Conditional deletion of smooth muscle Cullin-3 causes severe progressive hypertension
Larry N. Agbor, Anand R. Nair, Jing Wu, Ko-Ting Lu, Deborah R. Davis, Henry L. Keen, Frederick W. Quelle, James A. McCormick, Jeffrey D. Singer, Curt D. Sigmund
Larry N. Agbor, Anand R. Nair, Jing Wu, Ko-Ting Lu, Deborah R. Davis, Henry L. Keen, Frederick W. Quelle, James A. McCormick, Jeffrey D. Singer, Curt D. Sigmund
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Research Article Vascular biology

Conditional deletion of smooth muscle Cullin-3 causes severe progressive hypertension

Abstract

Patients with mutations in Cullin-3 (CUL3) exhibit severe early-onset hypertension, but the contribution of the smooth muscle remains unclear. Conditional genetic ablation of CUL3 in vascular smooth muscle (S-CUL3KO) causes progressive impairment in response to NO, rapid development of severe hypertension, and increased arterial stiffness. Loss of CUL3 in primary aortic smooth muscle cells or aorta resulted in decreased expression of the NO receptor soluble guanylate cyclase (sGC), a marked reduction in cGMP production, and impaired vasodilation to cGMP analogs. Vasodilation responses to a selective large-conductance Ca2+-activated K+ channel activator were normal, suggesting that downstream signals that promote smooth muscle–dependent relaxation remained intact. We conclude that smooth muscle–specific CUL3 ablation impairs both cGMP production and cGMP responses and that loss of CUL3 function selectively in smooth muscle is sufficient to cause severe hypertension by interfering with the NO/sGC/cGMP pathway. Our study provides evidence that vascular smooth muscle CUL3 is a major regulator of BP. CUL3 mutations cause severe vascular dysfunction, arterial stiffness, and hypertension due to defects in vascular smooth muscle.

Authors

Larry N. Agbor, Anand R. Nair, Jing Wu, Ko-Ting Lu, Deborah R. Davis, Henry L. Keen, Frederick W. Quelle, James A. McCormick, Jeffrey D. Singer, Curt D. Sigmund

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Figure 4

Vasocontraction of aorta: role of RhoA/ROCK.

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Vasocontraction of aorta: role of RhoA/ROCK. (A–D) Vasocontraction of th...
(A–D) Vasocontraction of thoracic aorta from the indicated mice (Con, control mice). Aorta rings were isolated and cleaned of perivascular adipose fat and were equilibrated for 45 minutes at 0.5 g tension; and vasocontraction responses to potassium chloride at 10 mmol/L (A, n = 5–7/genotype), prostaglandin F2α (PGF2α; B, n = 5–7/genotype), and endothelin 1 (D, ET-1, n = 4 per genotype) were assessed. For A–D, error bars represent mean ± SEM. *P < 0.05 by 1-way (A and C) or 2-way repeated-measures ANOVA (B and D). Note that the S-CUL3het graph in B was shifted 0.05 units in the x and y directions to clearly indicate the presence of 3 data sets on the graph because the control and S-CUL3 were essentially superimposed. Individual points for the 1 μM dose are shown in C. (E) Western blot showing expression levels of indicated proteins in aorta from control and S-CUL3KO mice. Aortas were cleaned of perivascular fat and adventitia. Representative of 3 independent experiments. MYPT, myosin light chain phosphatase. (F and G) Quantification of total RhoA (F) and P-MYPT1 (G) in aorta. For F and G, *P < 0.05, S-CUL3KO vs. control by Student’s t test. (H) ET-1–induced vasoconstriction at 10 μmol/L in the absence or presence of the Rho kinase inhibitor Y27632 (n = 6–8/genotype). For F and G, error bars represent mean ± SEM. *P < 0.05 by 1-way ANOVA. For H, *P < 0.05 vs. control; #P < 0.05 vs. baseline by one-way ANOVA. (I and J) Effect of the Rho kinase inhibitor Y27632 (1 μmol/L, 30 minutes) on acetylcholine-mediated (Ach) relaxation in aorta (I, n = 4–6/genotype) and basilar arteries (J, n = 5–7/genotype). For I and J, error bars represent mean ± SEM. *P < 0.05 by 2-way repeated-measures ANOVA.