(849.17) Rho-related BTB Domain-containing Protein 1 Reverses Established Arterial Stiffness in Angiotensin II-treated Mice

Poster Board Number: E17

Shi Fang (The Medical College of Wisconsin, University of Iowa), Jing Wu (The Medical College of Wisconsin), John Reho (The Medical College of Wisconsin), Ko-ting Lu (The Medical College of Wisconsin), Daniel Brozoski (The Medical College of Wisconsin), Alec Werthman (The Medical College of Wisconsin), Gaurav Kumar (The Medical College of Wisconsin), Sebastiao Silva (The Medical College of Wisconsin), Patricia Muskus (The Medical College of Wisconsin), Kelsey Wackman (The Medical College of Wisconsin), Angela Mathison (The Medical College of Wisconsin, Medical College of Wisconsin), Raul Urrutia (The Medical College of Wisconsin, Medical College of Wisconsin), Bi Qing Teng (The Medical College of Wisconsin), Chien-Wei Lin (The Medical College of Wisconsin), Fredrick Quelle (University of Iowa), Curt Sigmund (The Medical College of Wisconsin)

Rho-related BTB domain-containing protein 1 (RhoBTB1) is associated with blood pressure in GWAS studies, but its function is not well-understood. The level of RhoBTB1 mRNA was decreased in mice treated with angiotensin II (Ang-II) concomitant with hypertension, arterial stiffness, and impaired vasodilation. Here, we tested the hypothesis that restoration of RhoBTB1 would reverse hypertension, arterial stiffness, and improve vasodilation response caused by Ang-II infusion. Genetic complementation of RhoBTB1 in the vasculature was achieved by expressing a smooth muscle-specific, tamoxifen-inducible RhoBTB1 transgene (S-RhoBTB1 mice). Mice expressing smooth muscle-specific, tamoxifen-inducible Cre-recombinase without RhoBTB1 (ISM-Cre mice) were used as genotype control. ISM-Cre and S-RhoBTB1 mice received 6 weeks of Ang-II infusion, but their transgene was not activated until two weeks after the initiation of Ang-II treatment. Meanwhile, vehicle-treated ISM-Cre and S-RhoBTB1 mice were used as control. Blood pressure was continuously recorded by radiotelemetry during the entire protocol. Pulse wave velocity (PWV) and aortic compliance assay were used to evaluate arterial stiffness both during and at the end of the protocol. Vasodilation was measured in isolated carotid and mesenteric arteries using wire myography.  In vehicle-treated mice, aortic RhoBTB1 expression was increased in S-RhoBTB1 mice compared to ISM-Cre mice. Ang-II significantly decreased the level of RhoBTB1 in the aorta of ISM-Cre mice whereas S-RhoBTB1 mice normalized this effect. At baseline, overexpressing RhoBTB1 in the vasculature had no significant effect on any phenotype. Before transgene activation, Ang-II increased blood pressure, arterial stiffness, and impaired vasodilation in response to acetylcholine and sodium nitroprusside in both ISM-Cre and S-RhoBTB1 mice. Remarkably, in Ang-II-treated mice, RhoBTB1 restoration restored arterial stiffness nearly back to baseline without rescuing either hypertension or vasodilation. Since arterial stiffness is often observed to go hand-in-hand with hypertension, the current phenotype is particularly interesting as it provides novel insight into the cause-effect relationship between arterial stiffness and hypertension. Next, we utilized RNA-seq and biochemical assays to investigate the mechanism by which RhoBTB1 reversed arterial stiffness.  Classical contributors of arterial stiffness, for instance, collagen, elastin, and vascular smooth muscle remodeling, were not altered by RhoBTB1 restoration. However, actin polymerization was elevated by Ang-II and normalized upon RhoBTB1 restoration, as evident by both immunoblotting and fluorescent labeling. In conclusion, we identified a novel function of RhoBTB1 and showed its importance in protecting from and reversing arterial stiffness.

lt;agt;This study was funded by the National Institutes of Health (HL144807 to CDS, DK126792 to JW) and the American Heart Association (20PRE35120137 to SF).lt;/agt;