Traditionally thought of primarily as the predominant regulator of myocardial perfusion, it is becoming more accepted that the human coronary microvasculature also exerts a more direct influence on ...the surrounding myocardium. Coronary microvascular dysfunction (CMD) not only precedes large artery atherosclerosis, but is associated with other cardiovascular diseases such as heart failure with preserved ejection fraction and hypertrophic cardiomyopathy. It is also highly predictive of cardiovascular events in patients with or without atherosclerotic cardiovascular disease. This review focuses on this recent paradigm shift and delves into the clinical consequences of CMD. Concepts of how resistance arterioles contribute to disease will be discussed, highlighting how the microvasculature may serve as a potential target for novel therapies and interventions. Finally, both invasive and non‐invasive methods with which to assess the coronary microvasculature both for diagnostic and risk stratification purposes will be reviewed.
Chronic administration of exogenous adiponectin restores nitric oxide (NO) as the mediator of flow-induced dilation (FID) in arterioles collected from patients with coronary artery disease (CAD). ...Here we hypothesize that this effect as well as NO signaling during flow during health relies on activation of Adiponectin Receptor 1 (AdipoR1). We further posit that osmotin, a plant-derived protein and AdipoR1 activator, is capable of eliciting similar effects as adiponectin. Human arterioles (80-200 μm) collected from discarded surgical adipose specimens were cannulated, pressurized, and pre-constricted with endothelin-1 (ET-1). Changes in vessel internal diameters were measured during flow using videomicroscopy. Immunofluorescence was utilized to compare expression of AdipoR1 during both health and disease. Administration of exogenous adiponectin failed to restore NO-mediated FID in CAD arterioles treated with siRNA against AdipoR1 (siAdipoR1), compared to vessels treated with negative control siRNA. Osmotin treatment of arterioles from patients with CAD resulted in a partial restoration of NO as the mediator of FID, which was inhibited in arterioles with decreased expression of AdipoR1. Together these data highlight the critical role of AdipoR1 in adiponectin-induced NO signaling during shear. Further, osmotin may serve as a potential therapy to prevent microvascular endothelial dysfunction as well as restore endothelial homeostasis in patients with cardiovascular disease.
Background Elevated levels of ceramide, a sphingolipid known to cause a transition from nitric oxide (NO)- to hydrogen peroxide-dependent flow-induced dilation (FID) in human arterioles, correlate ...with adverse cardiac events. However, elevations of ceramide are associated with changed concentrations of other sphingolipid metabolites. The effects of sphingolipid metabolites generated through manipulation of this lipid pathway on microvascular function are unknown. We examined the hypothesis that inhibition or activation of the ceramide pathway would determine the mediator of FID. Methods and Results Using videomicroscopy, internal diameter changes were measured in human arterioles collected from discarded adipose tissue during surgery. Inhibition of neutral ceramidase, an enzyme responsible for the hydrolysis of ceramide, favored hydrogen peroxide-dependent FID in arterioles from healthy patients. Using adenoviral technology, overexpression of neutral ceramidase in microvessels from diseased patients resulted in restoration of NO-dependent FID. Exogenous sphingosine-1-phosphate, a sphingolipid with opposing effects of ceramide, also restored NO as the mediator of FID in diseased arterioles. Likewise, exogenous adiponectin, a known activator of neutral ceramidase, or, activation of adiponectin receptors, favored NO-dependent dilation in arterioles collected from patients with coronary artery disease. Conclusions Sphingolipid metabolites play a critical role in determining the mediator of FID in human resistance arterioles. Manipulating the sphingolipid balance towards ceramide versus sphingosine-1-phosphate favors microvascular dysfunction versus restoration of NO-mediated FID, respectively. Multiple targets exist within this biolipid pathway to treat microvascular dysfunction and potentially improve patient outcomes.
Abstract
Background
Hypotension that is resistant to phenylephrine is a complication that occurs in anesthetized patients treated with angiotensin converting enzyme (ACE) inhibitors. We tested the ...hypothesis that Ang 1–7 and the endothelial Mas receptor contribute to vasodilation produced by propofol in the presence of captopril.
Methods
The internal diameters of human adipose resistance arterioles were measured before and after administration of phenylephrine (10
–9
to 10
–5
M) in the presence and absence of propofol (10
–6
M; added 10 min before the phenylephrine) or the Mas receptor antagonist A779 (10
–5
M; added 30 min before phenylephrine) in separate experimental groups. Additional groups of arterioles were incubated for 16 to 20 h with captopril (10
–2
M) or Ang 1–7 (10
–9
M) before experimentation with phenylephrine, propofol, and A779.
Results
Propofol blunted phenylephrine-induced vasoconstriction in normal vessels. Captopril pretreatment alone did not affect vasoconstriction, but the addition of propofol markedly attenuated the vasomotor response to phenylephrine. A779 alone did not affect vasoconstriction in normal vessels, but it restored vasoreactivity in arterioles pretreated with captopril and exposed to propofol. Ang 1–7 reduced the vasoconstriction in response to phenylephrine. Addition of propofol to Ang 1–7-pretreated vessels further depressed phenylephrine-induced vasoconstriction to an equivalent degree as the combination of captopril and propofol, but A779 partially reversed this effect.
Conclusions
Mas receptor activation by Ang 1–7 contributes to phenylephrine-resistant vasodilation in resistance arterioles pretreated with captopril and exposed to propofol. These data suggest an alternative mechanism by which refractory hypotension may occur in anesthetized patients treated with ACE inhibitors.
The sphingolipid sphingosine‐1‐phosphate (S1P) has emerged as a regulator of microvascular tone in animals, however its role in the human microcirculation remains unknown. Pre‐clinical studies ...suggest that S1P‐induced activation of two endothelial‐specific receptors, S1PR1 and S1PR3,elicits nitric‐oxide (NO)‐mediated vasodilation. As opposed to S1PR1, the S1PR3 pathway also activates NADPH oxidase (NOX) to produce reactive oxygen species (ROS). We therefore hypothesized that S1P induces vasodilation in the human microvasculature through activation of S1PR1 and S1PR3 in both a ROS‐independent and ROS‐dependent manner, respectively. Human microvessels (100‐200µm in diameter) were prepared for videomicroscopy. Following equilibration, arterioles were pre‐constricted with endothelin‐1 to 30‐70% of their passive diameters. Luminal diameter was measured and recorded in regular intervals (1 min) in response to increasing concentrations of S1P (10‐12 to 10‐6 M) in the presence or absence of the S1PR1 receptor antagonist W146 (10‐5M), S1PR3 antagonist CAY10444 (10‐5 M), nitric oxide synthase inhibitor Nω‐nitro‐ʟ‐arginine (L‐NAME, 10‐4 M), NO scavenger 2‐4‐carboxyphenyl‐4,4,5,5‐tetramethylimidazoline‐1‐oxyl‐3‐oxide (c‐PTIO, 10‐4 M), polyethylene glycol‐catalase (peg‐Cat, 500U/ml), NADPH oxidase inhibitor apocynin (3x10‐4 M), NOX‐2 inhibitor GSK2795039 (10‐6 M) or the NOX‐4 inhibitor GKT137831 (10−6 M). S1P induced vasodilation in a dose‐responsive manner with a maximum dilation of 56.5%±4.9, n=12 (mean±SEM). Dilation was abolished during inhibition of S1PR1 (1.5%±5.2, n=4) and was reduced during inhibition of S1PR3 (19.7%±8.7, n=6). Both L‐NAME and c‐PTIO inhibited S1P‐induced dilation (13.0%±7.5, n=4 and 11.2%±3.3, n=4, respectively). Interestingly, dilation was nearly completely inhibited by peg‐Cat (11%±6.6, n=4), apocynin (12.5%±4.4, n=3) and the NOX‐4 inhibitor (7.2%±3.1, n=4). Dilation was also partly reduced during inhibition of NOX‐2 (25.6%±7.8, n=4). These data suggest that S1P‐induced dilation occurs through activation of S1PR1 and S1PR3 through formation of NO and NOX‐4‐generated H2O2. These translational studies highlight the inter‐species variation observed in vascular signaling and provide insight into the mechanism by which S1P regulates microvascular resistance in humans.
Increased plasma ceramide levels and microvascular dysfunction are both independent risk factors for major adverse cardiac events (MACE). We have previously shown that chronic exposure to exogenous ...ceramide promotes microvascular endothelial dysfunction, defined as hydrogen peroxide (H2O2)‐mediated flow‐induced dilation (FID) as opposed to dilation due to formation and release of endothelial nitric oxide (NO). Interestingly, ceramide and its metabolites (e.g. sphingosine‐1‐phosphate; S1P) have also been shown to stimulate production of NO. Our previous studies indicate that activation of the ceramide‐producing enzyme neutral sphingomyelinase (NSmase) in necessary for NO‐mediated FID. We therefore hypothesized that formation of S1P is responsible for cellular increases in NO from both acute exogenous administration as well as endogenous formation of ceramide from shear. Human resistance arterioles (100‐200µm) were dissected from adipose tissue collected from healthy patients and were prepared for videomicroscopy. Increasing doses of exogenous C2 ceramide (10‐9 to 10‐5 M) were administered in the absence or presence of the NO scavenger c‐PTIO (1µM) and a sphingosine kinase inhibitor (SpKi, 5µM). We observed a dose‐response increase in arteriolar dilation from ceramide (46.9%±10.3 of maximal dilator capacity±SEM, n=3) that was impaired by c‐PTIO (15%±6.0, n=3). Ceramide‐induced dilation was also decreased in the presence of the SpKi (20.8%±4.0, n=3). To examine the role of S1P formation in NO‐mediated FID, SpKi was administered to healthy arterioles (5µM, 30 min) prior to initiating flow (pressure gradient 5‐100cm H2O) and a dramatic decrease in overall dilation was observed (7.7%±8.0, n=3). Together these findings suggest that NO generated from ceramide is primarily due to the formation of S1P, a process also critical to maintain FID in arterioles from healthy individuals.
Abstract only
Precise regulation of vascular resistance is critical to maintain proper tissue perfusion. Recent data in animal models have shown that sphingolipids, a ubiquitous class of bioactive ...lipid messengers, can influence vasomotor tone via dilation or constriction. It has been suggested that sphingosine‐1‐phosphate (S1P) elicits vasodilation through generation of endothelial cell nitric oxide (NO) via activation of sphingosine‐1‐phosphate receptor 1 (S1PR
1
). We therefore examined the hypothesis that S1P is a regulator of human microvascular tone and elicits dilation via S1PR
1
‐induced increase in NO. Arterioles from healthy human adipose tissue were dissected and prepared for videomicroscopy, equilibrated and pre‐constricted with endothelin 1. Changes in arteriolar luminal diameter were recorded in response to increasing concentrations of S1P (10
−12
to 10
−6
M) in the presence or absence of the S1PR
1
receptor antagonist W146 (10
−5
M), nitric oxide synthase (NOS) inhibitor N
ω
‐nitro‐
l
‐arginine (L‐NAME, 10
−4
M) or hydrogen peroxide (H
2
O
2
) scavenger polyethylene glycol‐catalase (peg‐Cat, 500U/ml). Administration of exogenous S1P induced vasodilation in healthy vessels to a maximal dilation of 63.7%±5.0, n=8 (mean±SEM). As with previous animal studies, vascular constriction was observed with higher, but still physiological levels of S1P (10
−6
M). Vasodilation due to S1P was reduced following inhibition of S1PR
1
compared to S1P alone (1.8%±4, n=5). In the presence of L‐NAME, S1P‐induced dilation decreased (9%±7.9, n=6). Interestingly, addition of peg‐Cat, an H
2
O
2
scavenger, also decreased S1P‐induced dilation (11.1%±6.6, n=3). Together these data suggest that S1PR
1
is a critical mechanistic component of S1P‐induced vasodilation in the human vasculature. Further, the S1P vasodilatory pathway may require formation of H
2
O
2
in addition to NO.
Support or Funding Information
This research was supported by National Institute of Health (NHLBI) K08 HL141562‐02 (JKF)
Endothelial dysfunction, or the loss of nitric oxide (NO)‐mediated dilation to shear stress (flow‐induced dilation; FID), is observed in human adipose arterioles from patients with coronary artery ...disease (CAD). FID is maintained during CAD by release of the mitochondrial‐derived, pro‐inflammatory dilator, H2O2. Ceramide, a sphingolipid that when elevated in plasma is an independent risk factor for future cardiac events, also induces this transition in human arterioles (10µM, 16‐20hrs). The initial events leading to ceramide‐induced endothelial dysfunction are unknown. Here we hypothesize that activation of NADPH oxidase 2 (NOX2), an endothelial membrane enzyme activated by acute stress, occurs prior to the formation of mitochondrial H2O2 in arterioles exposed to chronic ceramide. Human arterioles (100‐250µm in diameter) were dissected from discarded surgical adipose tissues and prepared for videomicroscopy. Following preconstriction with endothelin‐1, internal diameters were measured in response to increased flow. To determine the minimum exposure time necessary to convert to H2O2‐dependent FID in arterioles exposed to ceramide, microvessels from healthy nonCAD adults were incubated with C2 ceramide for 30 min, 2 hrs, and 4 hrs. FID remained mediated by NO at 30min and 2hrs, however, following a 4hr incubation, FID was significantly impaired in the presence of PEG‐catalase (44.6% of maximal dilator capacity ±13.8 (SEM), n=8, p<0.05, one‐way ANOVA*) compared to vehicle control (83.5%±5.0, n=8). The source of H2O2 generated during FID in arterioles treated for 4hr with ceramide was not mitochondrial as increases in fluorescent intensity of mito peroxy yellow 1 (mitoPY1) were not observed during maximal flow. To test whether NOX2 is the initial source of H2O2 during FID in arterioles exposed to 4hr ceramide, nonCAD arterioles were first treated with the NOX2 inhibitor GSK2795039 (NOX2i, 10‐6 M) or underwent intraluminal administration of siRNA to decrease NOX2 expression prior to treatment with ceramide. FID was reduced in vessels treated with both ceramide and the NOX2 inhibitor in the presence of cPTIO (NO scavenger) (50.2%±30.1, n=4* vs. NOX2 inhibitor and ceramide alone 80.8%±6.4, n=7). L‐NAME also impaired FID in vessels with reduced expression of NOX2 and subsequently treated with ceramide (10.8%±38.9, n=3* compared to siNOX2 and ceramide alone 85.5%±5.8, n=3). This suggests that activation of NOX2 may be the initial event in the conversion of NO‐ to H2O2‐dependent FID due to ceramide and may serve as a potential therapeutic target in those at risk for developing cardiovascular disease due to increased plasma ceramide.
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