Vascular endothelial dysfunction in β-thalassemia occurs despite increased eNOS expression and preserved vascular smooth muscle cell reactivity to NO

Abstract

Aims: The hereditary β-thalassemia major condition requires regular lifelong blood transfusions. Transfusion-related iron overloading has been associated with the onset of cardiovascular complications, including cardiac dysfunction and vascular anomalies. By using an untransfused murine model of β-thalassemia major, we tested the hypothesis that vascular endothelial dysfunction, alterations of arterial structure and of its mechanical properties would occur despite the absence of treatments.

Methods and results: Vascular function and structure were evaluated ex vivo. Compared to the controls, endothelium-dependent vasodilation with acetylcholine was blunted in mesenteric resistance arteries of β-thalassemic mice while the endothelium-independent vasodilator (sodium nitroprusside) produced comparable vessel dilation, indicating endothelial cell impairment with preserved smooth muscle cell reactivity to nitric oxide (NO). While these findings suggest a decrease in NO bioavailability, Western blotting showed heightened expression of aortic endothelial NO synthase (eNOS) in β-thalassemia. Vascular remodeling of the common carotid arteries revealed increased medial elastin content. Under isobaric conditions, the carotid arteries of β-thalassemic mice exhibited decreased wall stress and softening due to structural changes of the vessel wall.

Conclusions: A complex vasculopathy was identified in untransfused β-thalassemic mice characterized by altered carotid artery structure and endothelial dysfunction of resistance arterioles, likely attributable to reduced NO bioavailability despite enhanced vascular eNOS expression.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1. Vasodilatory responses of mesenteric resistance arterioles to acetylcholine (ACh) (A) and sodium nitroprusside (SNP) (B), in control (○; n = 10) and homo-βthal (•; n = 9) mice.

Relaxation responses are expressed as a percentage increase in lumen diameter after norepinephrine pre-contraction. Data are means ± SEM. *p<0.05.

Figure 2. Endothelium-dependent vasodilatory responses of mesenteric resistance arterioles from control (n = 10) (A) and homo-βthal mice (n = 9) (B) to acetylcholine (ACh) in the absence (○) or presence (

▴) of L-NAME. Relaxation responses are expressed as a percentage increase in lumen diameter after norepinephrine pre-contraction. Data are means ± SEM. *p<0.05 and †p<0.001.

Figure 3. eNOS protein expression in aortae of control and homo-βthal mice.

(A) Representative Western blotting, and (B) densitometric analysis of eNOS protein expression (n = 6 mice per group). Measurements are expressed as % of eNOS bands in aortae of control mice. Data are means ± SEM. *p<0.05.

Figure 4. Comparison of structural characteristics in common carotid arteries from control (○; n = 13) and homo-βthal mice (•; n = 11).

(A) Luminal diameter, (B) external diameter, (C) wall cross-sectional area (CSA), and (D) wall-to-lumen ratio versus intraluminal pressure. Data are means ± SEM. †p<0.001.

Figure 5. Comparison of mechanical parameters in common carotid arteries from control (○; n = 13) and homo-βthal mice (•; n = 11).

Circumferential strain-intraluminal pressure (A), circumferential stress-intraluminal pressure (B), and stress-strain (C) relationships. Data are means ± SEM. †p<0.001. The curve in panel C exhibits a rightward shift in the homo-βthal group, as shown by a significant decrease in β (p<0.01) and the slope of the tangential elastic strain modulus-to-stress relationship.

Figure 6. Histological analysis of elastin content in media (A) and representative microscopy images of Verhoeff van Gieson-stained sections (B) of the left common carotid arteries in control (open bars) and homo-βthal mice (closed bars, n = 8 per group). Data are means ± SEM.

*p<0.05. L, vascular lumen.