Aortic stenosis (AS) is the most common valvular heart disease, and it is suspected that atherosclerotic mechanisms are involved in the development of this disorder. Therefore, the retention of lipids within the aortic valve may play a role in the pathobiology of AS. In this study, a gene expression microarray experiment was conducted on human aortic valves with and without AS. The expression levels of transcripts encoding proteoglycans and enzymes involved in lipid retention were compared between the two groups. The microarray results were subsequently replicated in a cohort of 87 AS valves and 36 control valves. In addition, the interaction between proteoglycan and lipid-modifying enzyme was documented in isolated valve interstitial cells (VICs). The microarray results indicated that only biglycan (BGN) and phospholipid transfer protein (PLTP) were overexpressed in the AS valves. These results were then confirmed by quantitative PCR. The immunohistochemical analysis revealed a colocalization of BGN, PLTP, and Toll-like receptor-2 (TLR 2) in AS valves. In vitro, we showed that BGN induces the production of PLTP in VICs via the stimulation of TLR 2. Thus, increased accumulation of BGN in AS valves contributes to the production of PLTP via TLR 2. These results suggest that intricate links between valve matrix proteins, inflammation, and lipid retention are involved in the pathobiology of AS.