BackgroundAortic stenosis (AS) is a chronic inflammatory disease, and calcification plays an important role in the progression of the disease. Galectin‐3 (Gal‐3) is a proinflammatory molecule involved in vascular osteogenesis in atherosclerosis. Therefore, we hypothesized that Gal‐3 could mediate valve calcification in AS.Methods and ResultsBlood samples and aortic valves (AVs) from 77 patients undergoing AV replacement were analyzed. As controls, noncalcified human AVs were obtained at autopsy (n=11). Gal‐3 was spontaneously expressed in valvular interstitial cells (VICs) from AVs and increased in AS as compared to control AVs. Positive correlations were found between circulating and valvular Gal‐3 levels. Valvular Gal‐3 colocalized with the VICs markers, alpha‐smooth muscle actin and vimentin, and with the osteogenic markers, osteopontin, bone morphogenetic protein 2, runt‐related transcription factor 2, and SRY (sex‐determining region Y)‐box 9. Gal‐3 also colocalized with the inflammatory markers cd68, cd80 and tumor necrosis factor alpha. In vitro, in VICs isolated from AVs, Gal‐3 induced expression of inflammatory, fibrotic, and osteogenic markers through the extracellular signal‐regulated kinase 1 and 2 pathway. Gal‐3 expression was blocked in VICs undergoing osteoblastic differentiation using its pharmacological inhibitor, modified citrus pectin, or the clustered regularly interspaced short palindromic repeats/Cas9 knockout system. Gal‐3 blockade and knockdown decreased the expression of inflammatory, fibrotic, and osteogenic markers in differentiated VICs.ConclusionsGal‐3, which is overexpressed in AVs from AS patients, appears to play a central role in calcification in AS. Gal‐3 could be a new therapeutic approach to delay the progression of AV calcification in AS.

ObjectiveWe aim to analyse sex-specific differences in aortic valves (AVs) and valve interstitial cells (VICs) from aortic stenosis (AS) patients.Approach and Results238 patients with severe AS undergoing surgical valve replacement were recruited. Two hundred and two AVs (39.1% women) were used for ex vivo analyses and 36 AVs (33.3% women) for in vitro experiments. AVs from men presented increased levels of the inflammatory molecules interleukin (IL)-1β, IL-6, Rantes, and CD45. Oxidative stress (eNOS, myeloperoxidase, malondialdehyde and nitrotyrosine) was upregulated in male AVs. Concerning fibrosis, similar levels of collagen type I, decreased levels of collagen type III and enhanced fibronectin, active Lox-1 and syndecan-1 expressions were found in AVs from men compared with women. Extracellular matrix (ECM) remodeling was characterized by reduced metalloproteinase-1 and 9 expression and increased tissue inhibitor of metalloproteinase-2 expression in male AVs. Importantly, osteogenic markers (bone morphogenetic protein-9, Rank-L, osteopontin, periostin, osteocalcin and Sox-9) and apoptosis (Bax, Caspase 3, p53, and PARP1) were enhanced in AVs from men as compared to women. Isolated male VICs presented higher myofibroblast-like phenotype than female VICs. Male VICs exhibited increased inflammatory, oxidative stress, fibrotic, apoptosis and osteogenic differentiation markers.ConclusionsOur results suggest that the mechanisms driving the pathogenesis of AS could be different in men and women. Male AVs and isolated VICs presented more inflammation, oxidative stress, ECM remodeling and calcification as compared to those from women. A better knowledge of the pathophysiological pathways in AVs and VICs will allow the development of sex-specific options for the treatment of AS.