Aging and Endothelial Dysfunction: The Role of NADPH Oxidase and the Insufficient Inhibition by PKG

Abstract

BackgroundNicotinamide adenine dinucleotide phosphate (NADPH) oxidase‐induced oxidative stress plays a critical role in the age‐related impairment of endothelium‐dependent dilation. However, the role of NADPH oxidase (i.e. which nitrogen oxides (NOX) may be involved) and the potential role of protein kinase G (PKG) inhibition of NADPH in human skeletal muscle feed arteries (SMFAs) has yet to be clarified. Therefore, this study sought to determine, in vitro, if treatment with NOX1 inhibitor (ML090), NOX2 inhibitor (gp91ds‐tat), and NOX4 inhibitor (plumbagin) could restore the age‐related endothelial dysfunction in human SMFAs.MethodsPKG, NOX1, NOX2, and NOX4 protein expression and cyclic guanosine monophosphate (cGMP) activity were analyzed in young (24±6 yrs, n=7) and old (77±8 yrs, n=8) subjects via Western blot analysis and an enzyme‐linked immunosorbent assay (ELISA). Using pressure myography, endothelium‐dependent and ‐independent vasodilation were assessed with and without ML090, gp91ds‐tat, and plumbagin in SMFAs obtained from 13 young and 16 old subjects in response to three stimuli 1) flow‐induced shear stress, 2) acetylcholine (ACh), and 3) sodium nitroprusside (SNP). Additionally, L‐NG‐monomethyl arginine citrate (L‐NMMA) was utilized to identify the impact of blocking nitric oxide (NO) synthase on these responses.ResultsNOX2 and NOX4, but not NOX1, protein expression were significantly increased in the old compared to the young SMFAs. In addition, cGMP activity and PKG protein expression were significantly lower in the old compared to the young SMFAs. Endothelial‐dependent vasodilatory dysfunction was evident in response to intraluminal flow (Young: 81 ± 15; Old: 27 ± 14%, P < 0.05) and ACh (Young: 95 ± 17, Old: 34 ± 15 %, P < 0.05). The NOX2 inhibitor, gp91ds‐tat, restored the vasodilation response in the old SMFAs to both flow and Ach (70 ± 11 and 89 ± 13 %, respectively, P < 0.05), while the NOX4 inhibitor, plumbagin, partially restored the vasodilation response in the old SMFAs to flow and ACh (45 ± 15 %, P=0.08; 59 ± 13 %, P=0.07). However, the NOX1 inhibitor, ML090, had no effect. Endothelium‐independent vasodilation (SNP) was not affected by age and, therefore, all NOX inhibitors had no effect on SNP‐induced vasodilation. L‐NMMA negated the restorative effects of NOX2 and NOX4 blockade.ConclusionsBoth NOX2 and to a lesser extent NOX4 appear to play an important role in the age‐associated endothelial dysfunction exhibited by SMFAs. Furthermore, in combination, these findings suggest that there is likely an age‐related weakening of the NO‐initiated link between cGMP, PKG, and NADPH oxidase, with PKG demonstrating insufficient NADPH oxidase inhibition with advancing age and, ultimately, attenuated vascular function.Support or Funding Informationthe National Heart, Lung, and Blood Institute at the National Institute of Health (PO1 HL1091830) and the Veteran's Administration Rehabilitation Research and Development Service (E6910‐R, E1697‐R, E1433‐P, E9275‐L and E1572‐P).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.