Neutrophil, not macrophage, infiltration precedes neointimal thickening in balloon-injured arteries

Abstract

Macrophages are abundant after stent-induced arterial injury. Inhibition of macrophage recruitment blocks neointimal growth in this model. In contrast, after superficial injury from balloon endothelial denudation, macrophages are sparse. However, many anti-inflammatory therapies remain effective against neointimal growth after balloon injury. To investigate further the role of leukocytes after injury, 41 New Zealand White rabbits underwent iliac artery balloon denudation. In 18, subcutaneous pumps were placed to deliver intravenous heparin (0.3 mg/kg per hour). Arteries were harvested at 6 hours and at 3, 7, and 14 days. In 8 animals, either M1/70 (a monoclonal antibody [mAb] against adhesion molecule Mac-1) or a nonspecific IgG was given (5 mg/kg IV bolus and then 1 mg/kg SC QOD), and arteries were harvested at 6 hours and 3 days. Computer-aided morphometry was performed as was immunohistochemistry to assess smooth muscle cell (SMC) proliferation (bromodeoxyuridine-positive cells), neutrophil content (RPN357, mAb against rabbit neutrophil/thymocyte), and macrophage content (RAM-11, mAb against rabbit macrophage). Heparin inhibited neointimal growth at 7 and 14 days (64% and 32.5% reduction, respectively; P:<0.05). Neutrophils were observed in the media early after balloon injury, and heparin and M1/70 inhibited this infiltration (82% and 83% reduction, respectively; P:<0.05 each) with a coincident inhibition of medial SMC proliferation at 3 days (49% and 84% reduction, respectively; P:<0.05 each). Macrophages were absent at all time points. Neutrophil, but not macrophage, infiltration occurs early after endothelial denudation. Inhibition of this process is associated with a reduction in medial SMC proliferation. These data suggest a central role for neutrophils in restenosis and help to explain prior reports of an inhibitory effect of anti-inflammatory therapies on neointimal growth after balloon injury.