SvO(2)-guided resuscitation for experimental septic shock: effects of fluid infusion and dobutamine on hemodynamics, inflammatory response, and cardiovascular oxidative stress.

The pathogenetic mechanisms associated to the beneficial effects of mixed venous oxygen saturation (SvO(2))-guided resuscitation during sepsis are unclear. Our purpose was to evaluate the effects of an algorithm of SvO(2)-driven resuscitation including fluids, norepinephrine and dobutamine on hemodynamics, inflammatory response, and cardiovascular oxidative stress during a clinically resembling experimental model of septic shock. Eighteen anesthetized and catheterized pigs (35-45 kg) were submitted to peritonitis by fecal inoculation (0.75 g/kg). After hypotension, antibiotics were administered, and the animals were randomized to two groups: control (n = 9), with hemodynamic support aiming central venous pressure 8 to 12 mmHg, urinary output 0.5 mL/kg per hour, and mean arterial pressure greater than 65 mmHg; and SvO(2) (n = 9), with the goals above, plus SvO(2) greater than 65%. The interventions lasted 12 h, and lactated Ringer's and norepinephrine (both groups) and dobutamine (SvO(2) group) were administered. Inflammatory response was evaluated by plasma concentration of cytokines, neutrophil CD14 expression, oxidant generation, and apoptosis. Oxidative stress was evaluated by plasma and myocardial nitrate concentrations, myocardial and vascular NADP(H) oxidase activity, myocardial glutathione content, and nitrotyrosine expression. Mixed venous oxygen saturation-driven resuscitation was associated with improved systolic index, oxygen delivery, and diuresis. Sepsis induced in both groups a significant increase on IL-6 concentrations and plasma nitrate concentrations and a persistent decrease in neutrophil CD14 expression. Apoptosis rate and neutrophil oxidant generation were not different between groups. Treatment strategies did not significantly modify oxidative stress parameters. Thus, an approach aiming SvO(2) during sepsis improves hemodynamics, without any significant effect on inflammatory response and oxidative stress. The beneficial effects associated with this strategy may be related to other mechanisms.

Differential expression of toll-like receptor signaling cascades in LPS-tolerant human peripheral blood mononuclear cells.

Pre-exposure to low doses of LPS induces resistance to a lethal challenge, a phenomenon known as endotoxin tolerance. In this study, tolerance was induced in human PBMC by culturing cells with 1 ng/mL LPS for 48 h. Cells were subsequently challenged with 100 ng/mL LPS for 2, 6 and 24 h, and the expression of 84 genes encoding proteins involved in the TLR signaling pathway was evaluated at each time point by PCR array. LPS pretreatment did not modulate the expression of TLR4 and CD14 on the surface of monocytes. A gene was defined as tolerized when LPS pretreatment reversed the effect of LPS challenge on the expression of the gene or as non-tolerized when LPS pretreatment did not reverse the effects of LPS challenge. We observed impaired signal transduction through the NF-κB, JNK, ERK and TRIF pathways, whereas expression of p38 pathway-related genes was preserved in LPS-tolerant cells. These results show a distinct regulation of the TLR pathway cascades during tolerance; this may account for the differential gene expression of some inflammatory mediators, such as up-regulation of IL-10 and COX2 as well as down-regulation of TNF-α and IL-12. Depending on the effect of LPS-induced gene up-regulation or down-regulation, tolerance, as a reversion of such LPS effects, may result in repression or induction of gene expression.

Toll-like receptor pathway signaling is differently regulated in neutrophils and peripheral mononuclear cells of patients with sepsis, severe sepsis, and septic shock.

OBJECTIVES: Up- and down-regulation of inflammatory response was described in blood cells from septic patients, according to the stage of sepsis and the cells evaluated. This study aimed to evaluate the Toll-like receptor (TLR) signaling pathway gene expression in peripheral blood mononuclear cells (PBMC) and neutrophils in patients throughout the different stages of sepsis. DESIGN: Prospective, observational study. SETTINGS: Two emergency rooms and two intensive care units in one university and one teaching hospital. PATIENTS AND CONTROLS: A total of 15 septic patients, five with sepsis, five with severe sepsis, and five with septic shock, in addition to five healthy volunteers were enrolled. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The Human-TLR Signaling Pathway, which comprises 84 genes related to TLR-mediated signal transduction, was evaluated by real time polymerase chain reaction in PBMC and neutrophils obtained from patients and controls. The fold change for each gene (2(-Delta DeltaCt)) was compared between the groups. Genes with fold changes greater than 2 and significant changes in DeltaCT are reported as differently expressed. The fold change ratios in PBMC gene expression between septic patients and healthy controls revealed a dynamic process according to the stage of sepsis, tending toward down-regulation of the TLR signaling pathway in PBMC in the more severe forms of the disease. However, the differential gene expression was restricted to five down-regulated genes in septic shock patients, which are found in the effector and downstream pathways. Neutrophils showed a different pattern of adaptation. Patients with sepsis, severe sepsis, and septic shock presented a broad gene up-regulation, which included all functional groups evaluated and persisted throughout the stages of the disease. CONCLUSIONS: TLR-signaling pathway genes are differently regulated in PBMC and neutrophils of septic patients, and are dynamically modulated throughout the different stages of sepsis.

TLR signaling pathway in patients with sepsis.

The pathogenesis of sepsis involves complex interaction between the host and the infecting microorganism. Bacterial recognition and signaling are essential functions of the cells of innate immune systems and drive a coordinated immune response. One of the more intriguing aspects of sepsis is the fact that the protective and damaging host response are part of the same process, that is, the inflammatory response that is aimed to control the infectious process also underscores many of the pathophysiological events of sepsis. The discovery of Toll-like receptors (TLRs) in humans, and the early recognition of TLR-4 as the receptor that signals LPS bioactivity were major breakthroughs not only in the field of sepsis but also in immunology as a whole. In this article, we aimed to review TLR expression and signaling in the context of sepsis. The results obtained by our group show that TLR and other cellular surface receptors may be differently regulated on mononuclear cells and neutrophils, and that they are dynamically modulated across the stages of sepsis. Toll-like receptor signaling gene expression in mononuclear cells is decreased in more severe forms of the disease. In contrast, up-regulated genes are seen along the clinical spectrum of sepsis in neutrophils.