Postshock mesenteric lymph induces endothelial NF-KB activation

Background. Posthemorrhagic shock mesenteric lymph (PSML) has been shown to activate pulmonary endothelial cells and cause lung injury. Although multiple mediators may be involved, most of these effects are mediated by nuclear factor-kappa B (NF-kappa B) activation. Degradation of the inhibitor of kappa B (I kappa B) is a key regulatory step in the activation of NF-kappa B. We therefore hypothesized that PSML would cause I kappa B degradation with subsequent NF-kappa B phosphorylation and nuclear translocation. Methods. Mesenteric lymph was collected from male rats before shock and each hour after shock for up to 3 h (n = 5). Buffer (control), buffer + 10% (v/v) lymph, or buffer + tumor necrosis factor (10 ng/mL) were incubated with human pulmonary endothelial cells for 30 min and then lysed. Immunoblots of lysates were probed for licB and phospho-p65. Immunohistochemistry was performed on cells grown on glass slides and then treated as above with the third PSML sample. Cells were fixed and then probed for p65. Statistical analysis was performed with Student's t-test and analysis of variance with significance was set at P < 0.05. Results. Western blots of cell lysates for I kappa B demonstrated a steady decrease in total I kappa B with each lymph sample. Phosphorylation of NF-kappa B, p65 component, steadily increased with each PSML sample, with a maximum reached during the third PSML sample, which also significantly increased translocation of NF kappa B to the nucleus. Conclusion. Postshock mesenteric lymph bioactivity is mediated by pathways which involved licB degradation. These pathways offer novel off targets for clinical intervention to prevent the distal organ injury caused by post-injury hemorrhagic shock. (c) 2007 Elsevier Inc. All rights reserved.