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Background: Although calcific aortic stenosis is common, calcification of the other three heart valves is not. The aortic valve interstitial cell (VIC) has been implicated in the pathogenesis of aortic stenosis. Proinflammatory stimulation of aortic VICs induces an osteogenic and inflammatory phenotypic change. We hypothesized that the VICs of the other heart valves do not undergo these changes. Using isolated human VICs from normal aortic, mitral, pulmonary, and tricuspid valves, our purpose was to compare the osteogenic response to proinflammatory stimulation via toll-like receptor 4 (TLR-4)., Materials and Methods: Aortic, pulmonic, mitral, and tricuspid (n=4 for each valve type) VICs were isolated from hearts valves explanted from patients undergoing cardiac transplantation. Cells were cultured and grown to confluence in passage 2-6 before treatment with Lipopolysaccharide (LPS) (100-200 ng/mL) for 24 or 48 h. Cells were characterized by immunofluorescent staining. TLR-4 expression was analyzed (immunoblotting, flow cytometry). Bone morphogenetic protein 2 and intercellular adhesion molecule 1 production were determined (immunoblotting). Monocyte chemoattractant protein 1 levels were determined by enzyme-linked immunosorbent assay. Statistics were by Mann-Whitney U test., Results: TLR-4 stimulation induced bone morphogenetic protein 2 production only in aortic VICs (P<0.05). Intra-cellular adhesion molecule 1 production and monocyte chemoattractant protein 1 secretion increased in a similar fashion among TLR-4-stimulated VICs from all four valves., Conclusions: Proinflammatory stimulation induces an osteogenic phenotype in aortic VICs but not mitral, pulmonic, or tricuspid VICs. We conclude that this differential osteogenic response of aortic VICs contributes to the pathogenesis of calcific aortic stenosis., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: The aortic valve interstitial cell (AVIC) has been implicated in the pathogenesis of calcific aortic stenosis. When appropriately stimulated, AVICs undergo a phenotypic change from that of a myofibroblast to that of a bone-forming-like cell. An elevated blood level of low-density lipoprotein (LDL) cholesterol is a clinical risk factor for aortic stenosis, and oxidized LDL (ox-LDL) cholesterol has been consistently found in calcified aortic valve leaflets. However, whether it plays a role in the pathogenesis of aortic stenosis is unknown. The process of aortic valve leaflet calcification has been associated with the deposition of calcium phosphate, mediated in part by the phosphate inorganic transporter 1 (PiT-1), a sodium-phosphate ion cotransporter. Therefore, we hypothesized that ox-LDL induces an osteogenic change in human AVICs marked by the induction of PiT-1. Using isolated human AVICs, the purpose of the present study was to examine the effect of ox-LDL on the expression of PiT-1 and the osteogenic factor bone morphogenetic protein 2 (BMP-2), which is a protein necessary for bone formation., Methods: Human AVICs were isolated from nonstenotic aortic valves obtained from the explanted hearts of patients undergoing cardiac transplantation (n = 4) and grown in culture. The cells were treated with serum-free media, serum-free media with dimethyl sulfoxide (vehicle control), 40 μg/mL of ox-LDL, or 40 μg/mL of ox-LDL plus 2.5 mM phosphonoformate hexahydrate acid. Phosphonoformate hexahydrate acid is a competitive inhibitor of PiT-1 by mimicking inorganic phosphate. Cell lysis was performed at 24 h after treatment. Cell lysates were analyzed using immunoblot and densitometry for PiT-1 and BMP-2. Statistical analysis was performed using analysis of variance. P < 0.05 was significant., Results: ox-LDL stimulation of AVICs induced an increase in PiT-1 and BMP-2. ox-LDL induced increased production of the phosphate transporter, PiT-1, and the osteogenic factor, BMP-2. Inhibition of PiT-1 with phosphonoformate hexahydrate acid prevented ox-LDL-induced BMP-2 expression., Conclusions: These data offer mechanistic insight into the pathogenesis of calcific aortic stenosis., (Copyright © 2013 Elsevier Inc. All rights reserved.)