Sepsis, the body's response to infection, is associated with extremely high mortality rates. Why a protective mechanism turns into a deadly clinical picture is a matter of debate, and goes largely unexplained. In previous work we demonstrated that platelet-derived microparticles (MP) can induce endothelial and vascular smooth muscle cell apoptosis in septic patients through NADPH oxidasedependent superoxide release [1]. In this work we sought to create a model for ex vivo generation of septic-like MP and to identify the pathways responsible for MP free radical release and effects. Septic shock is a condition related to the generation of high amounts of thrombin, TNFα and nitrogen reactive species. Human platelets exposed to the NO donors diethylamine-NONOate (0.5 mM) and nitroprusside (2 mM) for 20 minutes generated MP similar to those found in the blood of septic shock patients. Flow cytometry and western blot analysis of those MP, like their septic counterparts, revealed exposure of the tetraspanin markers CD9, CD63, and CD81, but little phosphatidylserine. Such a membrane exposure, associated with their size, characterizes them as exosomes. Furthermore, we identified the Nox2 and p22phox NADPH oxidase subunits and the inducible isoform of NO synthase (NOS), but not the NOS I and III isoforms. On the other hand, platelets exposed to thrombin or TNFα released particles with clearly distinct characteristics, such as high phosphatidylserine and low tetraspanin. Like the septic MP, the MP obtained by NO exposure generated the superoxide radical and NO, as disclosed by lucigenin (5 µM) and coelenterazine (5 µM) chemiluminescence and by 4,5-diaminofluorescein (10 mM) and 2′,7′-dichlorofluorescein (10 mM) fluorescence. As expected, NOS inhibitors or NADPH oxidase inhibitors significantly reduced signals. In addition, endothelial cells exposed to this type of MP underwent apoptotic death, while control MP had negligible effects. NADPH oxidase as well as NOS inhibition significantly reduced apoptosis rates. Concomitant generation of NO and superoxide suggests biological effects of the highly reactive radical peroxynitrite. In fact, the peroxynitrite scavenger urate (1 mM) showed an additive effect on fluorescent signal inhibition, as well as on endothelial apoptosis rate reduction. We thus propose that platelet-derived exosomes may be another class of actors in the complex play known as 'vascular redox signaling'. In this sense, an exosome-based approach can provide novel tools for further understanding and even treating vascular dysfunction related to sepsis. Introduction The intestinal hypothesis of sepsis has been attributed to bacterial translocation (BT), and the aggravation of sepsis is related to the increased vascular permeability state that potentates the BT index. In this study we examined the BT index during sepsis with or without mesenteric lymph exclusion. Materials and methods Wistar rats (±200 g) were submitted to the BT process (E. coli R6 10 ml of 10 10 CFU/ml) and nonlethal sepsis (E. cloacae 89 2 ml ...
Introduction Cardiac surgery with cardiopulmonary bypass (CPB) is a recognized trigger of systemic inflammatory response, usually related to postoperative acute lung injury (ALI). As an attempt to dampen inflammatory response, steroids have been perioperatively administered to patients. Macrophage migration inhibitory factor (MIF), a regulator of the endotoxin receptor, is implicated in the pathogenesis of ALI. We have previously detected peak circulating levels of MIF, 6 hours post CPB. Experimental data have shown that steroids may induce MIF secretion by mononuclear cells. This study aims to correlate levels of MIF assayed 6 hours post CPB to the intensity of postoperative pulmonary dysfunction, analysing the impact of perioperative steroid administration. MethodsWe included patients submitted to cardiac surgery with CPB, electively started in the morning, performed by the same team under a standard technique except for the addition of methylprednisolone (15 mg/kg) to the CPB priming solution for patients from group MP (n = 37), but not for the remaining patients -group NS (n = 37). MIF circulating levels were assayed at the anesthesia induction, 3, 6, and 24 hours after CPB. A standard weaning protocol with fast track strategy was adopted, and indicators of organ dysfunction and therapeutic intervention were registered during the first 72 hours postoperative.Results Levels of MIF assayed 6 hours post CPB correlated directly to the postoperative duration of mechanical ventilation (P = 0.014, rho = 0.282) and inversely to PaO 2 /FiO 2 ratio (P = 0.0021, rho = -0.265). No difference in MIF levels was noted between the groups. The duration of mechanical ventilation was higher (P = 0.005) in the group MP (7.92 ± 6.0 hours), compared with the group NS (4.92 ± 3.6 hours). ConclusionCirculating levels of MIF assayed 6 hours post CPB are correlated to postoperative pulmonary performance. Immunosuppressive doses of methylprednisolone did not affect circulating levels of MIF and may be related to prolonged mechanical ventilation. P2Immediate and short-term safety of catheter-based autologous bone marrow-derived mononuclear cell transplantation into myocardium of patients with severe ischemic heart failure
CARDIOLOGY P1 Impact of peroperative administration of steroid over inflammatory response and pulmonary dysfunction following cardiac surgery
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