Early fluid replacement with hypertonic isoncotic solution guided by mixed venous oxygen saturation in experimental hypodynamic sepsis
Critical Care 2007, 11(Suppl 3):P1 (doi: 10.1186/cc5788) 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 . In this work we
... ght 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. Reference 1. Janiszewski M, Do Carmo AO, Pedro MA, et al.: Crit Care Med 2004, 32:818-825. P2 Which is the worst factor in sepsis aggravation: translocated bacterial amount or gut-associated lymphoid tissue activation?