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

Background and purpose: Reduced NO levels and activity are signs of endothelial dysfunction, which is important in mediating BP changes. Previously, we demonstrated that transient receptor potential channel V4 (TRPV4) could form a functional complex with Other proteins to mediate vasodilation in 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⁺ TRPV4^high NOS3^high ECs. The TRPV4-eNOS interaction was verified by co-immunoprecipitation and immuno-FRET, and their binding site was found by site-directed mutagenesis. Endothelium-specific TRPV4 knockout (TRPV4_EC^-/- ) mice were used to study the effect of the TRPV4-eNOS interaction on BP. A small molecule, JNc-463, was designed through molecular docking technology.

Key results: We uncovered CD106⁺ TRPV4^high NOS3^high ECs in the mouse aorta, which could regulate vasodilation via a TRPV4-eNOS interaction, and were essential to regulate BP. The TRPV4-eNOS interaction markedly decreased during the process of hypertension. We further attempted to identify molecules involved in the TRPV4-eNOS interaction and developed 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⁺ TRPV4^high NOS3^high 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.