Sitting leg vasculopathy: potential adaptations beyond the endothelium

Abstract

Increased sitting time, the most common form of sedentary behavior, is an independent risk factor for all-cause and cardiovascular disease mortality; however, the mechanisms linking sitting to cardiovascular risk remain largely elusive. Studies over the last decade have led to the concept that excessive time spent in the sitting position and the ensuing reduction in leg blood flow-induced shear stress cause endothelial dysfunction. This conclusion has been mainly supported by studies using flow-mediated dilation in the lower extremities as the measured outcome. In this review, we summarize evidence from classic studies and more recent ones that collectively support the notion that prolonged sitting-induced leg vascular dysfunction is likely also attributable to changes occurring in vascular smooth muscle cells (VSMCs). Indeed, we provide evidence that prolonged constriction of resistance arteries can lead to modifications in the structural characteristics of the vascular wall, including polymerization of actin filaments in VSMCs and inward remodeling, and that these changes manifest in a time frame that is consistent with the vascular changes observed with prolonged sitting. We expect this review will stimulate future studies with a focus on VSMC cytoskeletal remodeling as a potential target to prevent the detrimental vascular ramifications of too much sitting.

Keywords: cytoskeletal remodeling; endothelium-independent vasodilation; inward remodeling leg vascular dysfunction; prolonged sitting; vascular smooth muscle.

Figure 1.

Prolonged sitting is associated with pronounced reduction in leg blood flow (A), impaired reactive hyperemia (B), and blunted flow-mediated dilation (FMD; C) in the popliteal artery in healthy young adults. Values are presented as means ± SE. *P < 0.05 vs. presitting. Figure redrawn from Restaino et al. (21) with permission.

Figure 2.

Evidence that sitting for 3-h blunts passive limb movement (PLM)-induced hyperemia in the leg of young healthy aerobically untrained adults. Values are presented as means ± SE. *P < 0.05 vs. presitting. Figure redrawn from Garten et al. (27) with permission.

Figure 3.

Nitroglycerin-mediated dilation (NMD), an assessment of endothelium-independent vasodilation, is impaired in the popliteal artery after 3 h of sitting in healthy young adults. Values are presented as means ± SE. *P < 0.05 vs. presitting. Figure redrawn from Liu et al. (30) with permission.

Figure 4.

Prolonged exposure to norepinephrine and angiotensin II (NE + ANG II) causes inward eutrophic remodeling in rat (male Sprague–Dawley; weight range, 250–350 g)-isolated cremaster arterioles. Luminal diameter of arterioles exposed to NE (10^−5.5 M) + ANG II (10⁻⁷ M) and their maximal relaxation responses under Ca²⁺-free conditions before and after exposure to NE + ANG II for 4 h. Mean trace is illustrated. *P < 0.05 vs. before agonist exposure. Figure redrawn from Martinez-Lemus et al. (104) with permission.

Figure 5.

Disruption of the actin cytoskeleton reverts the inward remodeling caused by prolonged exposure to norepinephrine (NE) and angiotensin-II (ANG II) in rat (male Sprague–Dawley; weight range, 250–350 g)-isolated cremaster arterioles. Before and after the 4-h incubation with NE (10^−5.5 M) + ANG II (10⁻⁷ M), arterioles were allowed to develop spontaneous myogenic tone and were exposed to Ca²⁺-free conditions to assess maximal relaxation. After 5 min under the second exposure to Ca⁺²-free conditions, arterioles were treated (1 h) with vehicle control or mycalolide-B (2 µM) to depolymerize actin fibers. Mean traces are illustrated. *P < 0.05 vs. control. Figure redrawn from Staiculescu et al. (111) with permission.

Figure 6.

Prolonged exposure to norepinephrine and angiotensin-II [NE (10^−5.5 M) + ANG II (10⁻⁷ M), 4 h] increases reactive oxygen species (ROS) formation and matrix metalloproteinases (MMPs) activation in rat (male Sprague–Dawley; weight range, 250–350 g)-isolated cremaster arterioles. A: arterioles with or without NE + ANG II were coincubated with 5- (and 6-)carboxy-2′,7′-dichlorodihydrofluorescein diacetate (DCFH; 30 µM) and dihydroethidium (DHE; 5 µM) and imaged with a multiphoton microscope. Bar graphs represent percent changes in DCFH and DHE fluorescence intensities from 5 min to 4 h of incubation. B: arterioles with or without NE + ANG II were subjected to gel zymography. Bands representing gelatinolytic activity were analyzed by densitometry and expressed as fold changes from control for the activity of latent (72 kDa) and active (64 kDa) forms of MMP2. Values are presented as means ± SE. *P < 0.05 vs. control. Figure redrawn from Martinez-Lemus et al. (110) with permission.

Figure 7.

Actin depolymerization reverts tissue transglutaminase (TG2) activation-induced inward remodeling in rat-isolated cremaster arterioles. A: confocal images of isolated arterioles incubated with Alexa Fluor 488-cadaverine (green) and exposed for 4 h to vehicle control (left), to 200 µM of TG2 activator [dithiothreitol (DTT), middle], or TG2 activator in the presence of 1 mM of TG2 inhibitor cystamine (right). B: passive pressure diameter curves of arterioles before and after exposure to 200 µM of TG2 activator and cystamine (1 mM) for 4 h. *P < 0.05 vs. before TG2 activator; #P < 0.05 vs. TG2 activator. C: confocal images of isolated arterioles exposed to vehicle control (left) or 2 µM mycalolide-B (right) and subsequently stained with phalloidin-Alexa 546 to visualize the actin cytoskeleton. D: pressure-diameter curves of TG2 activator inwardly remodeled arterioles before and after exposure to 2 µM of mycalolide-B or its vehicle control for 1 h. *P < 0.05 vs. remodeled + mycalolide-B or remodeled + vehicle. Figure redrawn from Castorena-Gonzalez et al. (112) with permission.

Figure 8.

Summary of proposed vascular smooth muscle cell (VSMC)-related mechanisms contributing to leg vascular dysfunction with prolonged sitting and likely amplified with superimposition of cardiovascular disease risk factors such as advanced age, obesity, and hypertension. EC, endothelial cell; ECM, extracellular matrix; Kir, inwardly rectifying potassium channel; NE, norepinephrine; ANG II, angiotensin II; ROS, reactive oxygen species; MMP, matrix metalloproteinase; NO, nitric oxide; TG2, tissue transglutaminase; RhoA, Ras homolog family member A; LIMK, LIM kinase; ATR, angiotensin receptor; ADR-α, α-adrenergic receptor.