Characterization of cellular density and determination of neointimal extracellular matrix constituents in human lower extremity vein graft stenoses

Abstract

Arterial restenosis has been attributed to a hyperproliferative smooth muscle cell response. Paradoxically, studies of human coronary atherectomy and vein graft stenotic lesions have demonstrated a relatively low nuclear proliferative rate with the majority of the neointimal mass consisting of extracellular matrix. The purpose of the present study was to characterize the cellular density and determine the relative composition of the extracellular matrix protein constituents in stenotic, human lower extremity vein-bypass graft lesions. Methods: Duplex surveillance of 148 consecutive infrainguinal bypass grafts identified 17 patients with 22 pre-occlusive autogenous vein graft stenoses (mean graft age 7 months). Morphological analyses of these stenotic lesions were compared with excised samples of 20 greater saphenous vein segments taken at the time of graft implantation from matched control patients. Intimal and medial areas were compared and cell density was determined with fluorescent nuclear (Bisbenzimide) staining. Differential light microscopy with pentachrome staining was performed to determine the relative percent composition of intimal matrix constituents by stereological morphometric (point-count) techniques. Results: The intimal areas for control and stenotic vein segments were 1.64×10 6 μm 2 and 3.85×10 6 μm 2, P<0.0001, whereas the intimal nuclear densities (cells/unit volume) were 1.42×10 3 and 1.70×10 3 cells/ μm 2, P=0.03, respectively. The corresponding medial area and medial nuclear densities were 5.01×10 6 μm 2, 3.31×10 6 μm 2; P=0.08, and 2.27×10 3, 3.29×10 3; P=0.001, for control and stenotic specimens, respectively. The intima:media area ratios were much greater, whereas the intimal and medial cell densities were only slightly greater in the stenotic compared with control veins. The relative composition of intimal extracellular matrix proteins of stenotic vein graft segments consisted of 21% cellular (fibrous) material, 33% collagen, and 46% glycosaminoglycan ground substance. Conclusion: The intimal lesions responsible for lower extremity vein graft stenosis are more hypertrophic than hyperplastic. Therapies aimed at preventing arterial and vein graft restenosis may thus need to inhibit matrix biosynthetic processes in addition to cellular proliferation.