EQUINE NEUTROPHIL APOPTOSIS IN INFLAMMATORY CONDITIONS
Horses are at high risk to develop systemic inflammation due to the release of bacterial endotoxin from an inflamed gastrointestinal tract. Neutrophils are critical for mounting an immune response to bacterial endotoxins. Neutrophil activation following engagement of bacterial endotoxin expands their lifespan through suppression of their constitutive apoptosis. The prolonged lifespan of neutrophils propagates acute inflammation and delays the resolution of inflammation. Since equine neutrophil lifespan has not been well-studied, I investigated the occurrence of equine neutrophil apoptosis in vitro and in vivo. First, I investigated the effect of Escherichia coli lipopolysaccharide (LPS) treatment on the occurrence of equine neutrophil apoptosis in vitro. LPS treatment delayed in vitro equine neutrophil apoptosis in a dose-dependent manner at concentrations of 0.1-10 μg/ml through toll-like receptor (TLR)-4 signaling and down-regulation of the intrinsic pathway of apoptosis, specifically through reduced caspase-9 activity. Next, I found that ex vivo neutrophil apoptosis was delayed in two models of intestinal inflammation, jejunal ischemia and reperfusion (IR) and oligofructose-induced colitis, through down-regulation of both the intrinsic and extrinsic apoptosis pathways via reduced caspase-3, -8, and -9 activities. Pulmonary intravascular macrophages (PIMs) depletion with systemic gadolinium chloride (GC) prevented the prolongation of ex vivo neutrophil lifespan in horses undergoing jejunal IR through modulation of caspase-3, -8 and -9 activities. PIM depletion in IR horses resulted in an earlier and greater increase in tumor necrosis factor-alpha and a concomitant decrease in interleukin-10 to suggest an enhanced systemic pro-inflammatory response. I examined the effect of neutrophil concentration and co-incubation with aged, apoptotic neutrophils on the occurrence of neutrophil apoptosis in vitro. Neutrophil apoptosis was delayed with increasing concentrations of neutrophils in vitro, which may contribute to delayed neutrophil apoptosis in systemic inflammation. However, co-incubation with aged, apoptotic neutrophils did not alter in vitro neutrophil lifespan. Taken together, the data show that LPS delays equine neutrophils apoptosis in vitro in a TLR4-dependent manner through inhibition of caspase-9. Ex vivo neutrophil apoptosis was also delayed with systemic inflammation via down-regulation of caspase activity. A novel finding of this work was the reversal of delayed neutrophil apoptosis by depletion of PIMs in horses experiencing intestinal IR.
DegreeDoctor of Philosophy (Ph.D.)
DepartmentVeterinary Biomedical Sciences
ProgramVeterinary Biomedical Sciences
CommitteeDuke, Tanya; Loewen, Matthew; Townsend, Hugh; Honamorooz, Ali
Copyright DateNovember 2015
horse, neutrophil, apoptosis, inflammation, intestine, colic