Cardiovascular iron load is the leading cause of morbidity and mortality in beta-thalassemia major (β-TM). However, many patients remain asymptomatic until the late stage. In this cross-sectional study, we investigated the role of three-dimensional (3D) echocardiography and endothelial dysfunction parameters in asymptomatic β-TM patients, and the relationship between these parameters and cardiac magnetic resonance imaging (MRI) T2* value.A total of 51 asymptomatic β-TM patients receiving regular blood transfusions were divided into two groups based on cardiac MRI-T2* values (MRI-T2*20 ms and ≥20 ms), which MRI-T2*20 ms determines myocardial iron load and evaluated by two-dimensional (2D) and 3D-echocardiography including endothelial dysfunction parameters. The relationships between ferritin levels, 2D and 3D-echocardiography measurements, endothelial dysfunction parameters, and cardiac MRI-T2* values were investigated.All left ventricle ejection fraction (LVEF) obtained by 2D-echocardiography were normal (≥50%). LVEF-3D (53.25 ± 2.33 vs. 58.81 + 1.02), SDI12 (6.53 ± 0.56 vs. 2.85 + 0.48), and SDI16 (7.65 ± 0.75 vs. 3.26 + 0.49) were significantly different and negatively correlated between groups with MRI-T2*20 ms and ≥20 ms, respectively. Flow-mediated dilatation (FMD) (6.08% ± 0.34% vs. 14.46% ± 1.12), aortic strain (7.79% ± 2.19% vs. 12.76% ± 4.19), ferritin levels were significantly different and negatively correlated between groups with MRI-T2*20 ms and ≥20 ms, respectively. Higher ferritin, SDI12/16 were significant independent predictors of MR-T2* 20 ms. SDI16 5.5, SDI12 4.3 predicted MRI-T2*20ms with a sensitivity of 92%, specificity of 81% (AUC 0.85, P .001), and sensitivity of 92%, specificity of 78% (AUC 0.83, P .001), respectively.SDI12/16 calculated by 3D-echocardiography may be a promising predictors of cardiovascular iron load and, decreased LVEF-3D, FMD, and aortic strain might be good indicators of subclinical cardiovascular involvement of β-TM.