Endothelial TRPV4-eNOS coupling as a vital therapy target for treatment of hypertension.

BACKGROUND AND PURPOSE Reduced nitric oxide (NO) level and activity are signs of endothelial dysfunction, which is important in mediating blood pressure up-regulation. Previously, we demonstrated that transient receptor potential channel V4 (TRPV4) could form functional complex with other proteins to mediate vasodilation in the Endothelial cells (ECs). But how TRPV4 interacts with the NO pathway in larger arteries requires further exploration. EXPERIMENTAL APPROACH We used single-cell RNA-sequencing to find the CD106+ TRPV4high NOS3high ECs. The TRPV4-eNOS interaction was verified by co-immunoprecipitation and Immunofluorescence resonance energy transfer (FRET), and their binding site was found by site-directed mutagenesis. Endothelium-specific TRPV4 knockout (TRPV4EC-/- ) mice were used to study the effect of the TRPV4-eNOS interaction on blood pressure. A small molecule, JNc-463 was designed through molecular docking technology. KEY RESULTS We uncovered CD106+ TRPV4high NOS3high ECs in the mouse aorta, which they could regulate vasodilation via a TRPV4-eNOS interaction, and they were essential to regulate blood pressure. The TRPV4-eNOS interaction markedly decreased during the process of hypertension. We further attempted to identify the molecules re-join the TRPV4-eNOS interaction and develop a small-molecule drug, JNc-463, which could increase the TRPV4-eNOS interaction to enhance vasodilation, and exert antihypertensive effects in mice. CONCLUSION AND IMPLICATIONS This is the first study integrating single-cell RNA-Seq, single-cell functional study and drug screening in aorta. We identified a subpopulation of CD106+ TRPV4high NOS3high ECs, in which an impaired TRPV4-eNOS interaction was important in the progress of hypertension and we designed a small molecule, JNc-463 to improve the impaired TRPV4-eNOS interaction in hypertension.