To evaluate the impact of histological factors on the appearance of the wall of the coronary artery by intravascular ultrasound (IVUS), we performed an in vitro study of 34 coronary artery segments from eight autopsied patients. We assumed the coronary cross section to be divided into four equal parts, and assessed the quadrants with maximal and minimal wall thickness by IVUS (30 MHz; 4.3 Fr; 1800 rpm) and by a histological study. The histological layer thickness and composition were also evaluated in terms of their contribution to the appearance of the ultrasound layer. Fifty-eight quadrants were clearly visible on ultrasound. A three-layered appearance, with inner echogenic, subjacent sonolucent, and outer echogenic layers, was observed in 32 quadrants, while 26 quadrants showed a two-layered appearance with inner and outer echogenic layers. The thickness of the inner echogenic layer (0.77 +/- 0.38 mm) was moderately correlated with the intimal thickness (0.51 +/- 0.45 mm; r = 0.85, standard error of estimate [SEE] = 0.24 mm); however, the correlation was significantly improved when the thickness of the inner echogenic plus sonolucent layers (0.89 +/- 0.47 mm) was compared with that of the intima plus media (0.69 +/- 0.47 mm; r = 0.94, SEE = 0.15 mm; P = 0.012 between the coefficients). Discriminant analysis showed that intimal hyalinization, associated with intimal thickening, was strongly related to the presence of the three-layered appearance on IVUS (F to enter 40.0, P < 0.0001). These results indicate that the ultrasound layered appearance of human coronary arteries varies with histological alterations. We suggest that the thickness of the inner echogenic plus sonolucent layers on IVUS represents the intimal plus medial thickness observed by histology, and that the use of this value may be appropriate in the assessment of coronary wall thickening associated with atherosclerosis.