Cell-driven processes are now considered of relevance for the pathogenesis of aortic stenosis. In particular, during calcific valve degeneration, interstitial valve cells (VIC) resident in the leaflet can acquire an osteogenic/pro-calcific profile and actively contribute to matrix mineralization. The proteomic study described in this chapter is undertaken to investigate modifications in the proteome of bovine aortic VIC acquiring a calcifying phenotype. This approach can be useful to clarify cellular pathways involved in VIC pro-calcific differentiation and identify innovative therapeutic targets.