Role of NADPH oxidases in TRPV4‐mediated vasodilation of human arterioles
Authors: Xie Yangjing , Nishijima Yoshinori , Zhang David 
Published: June 2021

NADPH oxidases (NOXs) are a major source of reactive oxygen species (ROS) in the vascular system and play diverse roles in cardiovascular health and disease. We recently reported that H2O2, a diffusible ROS, regulates protein phosphorylation of endothelial transient receptor potential vanilloid 4 (TRPV4) channels and TRPV4‐mediated vasodilation in human coronary arterioles (HCA) from subjects with coronary artery disease (CAD). The endogenous source of H2O2 involved in TRPV4 regulation remains unclear. In this study, we investigated the expression of different NOX isoforms in human arterioles and their potential role in the regulation of TRPV4‐mediated arteriolar dilation.

Human adipose arterioles (HAA) and HCA were dissected from fresh adipose and heart tissues obtained as discarded surgical specimens. The mRNA and protein expression of NOXs were detected by RT‐PCR and Western blotting. TRPV4 agonist‐induced vasodilation was examined in cannulated arterioles (100–200 μm internal diameter at 60 mmHg) by measuring diameter changes using video microscopy.

RT‐PCR analysis revealed consistent expression of NOX2 and 4 but low expression of other isoforms (NOX1, 3, and 5) in HAA and HCA. Using endothelium‐intact and denuded arterioles (n=3 tissues/each) as well as freshly isolated endothelial cells and smooth muscle cells (n=2 tissues/each), we further found that NOX2 and 4 are more abundantly expressed in endothelial cells and at a significantly higher level than other NOX isoforms. Consistent with mRNA expression, NOX2 and 4 proteins were detected in HAA and HCA using Western blot analysis. In cannulated arterioles, TRPV4 agonist GSK1016790A (10−9–10−7 M) induced potent and concentration‐dependent dilation in HAA (maximal dilation at 10−7 M, 76±6%, logEC50 −8.0±0.1, n=6) and HCA (maximal dilation, 69±8%, logEC50 −7.9±0.1, n=5). This dilation was significantly reduced by the small molecule NOX2 inhibitor GSK2795039 (1 μM) in both HAA (22±8%, n=6, p< 0.05 vs control) and HCA (22±10%, n=5, p<0.05 vs control). Interestingly, NOX4 inhibitor GKT137831 (1 μM) also significantly reduced the dilation in HAA (maximal dilation at 10−7 M, 18±8% vs. 72±8% in control, n=6, p<0.05). Both NOX2 and 4 inhibitors did not affect sodium nitroprusside (10−10–10−4 M, an endothelium‐independent vasodilator)‐induced dilation in HAA (maximal dilation at 10−4 M, 94±2% with GSK2795039 vs. 94±2% of control, and 96±3% with GKT137831 vs. 97±3% of control, n=4 and 3, respectively).

These results indicate that NOX2 and NOX4 represent two main isoforms of NOXs expressed in endothelial cells of human arterioles and play a potentially important role in TRPV4‐mediated endothelium‐dependent vasodilation. Future studies will explore whether the mechanisms of TRPV4 regulation involves NOX2/4‐derived and H2O2‐mediated channel protein phosphorylation or other signaling pathways in endothelial cells.

Support or Funding Information

National Heart, Lung and Blood Institute Grant RO1‐HL 096647

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.