Aortic valve stenosis (AS) is characterized by extensive calcification of the aortic valve leaflets and infiltration of inflammatory cells. Activated mast cells (MCs) may participate in the induction of fibrosis and calcification with ensuing valve stiffening. We sought to investigate whether the number of MCs within stenotic aortic valves is associated with the severity of AS. We studied 43 patients (19 men, 24 women) with dominant AS (age, 64.2 ± 5.9 years; mean transvalvular pressure gradient, 62.11 ± 24.47 mmHg) without atherosclerotic vascular disease, undergoing elective aortic valve replacement. MCs were detected in the excised valves by immunostaining. Aortic valves from five healthy subjects obtained on autopsy served as negative controls. The number of tryptase- and chymase-positive MCs was increased in AS valves compared with the control valves (6.9 [2.3-18.9]/mm(2) vs. 0.7 [0-2.2]/mm(2), P = 0.0001 and 3.2 [2.1-9.4]/mm(2) vs. 0.3 [0-1.9]/mm(2), P = 0.002, respectively). MCs that colocalized with macrophages and neovessels were detected mainly in the calcified regions of the leaflets. The number of MCs positively correlated with maximal (r = 0.73, P < 0.0001) and mean (r = 0.78, P < 0.0001) gradients and maximal aortic jet velocity (r = 0.68, P = 0.0005). An inverse correlation with aortic valve area (r = -0.71, P = 0.0001) was also observed. Multivariate regression analysis revealed that MC number and valve thickness were significantly associated with mean transvalvular gradient (R (2) = 0.74, P < 0.000001) in AS patients. Increased MC number within human stenotic aortic valves is associated with the severity of AS.