Circulating neutrophil priming and systemic inflammation in limb ischaemia-reperfusion injury

Recruitment and activation of neutrophils is a key step in the development of local and systemic injury in lower limb ischaemia-reperfusion. We hypothesis that increased circulating neutrophil priming is responsible for systemic inflammation.

Anaesthetised ventilated swine (n = 6 per group) underwent mid-line laparotomy and were randomised to control group or bilateral external iliac artery occlusion for two hours followed by two and a half hours reperfusion (I/R group). Using luminol, respiratory burst activity was assayed with a BioOrbit Luminometer to detect whole blood chemiluminescence (CL) by stimulation with phorbol 1,2-myristate 1,3-acetate (PMA) in the absence or presence of tumour necrosis factor (TNF) respectively. PMN priming is expressed as the ratio of whole blood CL in the presence of TNF to that without. We measured plasma interleukin(IL)-6 and tumour necrosis factor alpha by bioassay as a measure of systemic inflammation. The alveolar-arterial (A-a) gradient was measured using the formula [(A-a)gradient = fraction inspired O2 x 710-(arterial pCO2/0.8)-arterial pO2], it is a measure of lung function, a large gradient being indicative of impaired oxygen transport and hence lung injury.

Lower limb I/R caused significantly greater PMN priming, 0.83 +/- 0.14, compared to control group, 0.22 +/- 0.04, (p < 0.001). Plasma IL-6, a reliable indicator of systemic inflammation, was significantly increased in I/R group after two and a half hours of reperfusion, 1295.0 (833.9-2073.0) pg/L, compared to control, 382.9 (367.4-568.3) pg/L, (p < 0.005). Plasma tumour necrosis factor alpha was significantly elevated after one hour of reperfusion in the I/R group, 86.8 (48.7-106.6) pg/ml, compared to the control group, 32.7 (0.9-42.8) pg/ml, (p < 0.01). (A-a) gradient was significantly increased after IRI, 407.97 +/- 53.13, compared to the control, 183.19 +/- 45.75, (p < 0.005). Mean pulmonary artery pressure was significantly greater after IRI, 38.80 +/- 4.87 mmHg, compared to control, 27.86 +/- 1.92 mmHg, (p < 0.005). Data represents mean +/- standard error mean or median (interquartile range), statistical comparisons using one-way Anova with Student's "t"-test and Kruskall-Wallis Anova with the Mann-Whitney U test.

Priming of neutrophils increases their circulating respiratory burst activity and ability to induce tissue injury. Systemic PMN priming during hind limb ischaemia-reperfusion injury is associated with the systemic inflammatory response syndrome.