Eed to be stimulated.P104 p22phox-containing microparticles from plasma of septic patients are related to reactive

Eed to be stimulated.P104 p22phox-containing microparticles from plasma of septic patients are related to reactive Betulin oxygen species production and apoptosis of vascular cells PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/20725854 in cultureMA Pedro*, AO Carmo*, M Janiszewski*, E Silva*, E Knobel*, FRM Laurindo *Intensive Care Unit — Basic Research Laboratory, Hospital Israelita Albert Einstein, Av Albert Einstein, 627 CTI A – Sao Paulo SP 05651 901, Brazil; University of S Paulo, Medical School, Av Eneas C Aguiar, 44 S/S – S Paulo SP 05403 900, Brazil Reactive oxygen species (ROS) are related to vascular dysfunction in several pathologies and may be related to the pathophysiology of sepsis and multiple organ dysfunction. To investigate their role and sources we evaluated the effects of plasma of septic patients (SPP) on reactive ROS production and apoptosis of rabbit endothelial (REC) and aortic smooth muscle (RASM) cells in culture through NAD(P)H (0.3 mM)-driven lucigenin (5 ) luminescence and annexin V assay respectively. SPP alone showed intrinsic luminescence when compared to plasma from healthy controls (HCP) (51.4 ?9.5 cpm/min/mg protein vs 7.0 ?1.2, n = 5, P < 0.05). After incubation with REC and RASM cells SPP induced a two-fold increase in ROS production (n = 5, P < 0.05) when compared to incubation with HCP. This enhancement of ROS production by REC and RASM cells were paralleled by increase in apoptosis rates induced by SPP. The fraction responsible for ROS production and apoptosis induction was determined through filtration and ultracentrifugation to be over 50 kD and to contain microparticles (MP), similar to those from activated platelets. MP showed similar luminescence (60.6 ?4.4 vs 32.9 ?3.1, n = 11), blocked by DPI (60 ), a flavoenzyme inhibitor, and by SOD (80 ). Addition of MP from SPP to REC and RASM also induced increased luminescence when compared to the effect of MP from HCP; These signals were also inhibitable by SOD and DPI. Incubation of REC and RASM with MP from SPP doubled apoptosis rates when compared to incubation with MP from HCP. Apoptosis was also inhibited by addition of SOD or DPI. After lipid extraction, western-blot analysis of the MP was positive for the p22phox and gp91 subunits of the NAD(P)H oxidase. Thus, SPP has intrinsic and enhances REC and RASM ROS production, due to NAD(P)H oxidase activity. It causes apoptosis of REC and RASM, also inhibited by SOD and DPI. These effects may be attributable to MP containing the p22phox and gp91 subunits of NAD(P)H oxidase. ROS derived from microparticles may represent a new signaling pathway involved in the pathophysiology of severe sepsis.P105 LPS activation of leukocytes attenuates adhesion to endothelial cells under shear stressB Noh? V Schmidt, A Hientz, C Zanke, HJ Dieterich, K Unertl Department of Anaesthesiology and Critical Care, University Hospital, Hoppe-Seyler-Str 3, 72076 T ingen, Germany Leukocyte adhesion is triggered by upregulation of cell surface adhesion molecules and counteracted by the shear forces of the flowing blood. Since both factors, inflammatory response and flow dynamics are severely altered during endotoxemia, we studied the effects of endotoxin on leukocyte ndothelial interactions under different levels of shear stress in vitro. In order to perform the experiments at precisely adjustable levels of shear stress, we used a parallel plate flow chamber as has been employed in a number of adhesion studies previously. Within this chamber, endothelial cells (HUVEC) could be perfused w.