Magnitude Of Flow-mediated Dilation Research Articles

ISH ADA-01 Critical role of telomerase in regulating cerebral vascular function and redox environment

Flow mediated dilation (FMD) is the most physiological relevant form of endothelial-mediated vasodilation. Our laboratory has previously shown that telomerase, a ribo-nucleoprotein that counteracts telomere shortening, has a protective effect on endothelial function under conditions of oxidative stress in the human microcirculation. In the presence of coronary artery disease, decreased telomerase activity contributes to a shift in the mediator of FMD from atheroprotective nitric oxide (NO) to pro-inflammatory and atherogenic hydrogen peroxide (H2O2). Endothelial dysfunction in the cerebral vasculature has been directly linked to cerebral microbleeds and cognitive decline in models of stroke and dementia, thus we hypothesized that TERT plays a critical role in maintaining normal NO-mediated vasodilation in the cerebral vasculature. Using Crisp/Cas9 we generated a novel deletion model for TERT (the catalytic subunit of telomerase) in rats on the WKY background. Middle cerebral arteries (MCA) were isolated from wildtype (WT) and TERT−/− rats, prepared for videomicroscopy, and FMD was measured. The mediator of FMD was determined using a pharmacological inhibitor of NO-synthase (L-NAME) and a scavenger of H2O2 (PEG-catalase). No changes in the magnitude of FMD were observed in MCA from TERT−/− compared to WT rats (max dilation: TERT−/− 71.97 ± 14.7% vs. WT 80.9 ± 10.6%; n≥3; p > 0.05 two way ANOVA RM). In WT animals, pre-incubation with L-NAME abolished FMD while PEG-catalase had no effect on FMD (max dilation: L-NAME -5.8 ± 7%; Peg-catalase 76.6 ± 8.6%; n≥3; p < 0.05). Conversely, in TERT−/− animals FMD was not affected in the presence of L-NAME but abolished in presence of Peg-catalase (max dilation: L-NAME 57.8.0 ± 16.5%; Peg-catalase −4.3 ± 2.4%; n≥3; p < 0.05). Telomerase deficiency causes a switch in the vasoactive mediator of FMD from NO to H2O2, creating a pro-oxidative environment in the cerebral vasculature. Understanding the regulatory role of telomerase in mediating this mechanistic switch may provide a novel therapeutic strategy for the treatment or prevention of cerebrovascular dysfunction.

CO-04: Aging altered epoxyeicosatrienoic acid-mediated regulation of peripheral conduit artery diameter at rest and during sustained flow-mediated dilatation

Background We evaluated the evolution of NO and CYP450 epoxygenases-derived epoxyeicosatrienoic acids (EETs) availability in conduit arteries during aging. Methods Radial artery diameter and mean wall shear stress were determined in 83 subjects (19 to 71 years old) at rest and during sustained endothelium-dependent flow-mediated dilatation induced by hand skin heating. The role of EETs and NO was assessed using the brachial infusion of inhibitors of cytochrome P450 epoxygenases (fluconazole) and NO-synthase (L-NMMA). Results Aging increased the reduction in basal arterial diameter induced by L-NMMA and nitrite plasma levels, an indicator of NO bioavailability. At the opposite, aging decreased EETs plasma levels and tended to reduce the basal vasoconstriction induced by fluconazole. Moreover, aging blunted the synergistic vasoconstricting effect of L-NMMA+fluconazole. Furthermore, the magnitude of flow-mediated dilatation was independently and negativelyrelated with age, together with baseline diameter and systolic blood pressure, and positively correlated with shear stress variation during heating. The increase in local nitrite level during heating and the decrease in flow-mediated dilatation induced by L-NMMA were not affected by aging. In contrast, aging tended to reduce the release of EETs and reduced the inhibitory effect of fluconazole alone and combined with L-NMMA on flow-mediated dilatation. Conclusions These results show that aging primarily impairs EETs availability and alters NO/EETs balance, contributing to promote endothelial dysfunction at the level of peripheral conduit arteries.

Microarray-Based Characterization of a Colony Assay Used to Investigate Endothelial Progenitor Cells and Relevance to Endothelial Function in Humans

An assay proposed to quantify endothelial progenitor cell (EPC) colonies in humans was investigated to determine the phenotype of recovered cells and their relevance to in vivo endothelial function. Twelve sedentary subjects participating in a worksite wellness program underwent endothelial flow-mediated dilation (FMD) testing of the brachial artery and blood sampling for EPC colony assay. Microarray-based genotypic characterization of colonies showed surface markers consistent with T lymphocyte phenotype, but not with an EPC (CD34, CD133, VEGFR-2) or endothelial (CD146) phenotype. Gene expression patterns more closely matched T lymphocytes (r=0.87) than endothelial cells (r=0.66) in our microarray database. Flow cytometry of colonies confirmed large populations of CD3+CD45+ T cells (>75%) and few CD146+CD45- endothelial cells (<1%). Further, there was no correlation between colony number and the magnitude of FMD (r=-0.1512, P=0.6389). After exercise training, subjects improved FMD, from 6.7+/-2.0 to 8.7+/-1.9% (P=0.0043). Colonies also increased (P=0.0210), but without relation to FMD (r=0.1074, P=0.7396). T lymphocyte phenotype persisted after exercise (r=0.87). Cells in a commonly used EPC colony assay have a gene expression and cell surface marker profile consistent with a predominance of T lymphocytes and have an unclear relevance to endothelial function, either before or after exercise training.

Determinants of coronary arterial flow‐mediated dilatation following percutaneous coronary intervention

It has previously been observed that coronary diameter may increase following relief of flow-limiting obstruction. Flow mediated dilatation (FMD) is a fundamental adaptive mechanism for arteries, which is dependent on intact endothelial function. We thus aimed to characterize whether the degree of this flow-mediated dilatation was related to risk factors, which may impair endothelial function. We measured coronary diameter with quantitative angiography before and after relief of chronic total or subtotal (>or=99%) occlusion in 171 patients, in which TIMI-0 or TIMI-1 flow was rapidly restored to TIMI-3 (with attendant increase in flow hypothesized to result in FMD). Of the 171 patients, 73% were male, 62% were current or ex-smokers, 47% were diabetic, 53% had hypertension, 64% had dyslipidemia (documented hypercholesterolemia or total cholesterol >5.0 mg/dL) and 65% were taking statin therapy. Mean vessel diameter was 2.8 +/- 0.7 mm and flow-mediated dilatation measured 15.1% +/- 20.1% in target vessel, compared with 1.6 +/- 3.1 in control vessels (P < 0.05). FMD was strongly and inversely related to baseline vessel diameter (r = -0.48, P < 0.001). The degree of vessel dilation correlated negatively with the presence of diabetes (r = -0.33, P < 0.001), smoking (r = -0.30, P < 0.001) and extent of coronary artery disease (CAD, r = -0.17, P = 0.01) and positively with the use of statins (r = 0.27, P = 0.001). These factors, apart from extent of CAD, remained significant predictors of FMD on multivariate analysis. FMD occurs in human coronary arteries following restoration of flow. The magnitude of FMD appears related to vascular risk factors and their treatment.

The Magnitude of FMD is Determined by the Area Under the Curve of the Reactive Hyperemia

The reactive hyperemia (RH) test commonly used to assess flow mediated dilation (FMD) in human conduit arteries creates a large, transient shear stimulus. PURPOSE: To identify the relative importance of the peak vs. the duration of shear rate in determining the peak FMD response. METHODS: Brachial artery (BA) blood flow velocity was measured with Doppler ultrasound and echo ultrasound BA images were analyzed with automated edge detection software (shear rate = velocity/diameter). 1) Duration manipulation experiment (DME) - 10 healthy males underwent six RH trials with different hyperemic durations (10s–2 min). The duration of hyperemia was manipulated by re-inflating the occlusion cuff after 10s, 20s, 30s, 40s, or 50s. 2) Peak manipulation experiment (PME) - 8 healthy males underwent three RH trials. Three peak shear rate magnitudes (large (L), medium (M) and small (S)) were created by applying BA compression upon cuff release. The post-peak area under the curve (AUC) of shear rate was also controlled with arterial compression. RESULTS: DME - Peak shear rate (AUC in 1st 9s post cuff release) was not different between trials (P = 0.309). Re-inflation of the occlusion cuff at times indicated resulted in the remaining AUC being significantly different between trials (10 s −2025.37 ±504.21; 20 s −3393.54±857.03; 30s-4329.30±1198.47; 40s–4784.20±1345.00; 50s–5215.83±1466.63 2min −7124.39±2160.67 p<0.001). The respective peak % changes in diameter were: 10s-2.67±1.34; 20s-6.21±1.95; 30s-7.89±2.95; 40s-8.3±3.21; 50s-7.91±3.20; 2min-9.34±4.10, with 10 and 20 s being significantly less than 2min (p<0.001). PME - Peak shear rate (AUC in 1st 9s post cuff release) was significantly different between trials (L- 998.04±305.16, M-700.66±225.89, S-487.48±1 67.55 p<0.001). The AUC of the post-peak shear rate was not significantly different between trials (P = 0.695). The peak % change in diameter was not significantly different between trials (L-6.99±2.72%, M-7.56±2.6%, S-6.87±1.99%, P = 0.437). CONCLUSIONS: Manipulation of the duration of the shear rate had an impact on the peak FMD response while independent manipulation of peak shear rate did not. We conclude that the shear rate AUC following the peak is a critical determinant of the peak FMD response and needs to be accounted for in quantifying the stimulus for FMD. Supported by ACSM Doctoral Research Grant, NSERC, CFI New Opportunities Fund

Flow-mediated dilation in human brachial artery after different circulatory occlusion conditions.

Different magnitudes and durations of postocclusion reactive hyperemia were achieved by occluding different volumes of tissue with and without ischemic exercise to test the hypotheses that flow-mediated dilation (FMD) of the brachial artery would depend on the increase in peak flow rate or shear stress and that the position of the occlusion cuff would affect the response. The brachial artery FMD response was observed by high-frequency ultrasound imaging with curve fitting to minimize the effects of random measurement error in eight healthy, young, nonsmoking men. Reactive hyperemia was graded by 5-min occlusion distal to the measurement site at the wrist and the forearm and proximal to the site in the upper arm. Flow was further increased by exercise during occlusion at the wrist and forearm positions. For the two wrist occlusion conditions, flow increased eightfold and FMD was only 1 to 2% (P > 0.05). After the forearm and upper arm occlusions, blood flow was almost identical but FMD after forearm occlusions was 3.4% (P < 0.05), whereas it was significantly greater (6.6%, P < 0.05) and more prolonged after proximal occlusion. Forearm occlusion plus exercise caused a greater and more prolonged increase in blood flow, yet FMD (7.0%) was qualitatively and quantitatively similar to that after proximal occlusion. Overall, the magnitude of FMD was significantly correlated with peak forearm blood flow (r = 0.59, P < 0.001), peak shear rate (r = 0.49, P < 0.002), and total 5-min reactive hyperemia (r = 0.52, P < 0.001). The prolonged FMD after upper arm occlusion suggests that the mechanism for FMD differs with occlusion cuff position.

Morning attenuation of endothelium-dependent, flow-mediated dilation in healthy young men: possible connection to morning peak of cardiac events?

Brachial artery flow-mediated dilation (FMD), a noninvasive, widely used clinical index of endothelial function and magnitude of FMD, has been reported to be closely related to many coronary risk factors and coronary atherosclerosis. However, there has been no study that examines the diurnal change of FMD. We designed this study to reveal the diurnal variation of FMD in healthy volunteers. We examined FMD in response to reactive hyperemia by high resolution ultrasound in 13 healthy young men (age 25-32) at four different times over the course of a day. Mean measures of brachial artery FMD was 4.0% at 8:00, 5.3% at 12:00, 9.7% at 17:00, and 6.9% at 21:00 hours. Flow-mediated dilation at 8:00 and at 12:00 hours was significantly lower than that at 17:00 (p < 0.05). These results show that endothelial function has diurnal variation and is significantly attenuated in the morning. Morning attenuation of endothelial function should be recognized in clinical research and may play an important role in the circadian variation of the occurrence of acute cardiovascular events.