Clinical in vitro endothelialization has been shown to increase the patency of synthetic vascular grafts. The shear stress resistance of the cultured autologous endothelium represents a crucial cornerstone of the concept. We investigated whether an enrichment of the precoating matrix with adhesion sites can augment endothelial cell attachment. Adult human saphenous vein endothelial cells (AHSVECs) were seeded confluently ([58 +/- 11] x 10(3) AHSVECs/cm2) onto 10-cm-long ePTFE (expanded polytetrafluorethylene) vascular grafts (n = 24) precoated with commercial clinically approved fibrin gel (Tisseal) containing various concentrations of cross-linked RGD peptide (0.0, 4.0, 8.0, or 16.0 mg of RGD per milliliter of Tisseal fibrinogen component). Endothelialized grafts were postcultivated for 9 days before they were exposed to a pulsatile circulation model mimicking peak physiological shear stress conditions of the femoral artery (12 dyn/cm2; min/max, -60/+28 dyn/cm2). Cell loss after 24 h was quantitatively determined by image analysis of vital stains. Initial 24-h cell loss was 27.2 +/- 1.7% in grafts precoated with the non-RGD-enriched fibrin matrix. In contrast, cell loss was significantly less on fibrin containing 4.0 mg of RGD peptide per milliliter of Tisseal fibrinogen component (13.3 +/- 7.9%; p < 0.05). Cell loss on fibrin containing 8 and 16 mg of RGD per milliliter of Tisseal fibrinogen component was 41.0 +/- 27.4 and 43.0 +/- 23.2% (p > 0.05), respectively. We conclude that low concentrations of RGD peptide cross-linked into commercial fibrin matrices used for clinical in vitro lining of vascular grafts led to significantly increased endothelial cell retention. The failure of higher RGD concentrations to enhance endothelial cell attachment may be explained by competitive binding of endothelial cells to non-cross-linked RGD.