Glycolysis and hexosamine biosynthesis regulate inflammatory protein expression and maturation and key steps of monocyte recruitment by human aortic valve cells

Inflammation and metabolism reprogramming are hallmarks of calcific aortic valve disease (CAVD). Recent studies link hyperglycolysis, inflammation, and calcification in aortic valve interstitial cells (VICs). The metabolism of aortic valve endothelial cells (VECs) has received less attention despite the fact that both resident valve cells are exposed to alike inflammatory clues involved in the biosynthesis of pathologically relevant glycoproteins at early stages of CAVD. Given this, we investigated the outcomes of glucose metabolism rewiring on glycoprotein maturation and monocyte adhesion in human resident valve cells. Real-time metabolic analysis revealed that basal VECs are more glycolytic than VICs. Also, VECs and VICs exposed to inflammatory stimuli exhibited a distinct rewiring, with VECs shifting to a more energetic metabolism, despite a similar upregulation of glycolytic genes. Blunting glucose metabolism in VICs and VECs inhibited inflammatory routes canonically associated with glycolysis, and the expression of proteins associated with the inflammatory response, like interleukin 6 and cyclooxygenase-2. Moreover, the expression and posttranslational modifications of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 showed glucose dependency and sex-related differences. The ensuing process of monocyte-valve cell adhesion was glucose-dependent. Notably, inhibiting the branch route of hexosamine biosynthesis with 6-diazo-5-oxo-l-norleucine (DON) and N-glycosylation by tunicamycin disrupted adhesion molecule maturation. Under shear stress conditions, 2-deoxy-d-glucose impaired monocyte rolling and adhesion to VECs monolayers, whereas DON reduced rolling. In conclusion, glycolysis and its side-branch route of hexosamine biosynthesis are necessary for nutrient-driven posttranslational modifications of inflammatory proteins in inflamed valve cells and key steps of monocyte recruitment, a crucial process in early stages of CAVD. NEW & NOTEWORTHY The study uncovers a relevant role of glycolysis and its side-branch route of hexosamine biosynthesis by regulating sugar-driven posttranslational modifications that are critical for the inflammatory response and key steps of leukocyte adhesion to human aortic valve cells. These events are essential for the recruitment of cells of the monocytic lineage to aortic valve leaflets in the initial stages of calcific aortic valve disease.