Background: To determine the impact of circulating miRNA and protein activity on the severity of ascending aortic dilatation in patients with bicuspid (BAV) and tricuspid aortic valve (TAV). Methods: By reverse transcription polymerase chain reaction, exosomal circulating expression levels (versus healthy aorta) of miRNAs and absolute levels of transforming growth factor β (TGF-β), matrix metalloproteinases (MMP-2, -3 and -9), tissue inhibitors (TIMP-1, -2, -3 and -4), and soluble receptors for advanced glycation end products AGEs (sRAGE) were evaluated in ascending dilated aortas of 71 patients with different valve morphotype. Results: Less-dilated ascending aorta exhibited a specific miRNA signature (i.e., miR-126 miR-15b, miR-195, miR-221, miR24, miR-30b and miR-320a), which was statistically different from that of severely-dilated ascending aorta. Among these analytes, miR-15b was the most significant (p < 0.001) and resulted as an independent predictor of aortic dilatation (β = −1.099, p = 0.041). When patients were grouped according to aortic valve morphology, miRNAs and protein proteolytic activity were different between BAV and TAV in the expression level of miR-133a, miR-155, miR-320a, miR-34a (#000425) , miR-34a (#000426) , miR-494 and measurements of TGF-β and MMP-3, MMP-9, TIMP-4. The circulating level of miR-34a (#000426) was negatively correlated to the aortic wall elasticity of bicuspid patients (R = −0.653 and p = 0.011), suggesting an apparent different mechanism of aortic wall degeneration specific for BAV. Conclusions: Taken these biomarkers together, we demonstrated that the severity of aortic size and valve morphology differently modulates miRNA analytes and protein proteolytic activity in patients with ascending aortic dilatation, and this may be useful to design new therapies that inhibit miRNAs.