Intracellular NAD⁺ Depletion Increases Prostanoid Production via p38/COX2 Signalling in FK866-Induced Senescent Human...

Vascular endothelial cells maintain vascular homeostasis by releasing mediators with vasoconstrictive and vasorelaxant effects. Prostanoids are bioactive substances synthesised by cyclooxygenase2 (COX2); they preserve vascular function and can increase the risk of cardiovascular disease (CVD) associated with ageing. Age-related reductions in nicotinamide adenine dinucleotide (NAD⁺) have been implicated in CVD pathogenesis. However, the relationship between intracellular NAD⁺ levels and prostanoid production in vascular endothelial cells is unclear. Herein, reduced intracellular NAD⁺ levels upregulated COX2 protein expression by activating the p38 mitogen-activated protein kinase (MAPK) signalling pathway, increasing the production of prostaglandin F_1α (PGF_1α) and thromboxane B₂ (TXB₂) in human umbilical vein endothelial cells (HUVECs). Treatment with the nicotinamide phosphoribosyltransferase (NAMPT) inhibitor, FK866, decreased intracellular NAD⁺ levels, induced a cellular senescence-like phenotype characterised by the suppression of cell proliferation without cell death and promoted PGF_1α and TXB₂ production in HUVECs. FK866 treatment increased phosphorylated p38 and COX2 protein levels, whereas treatment with the p38 inhibitor, PD169316, suppressed the FK866-induced increase in COX2 expression. Supplementation with nicotinamide mononucleotide (NMN), a precursor of NAD⁺, following FK866 treatment restored intracellular NAD⁺ levels, reduced the cellular senescence-like phenotype and attenuated the production of PGF_1α and TXB₂. These results suggest that intracellular NAD⁺ appears to regulate prostanoid production in HUVECs under conditions of acute depletion.