Depletion of pulmonary intravascular macrophages rescues inflammation-induced delayed neutrophil apoptosis in horses

The objective of this study was to determine the effect of pulmonary intravascular macrophage depletion on systemic inflammation and ex vivo neutrophil apoptosis, using an experimental model of intestinal ischemia and reperfusion injury in horses. Neutrophils were isolated before and after surgery from horses that were randomized to 3 treatment groups: sham celiotomy (CEL, n=4), intestinal ischemia and reperfusion (IR, n=6), intestinal ischemia and reperfusion with gadolinium chloride treatment to deplete pulmonary intravascular macrophages (PIMs, IRGC, n=6). Neutrophil apoptosis was assessed with Annexin V and propidium iodide staining quantified with flow cytometry and caspase-3, -8, and -9 activities in neutrophil lysates. All horses experienced a systemic inflammatory response following surgery. Following surgery, ex vivo neutrophil apoptosis was significantly delayed after 12 or 24 h in culture, except in IRGC horses (12 h: CEL: P = 0.03, IR: P = 0.05, IRGC: P =0.2; 24 h: CEL: P = 0.001, IR: P = 0.004, IRGC: P = 0.3). Caspase-3, -8, and -9 activities were significantly reduced in neutrophils isolated after surgery and cultured for 12 h in IR horses, but not IRGC horses (IR caspase-3: P = 0.002, IR caspase-8: 0.002 P =, IR caspase-9: P = 0.04). Serum TNF-α concentration were increased in IRGC horses for 6-18 h following jejunal ischemia. Following surgery, ex vivo equine neutrophil apoptosis was delayed via down-regulation of caspase activity, which was ameliorated by PIM depletion potentially via upregulation of TNF-α.