Chronic senolytic treatment alleviates established vasomotor dysfunction in aged or atherosclerotic mice

Abstract

While reports suggest a single dose of senolytics may improve vasomotor function, the structural and functional impact of long-term senolytic treatment is unknown. To determine whether long-term senolytic treatment improves vasomotor function, vascular stiffness, and intimal plaque size and composition in aged or hypercholesterolemic mice with established disease. Senolytic treatment (intermittent treatment with Dasatinib + Quercetin via oral gavage) resulted in significant reductions in senescent cell markers (TAF(+) cells) in the medial layer of aorta from aged and hypercholesterolemic mice, but not in intimal atherosclerotic plaques. While senolytic treatment significantly improved vasomotor function (isolated organ chamber baths) in both groups of mice, this was due to increases in nitric oxide bioavailability in aged mice and increases in sensitivity to NO donors in hypercholesterolemic mice. Genetic clearance of senescent cells in aged normocholesterolemic INK-ATTAC mice phenocopied changes elicited by D+Q. Senolytics tended to reduce aortic calcification (alizarin red) and osteogenic signaling (qRT-PCR, immunohistochemistry) in aged mice, but both were significantly reduced by senolytic treatment in hypercholesterolemic mice. Intimal plaque fibrosis (picrosirius red) was not changed appreciably by chronic senolytic treatment. This is the first study to demonstrate that chronic clearance of senescent cells improves established vascular phenotypes associated with aging and chronic hypercholesterolemia, and may be a viable therapeutic intervention to reduce morbidity and mortality from cardiovascular diseases.

Keywords: aging; atherosclerosis; calcification; endothelial function; fibrosis; senescence.

Figure 1

Effects of senolytic treatment on senescent cell burden, DNA damage, vasomotor function, nitric oxide signaling, calcification, and osteogenic signaling in chronologically aged mice. Chronic treatment with Dasatinib + Quercetin reduced senescent cell burden (A–B) and DNA damage (C) in aorta (Panel A pseudocolor legend for high magnification lower panels: DAPI/nuclei (blue), γH2A.X (green), telomeres (red). Low magnification micrographs in upper images are provided as an anatomic frame of reference/origin). White squares mark regions that are magnified in subsequent micrographs, and telomere‐associated foci are numbered in the bottom panels. Chronic senescent cell clearance improved vascular relaxation to acetylcholine (D) independently from changes in sodium nitroprusside (E) and significantly increased p‐VASP ²³⁹ levels (F). Chronic senolytic treatment with Dasatinib + Quercetin also tended to reduce vascular calcification (G), which was associated with modest reductions in osterix immunofluorescence (H). In all panels, asterisk denotes p < 0.05.

Figure 2

Effects of senolytic treatment on senescent cell burden, DNA damage, vasomotor function, and nitric oxide signaling in hypercholesterolemic mice. Chronic treatment with Dasatinib + Quercetin reduced senescent cell burden (A–B) and DNA damage (C) in the media of atherosclerotic plaques but not the intima (Panel A pseudocolor legend for high magnification lower panels: DAPI/nuclei (blue), γH2A.X (green), telomeres (red). Low magnification micrographs in upper images are provided as an anatomic frame of reference/origin). Chronic senescent cell clearance modestly improved vascular relaxation to acetylcholine (D) but markedly improved relaxation to sodium nitroprusside (E), and significantly increased levels of p‐eNOS ^ser1177 (F) and p‐VASP ²³⁹ (G) in both the media and intima of atherosclerotic vessels. While chronic intermittent senolytic treatment with Dasatinib + Quercetin did not alter lipid composition of plaques (H), senolytic treatment did significantly reduce vascular calcification (I), which was associated with marked reductions in osterix immunofluorescence (J). In all panels, asterisk denotes p < 0.05.