Post-hemorrhagic shock mesenteric lymph lipids prime neutrophils for enhanced cytotoxicity via phospholipase A2

Hemorrhagic shock induced mesenteric hypoperfusion has long been implicated as a key event in the pathogenesis of the adult respiratory distress syndrome (ARDS) and multiple organ failure (MOF). Previous work links post-hemorrhagic shock mesenteric lymph (PHSML) lipids and neutrophil (PMN) priming in the pathogenesis of ARDS. We hypothesize that gut phospholipase A(2)(PLA(2)) liberates proinflammatory lipids following hemorrhagic shock, which are responsible for enhanced PMN cytotoxicity. Mesenteric lymph was collected from rats (n greater than or equal to 5) before hemorrhagic shock, during hemorrhagic shock (MAP 40 mm Hg x 30 min), and after resuscitation (shed blood + 2x lactated Ringers). PMNs were incubated with physiologic concentrations (1-5%, v:v) of (a) buffer control, (b) sham (c) pre-shock lymph, (c) PHSML, (d) PHSML lipid extracts, (e) heat-denatured PSHML, and (f)PHSML harvested after IV pretreatment with a known PLA(2) inhibitor (quinacrine,10 mg/kg). PMNs were activated with fMLP (1 pmol), and the maximal rate of superoxide production measured by reduction of cytochrome c. Gut morphology was assessed histologically using hematoxalin and eosin (HE) staining. PHSML and PHSML lipid extracts (5%, v:v) primed for enhanced superoxide production compared to buffer controls (2.5-fold and 3.6-fold), sham (2.5-fold) and pre-shock lymph (2.0-fold). Lymph collected after systemic PLA(2) inhibition, in contrast, abrogated the PMN priming response. Gut mucosal morphology, at end-resuscitation, was intact on HE staining both with and without PLA(2) inhibition. Heat denaturing the PHSML (eliminating cytokines and complement), on the other hand, did not reduce PMN priming. Physiologic concentrations of PHSML lipids prime the PMN respiratory burst. Lymph priming is diminished with systemic PLA(2) inhibition, implicating gut PLA(2) as a source of proinflammatory lipids that may be central in the pathogenesis of hemorrhagic shock induced ARDS/MOF.