Reperfused Gut Elaborates PAF That Chemoattracts and Primes Neutrophils

Abstract

Our in vivo model of mesenteric ischemia/reperfusion (I/R) has shown that the gut serves as a priming bed for neutrophils (PMN). Activation of phospholipase A 2 (PLA 2) during ischemia temporally precedes PMN sequestration in the gut and the appearance of primed PMN in the portal circulation. Therefore, we hypothesized that reperfused gut secretes platelet activating factor (PAF) via PLA 2 activation that is responsible for increased PMN chemotaxis and priming for superoxide (O - 2) generation. Sprague-Dawley rats underwent gut ischemia/reperfusion (45 min SMA occlusion/2 hr reperfusion) or sham laparotomy. Distal ileum was harvested, rinsed with bacteriostatic saline/neomycin, and incubated for 1 hr at 37°C in RPMI 1640 and the cellfree supernatant was collected. Normal human PMNs, isolated by plasma-Percoll gradients, were pretreated with or without a PAF receptor antagonist (WEB 2170). Chemotaxis toward gut supernatant was then measured by the agarose method. Additionally, PMNs were preincubated with or without WEB 2170 and their O - 2 release in response to 1 μ M FMLP was measured by the V max of SOD-inhibitable cytochrome c reduction. Reperfused gut produced a chemotactic index of 2.1 ± 0.1 compared to 0.2 ± 0.9 following sham laparotomy ( P < 0.05); this was reduced to 0.4 ± 0.9 with PAF receptor blockade. Similarly, gut I/R supernatant primed PMNs for O - 2 ( P < 0.05) compared to laparotomy, and this effect was abrogated by a PAF antagonist. These data suggest that reperfused gut can elaborate PAF which chemoattracts and primes PMNs for O - 2 generation.