PECAM-1/IgG attenuates peroxynitrite-mediated extremity reperfusion injury.
OBJECTIVE
Neutrophil transendothelial migration, a key feature of skeletal muscle ischemia and reperfusion (I/R) injury, is mediated by the platelet endothelial cell adhesion molecule-1 (PECAM-1). Peroxynitrite anion, a toxic product of neutrophil superoxide anion and nitric oxide, contributes to oxidative skeletal muscle injury and can be quantified by measurement of protein tyrosine nitration after I/R. This study hypothesizes that administration of the PECAM-1/IgG antibody chimera will inhibit peroxynitrite-mediated injury after I/R.


METHODS
The study was composed of five groups: an I/R group (n = 4), a sham treatment group anesthetic control (n = 3), a treatment group receiving the PECAM-1/immunoglobulin G (IgG) antibody chimera with I/R (n = 9), a treatment group receiving human IgG with I/R as an antibody control (n = 6), and a treatment group receiving normal saline solution with I/R as a vehicle control (n = 5). The right hind limb in male New Zealand white rabbits was rendered ischemic by occluding the iliac and femoral arteries for 3 hours, followed by 2 hours of reperfusion (I/R). Sham-treated rabbits underwent arterial dissection without arterial occlusion. PECAM-1/IgG-treated rabbits and IgG-treated rabbits received an infusion of 1 mg/kg in normal saline solution 20 mL via an ear vein catheter during the last 5 minutes of ischemia and the first 15 minutes of reperfusion. Saline solution-treated rabbits similarly received normal saline solution 20 mL. The anterior tibialis muscle was harvested after reperfusion. Immunohistochemical staining for nitrotyrosine was performed with monoclonal antinitrotyrosine antibodies and fluorescently labeled secondary antibodies. Computed morphometric study was performed to calculate relative fluorescence scores for each histologic section. Averaged fluorescence scores were analyzed by one-way analysis of variance with Bonferroni post hoc comparison.


RESULTS
The averaged fluorescence scores (mean +/- SEM) for the sham-treated (2.88 +/- 0.78) and PECAM-1/IgG-treated (6.16 +/- 0.43) groups demonstrated a significant reduction in quantitative fluorescence compared with the IgG- (15.17 +/- 2.01) and saline solution-treated (17.46 +/- 3.71) control groups, and the I/R-treated (18.52 +/- 3.00) group, (P <.05).


CONCLUSIONS
These results suggest that PECAM-1/IgG diminishes peroxynitrite-mediated oxidative skeletal muscle injury by inhibiting neutrophil transendothelial migration and may therefore prove a useful therapeutic agent in the treatment of reperfusion injury.
Farooq
M M
MM
Department of Vascular Surgery, Medical College of Wisconsin, Milwaukee, USA. mfarooq@mednet.ucla.edu
Serra
A
A
Newman
P J
PJ
Cambria
R A
RA
Seabrook
G R
GR
Towne
J B
JB
Freischlag
J A
JA
D000818:Animals; D006614:Hindlimb / Q000098:blood supply*; D007074:Immunoglobulin G / Q000627:therapeutic use*; D008297:Male; D009566:Nitrates; D016877:Oxidants; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000627:therapeutic use*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000627:therapeutic use; D015427:Reperfusion Injury / Q000209:etiology / Q000517:prevention & control*
D007074:Immunoglobulin G; D009566:Nitrates; D016877:Oxidants; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D011993:Recombinant Fusion Proteins; C039642:peroxynitric acid