Glycocalyx-targeted therapy ameliorates age-related arterial dysfunction

Abstract

Advanced age is accompanied by arterial dysfunction, as well as a diminished glycocalyx, which may be linked to reduced high molecular weight-hyaluronan (HMW-HA) synthesis. However, the impact of glycocalyx deterioration in age-related arterial dysfunction is unknown. We sought to determine if manipulations in glycocalyx properties would alter arterial function. Tamoxifen-induced hyaluronan synthase 2 (Has2) reduction was used to decrease glycocalyx properties. Three weeks post-tamoxifen treatment, glycocalyx thickness was lower in Has2 knockout compared to wild-type mice (P<0.05). Has2 reduction induced arterial dysfunction, demonstrated by impaired endothelium-dependent dilation (EDD) and elevated aortic stiffness (P<0.05). To augment glycocalyx properties, old mice received 10 weeks of a glycocalyx-targeted therapy via Endocalyx™ (old+ECX), which contains HMW-HA and other glycocalyx components. Compared to old control mice, glycocalyx properties and EDD were augmented, and aortic stiffness decreased in old+ECX mice (P<0.05). Old+ECX mice had a more youthful aortic phenotype, demonstrated by lower collagen content and higher elastin content than old control mice (P<0.05). Functional outcomes were repeated in old mice that underwent a diet supplemented solely with HMW-HA (old+HA). Compared to old controls, glycocalyx properties and EDD were augmented, and aortic stiffness was lower in old+HA mice (P<0.05). We did not observe any differences between old+HA and old+ECX mice (P>0.05). Has2 reduction phenocopies age-related arterial dysfunction, while 10 weeks of glycocalyx-targeted therapy that restores the glycocalyx also ameliorates age-related arterial dysfunction. These findings suggest that the glycocalyx may be a viable therapeutic target to ameliorate age-related arterial dysfunction.

Keywords: Aging; Arterial function; Endothelium; Hyaluronan.

Fig. 1

Effects of Has2 conditional reduction on arterial function. Aortic gene expression of hyaluronan synthase 2 (Has2; A), estimated glycocalyx thickness (B), and dose responses to acetylcholine in the absence (solid lines) and presence of the nitric oxide synthase inhibitor, L-NAME (dashed lines), in carotid arteries (C) measured 3 weeks post-tamoxifen in Has2^+/+ and Has2−/− mice. Aortic pulse-wave velocity (PWV; D) at pre- and 3 weeks post-tamoxifen time points in Has2^+/+ and Has2^−/− mice. *P < 0.05 vs. Has2^+/+. †P < 0.05 vs. acetylcholine ‡P < 0.05 vs. pre-tamoxifen. Data are mean ± SEM

Fig. 2

Glycocalyx components and barrier properties. Aortic gene expression of hyaluronan synthesis (Has) genes (A), plasma hyaluronan concentrations (B), glycocalyx thickness (C), perfused boundary region (D), microvascular density (E), red blood cell fraction (F), and capillary blood volume (CBV) absolute (G) in young, old, and old Endocalyx™ (ECX)-treated B6D2F1 mice. Representative images are provided. Black scale bars are equal to 100 µm. *P < 0.05 vs. young. †P < 0.05 vs. old. Data are mean ± SEM

Fig. 3

Endothelium-dependent dilation. Dose responses to acetylcholine (ACh) in the absence (solid lines) and presence of the nitric oxide synthase inhibitor, L-NAME (dashed lines), in carotid arteries from young, old, and old Endocalyx™ (ECX)-treated B6D2F1 mice (A). Nitric oxide–mediated vasodilation (B). Maximal vasodilation to ACh in the absence and presence of L-NAME and the superoxide scavenger, TEMPOL, in carotid arteries (C). Aortic gene expression of Sod isoforms (D). Nitrotyrosine content in histological sections of aortas from young, old, and old + ECX mice (E). Representative images are provided. White scale bars are equal to 100 µm. *P < 0.05 vs. young. †P < 0.05 vs. old. ‡P < 0.05 vs. without L-NAME present within group. §P < 0.05 vs. ACh within group. Data are mean ± SEM

Fig. 4

Large artery stiffness and aortic structural characteristics. Aortic pulse-wave velocity (PWV; A), anesthetized heart rate (B), and systolic blood pressure (BP; C) at pre- and post-intervention time points in young and old time-controlled mice, as well as in old Endocalyx™ (ECX)-treated B6D2F1 mice. Medial cross-sectional area (CSA; D), media-to-lumen ratio (E), elastin (F), and collagen (G) expression in histological sections of aortas from young, old, and old + ECX mice. Representative images are provided. Black scale bars are equal to 100 µm. *P < 0.05 vs. young. †P < 0.05 vs. old. ‡P < 0.05 vs. pre. Data are mean ± SEM

Fig. 5

Arterial function in old, old with high molecular weight hyaluronan (HA) or Endocalyx™ (ECX) diet-treated C57BL/6 mice. Glycocalyx thickness (A). Dose responses to acetylcholine (ACh) in the absence (solid lines) and presence of the nitric oxide synthase inhibitor, L-NAME (dashed lines), in carotid arteries from old vs. old + HA (B), and old vs old + ECX (C) mice. Dose responses to sodium nitroprusside in carotid arteries (D). Aortic pulse-wave velocity (PWV; E) at post-intervention time point. *P < 0.05 vs. old. †P < 0.05 vs. ACh. Data are mean ± SEM