Nitric oxide is a radical with vasodilating properties that protects tissues from neutrophil-mediated
ischemia-reperfusion injury in the heart and intestine. Previous studies in our laboratory suggested that
L-arginine, a
nitric oxide precursor, can protect skin flaps from
ischemia-reperfusion injury. In this study, we examined the effects of
L-arginine on the survival of
myocutaneous flaps in a large animal model and established whether this effect was mediated by
nitric oxide and neutrophils. Two superiorly based 15 x 7.5 cm epigastric myocutaneous
island flaps were dissected in 15 Yorkshire pigs weighing 45 to 50 kg. One of the flaps was subjected to 6 hours of arterial
ischemia and then reperfused for 4 hours (
ischemia-reperfusion flaps), whereas the other flap was used as a non-ischemic control (non-
ischemia-reperfusion flaps). The flaps were divided into four groups: control non-
ischemia-reperfusion flaps that received only saline (group I);
ischemia-reperfusion flaps that were treated with saline (group II); and flaps treated with either
L-arginine (group III) or Nomega-nitro-
L-arginine methylester (
L-NAME), a
nitric oxide synthase competitive inhibitor, plus
L-arginine in equimolar amounts (group IV). These drugs were administered as an intravenous bolus 10 minutes before the onset of reperfusion, followed by a 1-hour continuous
intravenous infusion. Full-thickness muscle biopsies were taken at baseline, 3 and 6 hours of
ischemia, and 1 and 4 hours of reperfusion. The biopsies were evaluated by counting neutrophils and measuring myelo-
peroxidase activity. At the end of the experiment, skeletal muscle
necrosis was quantified using the
nitroblue tetrazolium staining technique, and a full-thickness biopsy of each flap was used for determination of water content. Statistical analysis was performed using analysis of variance and the Newman-Keuls test. Non-
ischemia-reperfusion flaps showed no muscle
necrosis.
Ischemia-reperfusion flaps treated with saline had 68.7 +/- 9.1 percent
necrosis, which was reduced to 21.9 +/- 13.6 percent with
L-arginine (p < 0.05).
L-NAME administered concomitantly with
L-arginine demonstrated a
necrosis rate similar to that of saline-treated
ischemia-reperfusion flaps (61.0 +/- 17.6 percent). Neutrophil counts and
myeloperoxidase activity after 4 hours of reperfusion were significantly higher in
ischemia-reperfusion flaps treated with
L-NAME and
L-arginine as compared with the other three groups (p < 0.05). Flap water content increased significantly in
ischemia-reperfusion flaps treated with saline and
L-NAME plus
L-arginine versus non-
ischemia-reperfusion flaps (p < 0.02) and
L-arginine-treated
ischemia-reperfusion flaps (p < 0.05). There was no difference in flap water content between
ischemia-reperfusion flaps treated with
L-arginine and non-
ischemia-reperfusion flaps. Administration of
L-arginine before and during the initial hour of reperfusion significantly reduced the extent of flap
necrosis, neutrophil accumulation, and
edema due to
ischemia-reperfusion injury in a large animal model. This protective effect is completely negated by the use of the
nitric oxide synthase blocker
L-NAME. The mechanism of action seems to be related to
nitric oxide-mediated suppression of
ischemia-reperfusion injury through neutrophil activity inhibition.