Induction of Tolerance to Hemorrhagic or Endotoxic Shock Involves Activation of NF-κB

Abstract

Background.Tolerance to hemorrhagic or endotoxic shock can be induced by prior sublethal hemorrhage (SLH). The purpose of this study was to explore whether alterations in signal transduction pathways involving NF-κB occur in macrophages (Mφ) following induction of tolerance by SLH.Methods.Using a model of SLH previously shown in our lab to impart a survival benefit to subsequent hemorrhagic or endotoxic shock, rats (n= 30) were conditioned by SLH. Peritoneal Mφ were harvested 24 h after conditioning and stimulated with lipopolysaccharide (LPS) (10 μg/mL). Nuclear and cytosolic proteins were isolated 1 h later for determination of NF-κB activation by gel-shift assay and IκB-α by Western blot. TNF mRNA gene expression was measured 4 h after LPS stimulation by reverse transcription/polymerase chain reaction (RT/PCR). TNF protein levels were measured in cellular supernatants by enzyme-linked immunosorbent assay (ELISA) 18 h after LPS.Results.LPS stimulation of sham Mφ increased NF-κB activation with corresponding loss of its inhibitor IκB-α. In contrast, IκB-α was not detectable following conditioning, and conditioned Mφ had NF-κB activation at baseline which increased minimally with LPS stimulation. LPS increased TNF gene expression and significantly increased protein production by both sham and conditioned Mφ, but this increase was greater in the sham-conditioned group.Conclusions.The ability of Mφ from animals made tolerant by SLH to produce TNFin vitrois conserved. Nevertheless, these same Mφ exhibit alterations in TNF gene induction and expression as well as signal transduction, specifically, changes in IκB-α and NF-κB activation. This suggests a role for activation of NF-κB in the induction of tolerance.