute Heart Failure 2009. In the diagnostic evaluation, the diagnostic flowchart was simplified and the role of clinical assessment and echocardiography was enhanced. In the clinical-hemodynamic evaluation on admission, the hemodynamic echocardiography gained prominence as an aid to define this condition in patients with acute HF in the emergency room. In the prognostic evaluation, the role of biomarkers was better established and the criteria and prognostic value of the cardiorenal syndrome was better defined. The therapeutic approach flowcharts were revised, and are now simpler and more objective. Among the advances in drug therapy, the safety and importance of the maintenance or introduction of beta-blockers in the admission treatment are highlighted. Anticoagulation, according to new evidence, gained a wider range of indications. The presentation hemodynamic models of acute pulmonary edema were well established, with their different therapeutic approaches, as well as new levels of indication and evidence. In the surgical treatment of acute HF, CABG, the approach to mechanical lesions and heart transplantation were reviewed and updated. This update strengthens the II Brazilian Guidelines on Acute Heart Failure to keep it updated and refreshed. All clinical cardiologists who deal with patients with acute HF will find, in the guidelines and its summary, important tools to help them with the clinical practice for better diagnosis and treatment of their patients.

Purpose: Metabolic disturbances are quite common in critically ill patients. Glycemic control appears to be an important adjuvant therapy in such patients. In addition, disorders of lipid metabolism are associated with worse prognoses. The purpose of this study was to investigate the effects that two different glycemic control protocols have on lipid profile and metabolism., Methods: We evaluated 63 patients hospitalized for severe sepsis or septic shock, over the first 72 h of intensive care. Patients were randomly allocated to receive conservative glycemic control (target range 140–180 mg/dl) or intensive glycemic control (target range 80–110 mg/dl). Serum levels of lowdensity lipoprotein, high-density lipoprotein, triglycerides, total cholesterol, free fatty acids, and oxidized low-density lipoprotein were determined., Results: In both groups, serum levels of low-density lipoprotein, high-density lipoprotein, and total cholesterol were below normal, whereas those of free fatty acids, triglycerides, and oxidized lowdensity lipoprotein were above normal. At 4 h after admission, free fatty acid levels were higher in the conservative group than in the intensive group, progressively decreasing in both groups until hour 48 and continuing to decrease until hour 72 only in the intensive group. Oxidized low-density lipoprotein levels were elevated in both groups throughout the study period., Conclusions: Free fatty acids respond to intensive glycemic control and, because of their high toxicity, can be a therapeutic target in patients with sepsis.

Sepsis and septic shock are associated with cardiac depression. Cardiovascular instability is a major cause of death in patients with sepsis. Focal adhesion kinase (FAK) is a potential mediator of cardiomyocyte responses to oxidative and mechanical stress. Myocardial collagen deposition can affect cardiac compliance and contractility. The aim of the present study was to determine whether the silencing of FAK is protective against endotoxemia-induced alterations of cardiac structure and function. In male Wistar rats, endotoxemia was induced by intraperitoneal injection of lipopolysaccharide (10 mg/kg). Cardiac morphometry and function were studied in vivo by left ventricular catheterization and histology. Intravenous injection of small interfering RNA targeting FAK was used to silence myocardial expression of the kinase. The hearts of lipopolysaccharide-injected rats showed collagen deposition, increased matrix metalloproteinase 2 activity, and myocyte hypertrophy, as well as reduced 24-h +dP/dt and -dP/dt, together with hypotension, increased left ventricular end-diastolic pressure, and elevated levels of FAK (phosphorylated and unphosphorylated). Focal adhesion kinase silencing reduced the expression and activation of the kinase in cardiac tissue, as well as protecting against the increased collagen deposition, greater matrix metalloproteinase 2 activity, and reduced cardiac contractility that occur during endotoxemia. In conclusion, FAK is activated in endotoxemia, playing a role in cardiac remodeling and in the impairment of cardiac function. This kinase represents a potential therapeutic target for the protection of cardiac function in patients with sepsis.

Lipid emulsion (LE) containing medium/ω-6 long chain triglyceride-based emulsion (MCT/ω-6 LCT LE) has been recommended in the place of ω-6 LCT-based emulsion to prevent impairment of immune function. The impact of MCT/ω-6 LCT LE on lymphocyte and neutrophil death and expression of genes related to inflammation was investigated. Seven volunteers were recruited and infusion of MCT/ω-6 LCT LE was performed for 6 h. Four volunteers received saline and no change was found. Blood samples were collected before, immediately afterwards and 18 h after LE infusion. Lymphocytes and neutrophils were studied immediately after isolation and after 24 and 48 h in culture. The following determinations were carried out: plasma-free fatty acids, triacylglycerol and cholesterol concentrations, plasma fatty acid composition, neutral lipid accumulation in lymphocytes and neutrophils, signs of lymphocyte and neutrophil death and lymphocyte expression of genes related to inflammation. MCT/ω-6 LCT LE induced lymphocyte and neutrophil death. The mechanism for MCT/ω-6 LCT LE-dependent induction of leucocyte death may involve changes in neutral lipid content and modulation of expression of genes related to cell death, proteolysis, cell signalling, inflammatory response, oxidative stress and transcription., (© 2011 The Authors. Clinical and Experimental Immunology © 2011 British Society for Immunology.)

Excessive free-radical production due to various bacterial components released during bacterial infection has been linked to cell death and tissue injury. Peroxynitrite is a highly reactive oxidant produced by the combination of nitric oxide (NO) and superoxide anion, which has been implicated in cell death and tissue injury in various forms of critical illness. Pharmacological decomposition of peroxynitrite may represent a potential therapeutic approach in diseases associated with the overproduction of NO and superoxide. In the present study, we tested the effect of a potent peroxynitrite decomposition catalyst in murine models of endotoxemia and sepsis. Mice were injected i.p. with LPS 40 mg/kg with or without FP15 [Fe(III) tetrakis-2-(N-triethylene glycol monomethyl ether)pyridyl porphyrin] (0.1, 0.3, 1, 3, or 10 mg/kg per hour). Mice were killed 12 h later, followed by the harvesting of samples from the lung, liver, and gut for malondialdehyde and myeloperoxidase measurements. In other subsets of animals, blood samples were obtained by cardiac puncture at 1.5, 4, and 8 h after LPS administration for cytokine (TNF-α, IL-1β, and IL-10), nitrite/nitrate, alanine aminotransferase, and blood urea nitrogen measurements. Endotoxemic animals showed an increase in survival from 25% to 80% at the FP15 doses of 0.3 and 1 mg/kg per hour. The same dose of FP15 had no effect on plasma levels of nitrite/nitrate. There was a reduction in liver and lung malondialdehyde in the endotoxemic animals pretreated with FP15, as well as in hepatic myeloperoxidase and biochemical markers of liver and kidney damage (alanine aminotransferase and blood urea nitrogen). In a bacterial model of sepsis induced by cecal ligation and puncture, FP15 treatment (0.3 mg/kg per day) significantly protected against mortality. The current data support the view that peroxynitrite is a critical factor mediating liver, gut, and lung injury in endotoxemia and septic shock: its pharmacological neutralization may be of therapeutic benefit.

Objective: To investigate the relationship between adequacy of energy intake and intensive care unit mortality in patients receiving exclusive enteral nutrition therapy., Methods: Observational and prospective study conducted during 2008 and 2009. Patients above 18 years with exclusive enteral nutrition therapy for at least 72 hours were included. The adequacy of energy intake was estimated by the administered/prescribed ratio. Non-conditional logistic regression was used to assess the relationship between predictive variables (adequacy of energy intake, APACHE II, gender, age, and intensive care unit length of stay) and intensive care unit mortality., Results: Sixty-three patients (mean 58 years, 27% mortality) were included, 47.6% of whom received more than 90% of the energy prescribed (mean adequacy 88.2%). Mean energy balance was -190 kcal/day. Significant associations between death in the intensive care unit and the variables age and intensive care unit length of stay were observed, after removing the variables adequacy of energy intake, APACHE II, gender and age during the modeling process., Conclusion: In our study, adequacy of energy intake did not affect intensive care unit mortality. Carefully followed enteral nutrition protocols, resulting in an administered/prescribed ratio above 70%, are apparently not sufficient to impact the mortality rates in the intensive care unit. Therefore, it may not be necessary to achieve 100% of the targeted energy, considering the high frequency of enteral feeding interruptions due to gastrointestinal intolerance and fasting for tests and procedures. Additional research is needed to identify the optimal energy intake for improved outcomes and reduced costs.

Sepsis syndrome is caused by inappropriate immune activation due to bacteria and bacterial components released during infection. This syndrome is the leading cause of death in intensive care units. Specialized B-lymphocytes located in the peritoneal and pleural cavities are known as B-1 cells. These cells produce IgM and IL-10, both of which are potent regulators of cell-mediated immunity. It has been suggested that B-1 cells modulate the systemic inflammatory response in sepsis. In this study, we conducted in vitro and in vivo experiments in order to investigate a putative role of B-1 cells in a murine model of LPS-induced sepsis. Macrophages and B-1 cells were studied in monocultures and in co-cultures. The B-1 cells produced the anti-inflammatory cytokine IL-10 in response to LPS. In the B-1 cell-macrophage co-cultures, production of proinflammatory mediators (TNF-α, IL-6 and nitrite) was lower than in the macrophage monocultures, whereas that of IL-10 was higher in the co-cultures. Co-culture of B-1 IL-10(-/-) cells and macrophages did not reduce the production of the proinflammatory mediators (TNF-α, IL-6 and nitrite). After LPS injection, the mortality rate was higher among Balb/Xid mice, which are B-1 cell deficient, than among wild-type mice (65.0% vs. 0.0%). The Balb/Xid mice also presented a proinflammatory profile of TNF-α, IL-6 and nitrite, as well as lower levels of IL-10. In the early phase of LPS stimulation, B-1 cells modulate the macrophage inflammatory response, and the main molecular pathway of that modulation is based on IL-10-mediated intracellular signaling., (Copyright © 2010 Elsevier GmbH. All rights reserved.)

Sepsis induces a systemic inflammatory response leading to tissue damage and cell death. LPS tolerance affects inflammatory response. To comprehend potential new mechanisms of immune regulation in endotoxemia, we examined macrophage mRNA expression by macroarray affected by LPS tolerance. LPS tolerance was induced with subcutaneous administration of 1 mg/kg/day of LPS over 5 days. Macrophages were isolated from the spleen and the expression of 1200 genes was quantitatively analyzed by the macroarray technique. The tolerant group displayed relevant changes in the expression of 84 mRNA when compared to naïve mice. A functional group of genes related to cell death regulation was identified. PARP-1, caspase 3, FASL and TRAIL genes were confirmed by RT-PCR to present lower expression in tolerant mice. In addition, reduced expression of the pro-inflammatory genes TNF-α and IFN-γ in the tolerant group was demonstrated. Following this, animals were challenged with polymicrobial sepsis. Flow cytometry analysis showed reduced necrosis and apoptosis in macrophages from the tolerant group compared to the naïve group. Finally, a survival study showed a significant reduction in mortality in the tolerant group. Thus, in the current study we provide evidence for the selective reprogramming of the gene expression of cell death pathways during LPS tolerance and link these changes to protection from cell death and enhanced survival rates., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Sepsis remains one of the leading causes of death in intensive care units. Progressive cardiovascular failure is an important cause of the mortality. Septic patients with myocardial dysfunction have significantly higher mortality compared with patients without cardiovascular impairment. Myocardial dysfunction in sepsis is characterized by decreased contractility and impaired myocardial compliance. Experimental studies of sepsis showed heterogeneity of microvascular perfusion, as well as impaired myocardial oxygen extraction. The underlying cellular mechanisms include increased neutrophil adhesion to the endothelium, production of reactive free radicals and oxidants, and endothelial dysfunction. Superoxide, nitric oxide and peroxynitrite cardiac formation has been demonstrated in septic hearts, which has been implicated in the pathogenesis of the myocardial depression and cell death in sepsis. Nitric oxide, carbon monoxide and hydrogen sulfide are gaseotransmitters that may exert protective effects in the septic heart.

Background: Fatty liver disease is a problem in both bariatric patients and in patients with moderate obesity. Tumor necrosis factor (TNF)-alpha has been frequently measured in nonalcoholic steatohepatitis (NASH) with or without diabetes, but less is known about interleukin (IL)-6 and IL-10., Methods: Moderately obese patients (n = 80) with histologically proven steatosis (n = 29) and NASH (n = 51) were recruited. Serum levels of cytokines were documented along with clinical information. The aim was to identify the correlates of such biomolecules in a stable population., Results: Diabetes tended to be more associated with NASH (52.5% instead of 41.4%, P = 0.015), with no difference of age, gender, or body mass index regarding steatosis. For the entire population, cytokine changes were not significant, including TNF-alpha and IL-6. In diabetics only, all markers tended to diminish with NASH, especially IL-10 (P = 0.000). IL-10 correlated with homeostatic model assessment index (P = 0.000) and other variables of glucose homeostasis in diabetes, thus representing a major marker of the disease., Conclusions: (1) Generally inconsistent changes in pro- and anti-inflammatory cytokines occurred when NASH was globally compared to steatosis. (2) In contrast, downregulation of IL-6 and IL-10 was perceived in diabetics with NASH. (3) Arterial hypertension did not play a role in these circumstances. (4) IL-10 maintained strong correlations with glucose metabolism indices. (5) TNF-alpha could not be incriminated for progressive liver damage, as values failed to increase in NASH. (6) Investigations of IL-10 and other counterregulatory cytokines are lacking in this context and deserve further studies.

Extensive lymphocyte apoptosis may be an important cause of immune suppression in sepsis. Here we investigated the effect of LPS tolerance on lymphocyte apoptosis in an experimental model of polymicrobial infection. Tolerance was induced by the injection of lipopolysaccharide (1.0mg/kg/subcutaneously) once a day for 5 days. Macroarray analysis of mRNA isolated from T-(CD4) lymphocytes was used to identify genes that are differentially expressed during LPS tolerance. In addition, assessment of the expression of apoptosis-associated lymphocyte gene products and apoptotic events was performed on the 8th day; 6h after the terminal challenge with polymicrobial infection or high-dose LPS administration. Survival studies with polymicrobial infection were also conducted. LPS tolerance induced a broad reprogramming of cell death pathways, including a suppression of receptor-mediated and mitochondrial apoptotic pathways, inflammatory caspases, alternate apoptotic pathways, as well as reduced expression of genes involved in necrosis. These alterations led to a marked resistance of lymphocytes against cell death during the subsequent period of sepsis. In addition, LPS tolerance produced an increased differentiation of T-lymphocytes to T(H)1 and T(H)2, with a T(H)1 differentiation predominance. Thus, in the current study we provide an evidence for a marked reprogramming of gene expression of multiple cell death pathways during LPS tolerance. These alterations may play a significant role in the observed protection of the animals from a subsequent lethal polymicrobial sepsis challenge., (Copyright 2009 Elsevier GmbH. All rights reserved.)

Sepsis is a systemic inflammatory response that can lead to tissue damage and death. In order to increase our understanding of sepsis, experimental models are needed that produce relevant immune and inflammatory responses during a septic event. We describe a lipopolysaccharide tolerance mouse model to characterize the cellular and molecular alterations of immune cells during sepsis. The model presents a typical lipopolysaccharide tolerance pattern in which tolerance is related to decreased production and secretion of cytokines after a subsequent exposure to a lethal dose of lipopolysaccharide. The initial lipopolysaccharide exposure also altered the expression patterns of cytokines and was followed by an 8- and a 1.5-fold increase in the T helper 1 and 2 cell subpopulations. Behavioral data indicate a decrease in spontaneous activity and an increase in body temperature following exposure to lipopolysaccharide. In contrast, tolerant animals maintained production of reactive oxygen species and nitric oxide when terminally challenged by cecal ligation and puncture (CLP). Survival study after CLP showed protection in tolerant compared to naive animals. Spleen mass increased in tolerant animals followed by increases of B lymphocytes and subpopulation Th1 cells. An increase in the number of stem cells was found in spleen and bone marrow. We also showed that administration of spleen or bone marrow cells from tolerant to naive animals transfers the acquired resistance status. In conclusion, lipopolysaccharide tolerance is a natural reprogramming of the immune system that increases the number of immune cells, particularly T helper 1 cells, and does not reduce oxidative stress.

1. We recently demonstrated that hypertonic saline reduces inflammation and mortality in acute pancreatitis. The present study investigated the effects of hypertonic saline in metalloproteinase (MMP) regulation and pancreatitis-associated hepatic injury. 2. Wistar rats were divided into four groups: (i) control, not subjected to insult or treatment; (ii) no treatment (NT), induction of pancreatitis (retrograde infusion of 2.5% sodium taurocholate (1.0 mL/kg)), but no further treatment; (iii) normal saline (NS), induction of pancreatitis and treatment with normal saline (0.9% NaCl, 34 mL/kg, i.v. bolus, 1 h after the induction of pancreatitis); and (iv) hypertonic saline (HS), induction of pancreatitis and treatment with hypertonic saline (7.5% NaCl, 4 mL/kg administered over a period of 5 min, 1 h after the induction of pancreatitis). In all four groups, 4, 12 and 24 h after the induction of pancreatitis, liver tissue samples were assayed to determine levels of MMP-2, MMP-9, 47 kDa heat shock protein (HSP47) and collagen (Type I and III). 3. Compared with the control group, MMP-9 expression and activity was increased twofold in the NS and NT groups 4 and 12 h after the induction of pancreatitis, but remained at basal levels in the HS group. In contrast, MMP-2 expression was increased twofold 12 h after the induction of pancreatitis only in the NS group, whereas the expression of HSP47 was increased 4 h after the induction of pancreatitis in the NS and NT groups. Greater extracellular matrix remodelling occurred in the NS and NT groups compared with the HS group, probably as a result of the hepatic wound-healing response to repeated injury. However, the collagen content in hepatic tissue remained at basal levels in the HS group. 4. In conclusion, the results of the present study indicate that hypertonic saline is hepatoprotective and reduces hepatic remodelling, maintaining the integrity of the hepatic extracellular matrix during pancreatitis. Hypertonic saline-mediated regulation of MMP expression may have clinical relevance in pancreatitis-associated liver injury.

Objective: Monitor the adequacy of enteral nutritional therapy at the intensive care unit aiming to improve the quality of nutritional assistance., Methods: Prospective and observational study developed at the adult intensive care unit from 2005 to 2008. Patients over 18 years of age with exclusive enteral nutritional therapy for over 72h participated in the sample. The average values and the percentile adequacy of energy and proteins calculated, prescribed and administered in each year were analyzed. The factors responsible for the non-conformity of the administration planned were classified into intensive care unit extrinsic or intrinsic causes. The quality indicators proposed by the ILSI Brazil were applied, and expressed into percentile goals. In the statistic analyses, confidence interval and the t Student e Mann-Whitney (p<0.05) tests were used, according to the Epi Info program., Results: One hundred and sixteen patients were followed up. There were statically difference in values of energy and protein administered in 2005 and in 2006, when compared to those in 2008. The adequacy calculated/prescribed remained close to 100% in all the surveys and the adequacy administered/prescribed increased from 74% in 2005, to 89% in 2008. An increase in interruptions of enteral nutritional therapy for external factors and the decrease in interruptions for intensive care unit internal factors were verified. The quality indicators equally reflect the evolution of the patient care., Conclusion: In the four yearly surveys, a progressive enhancement of nutritional support was verified. Quality indicators allow nutritional care evolution monitoring, the comparison to other services data, and are a new perspective for enteral nutritional therapy assessment.

Sepsis involves a systemic inflammatory response of multiple endogenous mediators, resulting in many of the injurious and sometimes fatal physiological symptoms of the disease. This systemic activation leads to a compromised vascular response and endothelial dysfunction. Purine nucleotides interact with purinoceptors and initiate a variety of physiological processes that play an important role in maintaining cardiovascular function. The purpose of the present study was to investigate the effects of ATP on vascular function in a lipopolysaccharide (LPS) model of sepsis. LPS induced a significant increase in aortic superoxide production 16 h after injection. Addition of ATP to the organ bath incubation solution reduced superoxide production by the aortas of endotoxemic animals. Reactive Blue, an antagonist of the P2Y receptor, blocked the effect of ATP on superoxide production, and the nonselective P2Y agonist MeSATP inhibited superoxide production. Nitric oxide synthase (NOS) inhibition by L-NAME blocked vascular relaxation and reduced superoxide production in LPS-treated animals. In the presence of L-NAME there was no ATP effect on superoxide production. A vascular reactivity study showed that ATP increased maximal relaxation in LPS-treated animals compared to controls. The presence of ATP induced increases in Akt and endothelial NOS phosphorylated proteins in the aorta of septic animals. ATP reduces superoxide release resulting in an improved vasorelaxant response. Sepsis may uncouple NOS to produce superoxide. We showed that ATP through Akt pathway phosphorylated endothelial NOS and "re-couples" NOS function.

Objective: To describe the composition of metabolic acidosis in patients with severe sepsis and septic shock at intensive care unit admission and throughout the first 5 days of intensive care unit stay., Design: Prospective, observational study., Setting: Twelve-bed intensive care unit., Patients: Sixty patients with either severe sepsis or septic shock., Interventions: None., Measurements and Main Results: Data were collected until 5 days after intensive care unit admission. We studied the contribution of inorganic ion difference, lactate, albumin, phosphate, and strong ion gap to metabolic acidosis. At admission, standard base excess was -6.69 +/- 4.19 mEq/L in survivors vs. -11.63 +/- 4.87 mEq/L in nonsurvivors (p < .05); inorganic ion difference (mainly resulting from hyperchloremia) was responsible for a decrease in standard base excess by 5.64 +/- 4.96 mEq/L in survivors vs. 8.94 +/- 7.06 mEq/L in nonsurvivors (p < .05); strong ion gap was responsible for a decrease in standard base excess by 4.07 +/- 3.57 mEq/L in survivors vs. 4.92 +/- 5.55 mEq/L in nonsurvivors with a nonsignificant probability value; and lactate was responsible for a decrease in standard base excess to 1.34 +/- 2.07 mEq/L in survivors vs. 1.61 +/- 2.25 mEq/L in nonsurvivors with a nonsignificant probability value. Albumin had an important alkalinizing effect in both groups; phosphate had a minimal acid-base effect. Acidosis in survivors was corrected during the study period as a result of a decrease in lactate and strong ion gap levels, whereas nonsurvivors did not correct their metabolic acidosis. In addition to Acute Physiology and Chronic Health Evaluation II score and serum creatinine level,inorganic ion difference acidosis magnitude at intensive care unit admission was independently associated with a worse outcome., Conclusions: Patients with severe sepsis and septic shock exhibit a complex metabolic acidosis at intensive care unit admission, caused predominantly by hyperchloremic acidosis,which was more pronounced in nonsurvivors. Acidosis resolution in survivors was attributable to a decrease in strong ion gap and lactate levels.

Shock and resuscitation render patients more susceptible to acute lung injury due to an exacerbated immune response to subsequent inflammatory stimuli. To study the role of innate immunity in this situation, we investigated acute lung injury in an experimental model of ischemia-reperfusion (I-R) followed by an early challenge with live bacteria. Conscious rats (N = 8 in each group) were submitted to controlled hemorrhage and resuscitated with isotonic saline (SS, 0.9% NaCl) or hypertonic saline (HS, 7.5% NaCl) solution, followed by intratracheal or intraperitoneal inoculation of Escherichia coli. After infection, toll-like receptor (TLR) 2 and 4 mRNA expression was monitored by RT-PCR in infected tissues. Plasma levels of tumor necrosis factor alpha and interleukins 6 and 10 were determined by ELISA. All animals showed similar hemodynamic variables, with mean arterial pressure decreasing to nearly 40 mmHg after bleeding. HS or SS used as resuscitation fluid yielded equal hemodynamic results. Intratracheal E. coli inoculation per se induced a marked neutrophil infiltration in septa and inside the alveoli, while intraperitoneal inoculation-associated neutrophils and edema were restricted to the interseptal space. Previous I-R enhanced lung neutrophil infiltration upon bacterial challenge when SS was used as reperfusion fluid, whereas neutrophil influx was unchanged in HS-treated animals. No difference in TLR expression or cytokine secretion was detected between groups receiving HS or SS. We conclude that HS is effective in reducing the early inflammatory response to infection after I-R, and that this phenomenon is achieved by modulation of factors other than expression of innate immunity components.

Objectives: Acute pancreatitis (AP) protease release induces lung parenchymal destruction via matrix metalloproteinases (MMPs), a neutrophil (polymorphonuclear leukocyte)-dependent process. Recent studies in hemorrhagic shock revealed that hypertonic saline (HTS) has an anti-inflammatory effect and can inhibit a variety of neutrophil functions. The aim of this study was to determine whether HTS and its actions in the pathway of neutrophil migration, MMPs, and heat shock proteins (HSPs) are effective in protecting the lung from injury associated with AP., Methods: We determined neutrophil infiltration and expressions of MMPs and HSPs in the lung tissue after AP induced by retrograde infusion of 2.5% of sodium taurocholate., Results: Animals submitted to AP that received HTS compared with those who received normal saline presented with increased HSP70 and HSP90 expressions and reduced myeloperoxidase levels and MMP-9 expression and activity., Conclusions: Our data raised the hypothesis that a sequence of HTS lung protection events increases HSP70 and HSP90, inhibiting infiltration of neutrophils and their protease actions in the lung.

Background: Dobutamine is the agent of choice for increasing cardiac output during myocardial depression in humans with septic shock. Studies have shown that beta-adrenoceptor agonists influence nitric oxide generation, probably by modulating cyclic adenosine monophosphate. We investigated the effects of dobutamine on the systemic and luminal gut release of nitric oxide during endotoxic shock in rabbits., Material/methods: Twenty anesthetized and ventilated New Zealand rabbits received placebo or intravenous lipopolysaccharide with or without dobutamine (5 micro g/kg/min). Ultrasonic flow probes placed around the superior mesenteric artery and the abdominal aorta continuously estimated the flow. A segment from the ileum was isolated and perfused, and serum nitrate/nitrite levels were measured in the perfusate solution and the serum every hour., Results: The mean arterial pressure decreased with statistical significance in the lipopolysaccharide group but not in the lipopolysaccharide/dobutamine group. The abdominal aortic flow decreased statistically significantly after lipopolysaccharide administration in both groups but recovered to baseline in the lipopolysaccharide/dobutamine group. The flow in the superior mesenteric artery was statistically significantly higher in the lipopolysaccharide/dobutamine group than in the lipopolysaccharide group at 2 hours. The serum nitrate/nitrite levels were higher in the lipopolysaccharide group and lower in the lipopolysaccharide/dobutamine group than those in the control group. The gut luminal perfusate serum nitrate/nitrite level was higher in the lipopolysaccharide group than in the lipopolysaccharide/dobutamine group., Conclusions: Dobutamine can decrease total and intestinal nitric oxide production in vivo. Those effects seem to be inversely proportional to the changes in blood flow.

This paper analyzes the discussions in the field of legal medicine in Mexico about the prudence of regulations concerning intermarriage that were decreed in the civil codes for the Federal District of 1871 and 1884. It shows that the heated debate forced the authors of the regulations to struggle for the need for medical vigilance of marriages between relatives, as a preventive measure sustained in a nihilistic vision of the pathological inheritance. The paper concludes by proposing a philosophical analysis that abandons the old fields of the "external" and the "internal".

The mechanisms that initiate an inflammatory systemic response to a bacterial infection lead to a high mortality and constitute the first cause of death in Critical Care Units (ICU's). Sepsis is a poorly understood disease and despite life support techniques and the administration of antibiotics, not much more can be done to improve its diagnosis and treatment. The present article has as main objective to discuss the role of neutrophils recruitment in sepsis, dissecting the molecular mechanisms implicated in this complex process and its importance to the pathogenesis of this outstanding cause of death.

Mice expressing human cholesteryl ester transfer protein (huCETP) are more resistant to Escherichia coli bacterial wall LPS because death rates 5 days after intraperitoneal inoculation of LPS were higher in wild-type than in huCETP+/+ mice, whereas all huCETP+/+ mice remained alive. After LPS inoculation, plasma concentrations of TNF-alpha and IL-6 increased less in huCETP+/+ than in wild-type mice. LPS in vitro elicited lower TNF-alpha production by CETP expressing than by wild-type macrophages. In addition, TNF-alpha production by RAW 264.7 murine macrophages increased on incubation with LPS but decreased in a dose-dependent manner when human CETP was added to the medium. Human CETP in vitro enhanced the LPS binding to plasma high-density lipoprotein/low-density lipoprotein. The liver uptake of intravenous infused 14C-LPS from Salmonella typhimurium was greater in huCETP+/+ than in wild-type mice. Present data indicate for the first time that CETP is an endogenous component involved in the first line of defense against an exacerbated production of proinflammatory mediators.