Circulating factors from old mice directly induce endothelial dysfunction and aortic stiffening in young mouse arteries: A novel ex vivo experimental approach

Abstract

Aging is the primary risk factor for the development of cardiovascular diseases, largely as a result of reduced arterial function, characterized by endothelial arterial function, characterized by endothelial dysfunction and large elastic artery (e.g., aorta) stiffening. Changes in circulating factors (assessed in serum or plasma) may contribute to arterial dysfunction with aging; however, this has not yet been directly investigated. Heterochronic parabiosis could be used to study the influence of circulating factors from old animals on young arteries (and vice versa) but is limited by potential confounding effects of other organ systems. Thus, a highly controlled experimental model is necessary to determine the direct effects of changes in circulating factors with aging on arterial function. PURPOSE: To develop an experimental model that allows for direct assessment of circulating factors in mediating endothelial dysfunction and arterial stiffening with aging. METHODS & RESULTS: Endothelial Function. Common carotid arteries fromyoung (3-4mo) male C57BL/6 mice were cannulated in a tissue culture pressure myograph and incubated 24h with modified Krebs buffer containing 5% serum from either young (YS; 5mo, N=4) or old (OS; 24mo, N=4) male donor mice. Endothelial function was measured as endothelium-dependent dilation (EDD) to increasing doses of acetylcholine (10 to 10 M). Endothelium independent dilation (EID) was assessed as dilation to the nitric oxide (NO) donor sodium nitroprusside (10 to 10 M).Peak EDD was lower in arteries incubated with OS compared with YS (OS: 75.3±2.4%; YS: 89.4±1.3%; p=0.008). There were no differences in EID between treatments (OS: 91.7±2.5%; YS: 97.4±1.5%; p=0.06), suggesting circulating factors from old donors induced dysfunction in an endothelium-specific manner and were not attributable to changes in smooth muscle sensitivity to NO. Arterial Stiffness. Aortic rings from young (6mo) male mice were incubated 48h in standard culture media with 5% fetal bovine serum (control; N=8) or plasma from either young (YP; 6mo, N=4) or old (OP; 27mo, N=8) male donor mice and then tested for collagen elastic modulus (EM; an ex vivo measure of intrinsic mechanical wall stiffness) on an organ bath pin myograph.Collagen EM was higher (fold-difference from control) in aortic rings incubated with OP but no different in those incubated with YP (OP: 1.41±0.13, p=0.043; YP: 1.04±0.17, p=0.97), indicating higher intrinsic wall stiffness after exposure to plasma from old donors. Moreover, plasma-induced changes in aortic collagen EM were positively associated with aortic pulse wave velocity (an in vivo measure of aortic stiffness) of donor mice (r =0.34, p=0.047), suggesting that aortic stiffening with aging is in part due to changes in circulating factors. CONCLUSIONS: These data serve as a successful demonstration of a novel ex vivo model for determining the role of circulating factors in mediating endothelial dysfunction and arterial stiffening with aging. R21 AG078408, K99 HL159241, K99/R00 HL151818. VEB & ZSC contributed equally.