Abstract P086: Endothelial Cullin3 Mutation Causes Vascular Dysfunction, Arterial Stiffening, And Hypertension

Abstract

Mutations in CULLIN3 gene (causing in-frame deletion of exon 9, termed Cul3Δ9) cause human hypertension (HT), which is driven by a combination of renal tubular and vascular mechanisms. We have previously shown that disruption of Cullin3 (CUL3) in vascular smooth muscle impairs nitric oxide (NO) signaling and vasodilation through decreased cGMP bioavailability, strongly supporting a role of vascular CUL3 in blood pressure (BP) regulation. To test the importance of endothelial Cul3 in vivo , we bred the conditionally activatable Cul3Δ9 mice with tamoxifen-inducible Tie2-CRE ERT2 mice. Four weeks after tamoxifen, the resultant mice (E-Cul3Δ9) developed nocturnal HT (Night time peak systolic BP, E-Cul3Δ9: 135±3 vs Control: 124±3 mmHg) and arterial stiffening (pulse wave velocity, 3.7±0.3 vs 2.7±0.1 m/s). No difference was seen in daytime BP. To determine whether vascular remodeling impairs baroreflex function, we performed power spectral analysis. Heart rate (HR), low frequency/high frequency ratio of HR variability, and baroreflex gain were comparable between control and E-Cul3Δ9 mice, suggesting there was no change in cardiac sympathetic nerve activity. However, low frequency amplitude of arterial pressure variability (16±4 vs 7±2 mmHg 2 ) at night was markedly augmented in E-Cul3Δ9 mice, suggesting increased sympathetic activity in vascular tone regulation. Consistently, E-Cul3Δ9 mice exhibited impaired endothelial-dependent relaxation in carotid artery (Max ACh relaxation: 69% vs 84%) and cerebral resistance basilar artery (41% vs 77%). No difference in smooth muscle function was observed. Expression of Cul3Δ9 in primary mouse aortic endothelial cells markedly decreased wild type Cul3 protein, phosphorylated eNOS, and NO production. Because protein phosphatase 2A (PP2A) is a known Cul3 substrate which dephosphorylates eNOS, we determined whether impaired eNOS activity was attributable to PP2A. Cul3Δ9-induced impairment of eNOS activity was rescued by a selective PP2A inhibitor Okadaic Acid (4 nM), but not by a Protein Phosphatase 1 inhibitor Tautomycetin (4 nM). Thus, CUL3 mutations in the endothelium contribute to human HT in part through decreased NO bioavailability, endothelial dysfunction and secondary sympathoexcitation.