Obesity-induced vascular dysfunction and arterial stiffening requires endothelial cell arginase 1

Abstract

Aims: Elevation of arginase activity has been linked to vascular dysfunction in diabetes and hypertension by a mechanism involving decreased nitric oxide (NO) bioavailability due to L-arginine depletion. Excessive arginase activity also can drive L-arginine metabolism towards the production of ornithine, polyamines, and proline, promoting proliferation of vascular smooth muscle cells and collagen formation, leading to perivascular fibrosis. We hypothesized that there is a specific involvement of arginase 1 expression within the vascular endothelial cells in this pathology.

Methods and results: To test this proposition, we used models of type 2 diabetes and metabolic syndrome. Studies were performed using wild type (WT), endothelial-specific arginase 1 knockout (EC-A1-/-) and littermate controls(A1con) mice fed high fat-high sucrose (HFHS) or normal diet (ND) for 6 months and isolated vessels exposed to palmitate-high glucose (PA/HG) media. Some WT mice or isolated vessels were treated with an arginase inhibitor, ABH [2-(S)-amino-6-boronohexanoic acid. In WT mice, the HFHS diet promoted increases in body weight, fasting blood glucose, and post-prandial insulin levels along with arterial stiffening and fibrosis, elevated blood pressure, decreased plasma levels of L-arginine, and elevated L-ornithine. The HFHS diet or PA/HG treatment also induced increases in vascular arginase activity along with oxidative stress, reduced vascular NO levels, and impaired endothelial-dependent vasorelaxation. All of these effects except obesity and hypercholesterolemia were prevented or significantly reduced by endothelial-specific deletion of arginase 1 or ABH treatment.

Conclusion: Vascular dysfunctions in diet-induced obesity are prevented by deletion of arginase 1 in vascular endothelial cells or arginase inhibition. These findings indicate that upregulation of arginase 1 expression/activity in vascular endothelial cells has an integral role in diet-induced cardiovascular dysfunction and metabolic syndrome.

Keywords: Arginase; Diabetes; Fibrosis; Obesity; Vascular dysfunction.

Figure 1

EC arginase 1 deletion or arginase inhibition prevents HFHS-induced decreases in endothelium-dependent vasorelaxation to acetylcholine (ACh). (A) Effects of EC arginase 1 deletion or ABH (C) on endothelium-dependent vasorelaxation. Effects on endothelium-independent vasorelaxation to sodium nitroprusside (SNP, B, D). Values are mean ± SEM, n = 6–8. *P < 0.05 vs. WT ND or A1^con ND groups.

Figure 2

EC arginase 1 deletion prevents palmitic acid and high glucose (PA/HG)-induced decreases in endothelium-dependent vasorelaxation to acetylcholine (ACh). Effects of EC arginase 1 deletion on endothelium-dependent vasorelaxation in aorta (A) or mesenteric arteries (C) exposed to PA/HG or normal glucose control media (NG). Effects on endothelium-independent vasorelaxation to sodium nitroprusside (SNP, B, D). Values are mean ± SEM, n = 5. *P < 0.05, vs. NG.

Figure 3

EC arginase 1 deletion or arginase inhibition prevents HFHS-induced aortic stiffening and fibrosis. Effects of arginase 1 deletion or ABH on pulse wave velocity (A) and collagen deposition (B, C). Representative sections stained with Picrosirius Red (C). Values are mean ± SEM, n = 5–6. *P < 0.05 vs. ND.

Figure 4

EC arginase 1 deletion or arginase inhibition prevents HFHS-induced increases in arginase activity and decreases in L-arginine bioavailability. Effects of arginase 1 deletion or ABH on aortic (A) and plasma arginase activity (B). Effects of ABH on plasma L-arginine (C), the ratio of plasma L-ornithine to L-arginine (D) (a measure of systemic arginase activity) and ratio of plasma L-arginine to [L-ornithine + L-citrulline] (a measure of L-arginine bioavailability (E). Values are mean ± SEM, n = 5–7. *P < 0.05 vs. ND.

Figure 5

EC arginase 1 deletion or arginase inhibition prevents HFHS-induced increases in arginase 1 expression in the aorta. Effects of EC arginase 1 deletion (A) and ABH (B) on arginase 1 (A1) protein or arginase 1 (A1) and arginase 2 (A2) mRNA (C, D). Values are mean ± SEM, n = 4–6. *P < 0.05.

Figure 6

EC arginase 1 deletion or arginase inhibition prevents HFHS or PA/HG -induced decreases in vascular nitric oxide. Effects of EC arginase 1 deletion or ABH on fluorescence for the NO indicator DAF2-DA (A). Representative images of DAF2-DA fluorescence (B). Values are mean ± SEM, n = 5–6. *P < 0.05 vs. ND. Effects of EC arginase 1 deletion on NO production in aorta exposed to PA/HG ex vivo (C). Values are mean ± SEM, n = 5. *P < 0.05 vs. control media (NG).

Figure 7

EC arginase 1 deletion or arginase inhibition prevents HFHS-induced increases in ROS levels. (A) Effects of arginase 1 deletion or ABH on plasma malonaldehyde (MDA) and (B) aortic fluorescence for the ROS indicator dihydroethidium (DHE). (C) Representative images of DHE fluorescence. Values are mean ± SEM, n = 5–6. *P< 0.05 vs. ND. (D) Levels of the peroxynitrite marker—3-nitrotyrosine in aorta from A1con and ECA1^−/− mice exposed to normal glucose control media (CM) or palmitic acid and high glucose (PA/HG) for 24 h. Values are mean ± SEM, n = 5. *P< 0.05 vs. NG.