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Short-axis magnetic resonance images of the cardiac left ventricle, acquired in multiple slices and phases, may be used for the quantitative assessment of regional wall parameters. Conventional two-dimensional(2-D) methods for wall thickness measurement rely on information within one imaging plane, which may result in overestimation of the true thickness depending on the local direction of myocardial wall with respect to the imaging plane.To perform wall thickness measurements truly perpendicular to the myocardial wall, a three-dimensional(3-D) wall thickness calculation algorithm has been developed based on the 2-D improved centerline method. An evaluation was performed on left ventricular-shaped software phantoms, and on the magnetic resonance (MR) imaging data obtained from 20 healthy individuals.The 3-D method applied to software phantoms with an angulation within 20° of the true short-axis orientation demonstrated only a 1.6% overestimation of wall thickness at the mid to low slices, and a 10.6% error at the apex(2-D measurements: 8.1% and 28.6%, respectively). Three-dimensionally calculated wall thickness in the healthy individuals was systematically and significantly smaller than corresponding 2-D wall thickness (by 11.2%, 8.7%, and 2.6% at the apical, low, and mid slices, respectively).