The authors would like to thank the many subjects and patients who have participated in our studies in which we have evaluated endothelial function using the FMD test.

NOTE: The following FMD procedure is routinely conducted during vascular assessment studies in the Laboratory of Integrative Vascular and Exercise Physiology (LIVEP). All procedures followed the principles of the Declaration of Helsinki and were approved by the Institutional Review Board at Georgia Regents University. All participants were informed of the objectives and possible risks of the technique before written consent for participation was obtained. Figure 1 illustrates a schematic summary of the essential elements that should be considered for the ultrasound assessment of brachial artery FMD.
1. Subject Preparation (Prior to Arrival)
<ol>
	<li>Confirm that the participant has abstained from practicing exercise (&#8805;12 hr), caffeine (&#8805;12 hr), smoking or smoke exposure (&#8805;12 hr), vitamin supplementation (&#62;72 hr) and any medication (&#8805;4 hr half-lives of the drug, non-steroidal anti-inflammatory agents for 1 day and aspirins for 3 days).</li>
	<li>Ensure that the participant is under fasting conditions or has only consumed low-fat meals4 prior to testing.</li>
	<li>When testing premenopausal women, it is suggested to conduct the FMD protocol during the menses phase of the menstrual cycle to limit the impact of endogenous estrogens and progesterones8,32,33.</li>
</ol>
2. Subject Preparation (Upon Arrival)
<ol>
	<li>Prior to measurement acquisition, verify that the subject is resting in a supine position in a quiet, temperature-controlled (22 &#176;C to 24 &#176;C) room for approximately 20 min to achieve a hemodynamic steady state.</li>
	<li>Attach a 3-lead ECG in the standard limb lead II position. Using American standard instrumentation, place the white/negative polarity lead just below the clavicle on the right shoulder. Connect the black/ dual polarity lead below the left clavicle near the shoulder and connect the red/positive polarity lead below the left pectoral muscle in the lateral base of the chest.</li>
	<li>Extend the subject&#39;s arm laterally at about 80&#176; of shoulder abduction and secure the distal forearm in a vacuum packed pillow to maintain accurate position of the arm during the measurement (Figure 2).</li>
	<li>Place the forearm cuff immediately distal to the medial epicondyle and ensure that nothing is touching the cuff, including the table below (Figure 2).</li>
</ol>
3. Baseline Measurements
<ol>
	<li>Mapping the Brachial Artery with the Ultrasound:
	<ol>
		<li>While holding the probe with the hand, position it cross-sectionally and start scanning the inner side of the upper arm beginning at the insertion of the bicep and proceeding proximally.</li>
		<li>Within B-mode (gray-scale), identify the brachial artery and collateral vessels and use color flow (CF) mode to help confirm the location of the artery. Interpret the color and pulsatility carefully considering the direction of the transducer to ensure assessment of the artery and not the vein. 
		NOTE: With the probe indicator facing the head, red color means flow toward the transducer (arterial flow), while blue means flow away (venous flow).</li>
	</ol>
	</li>
	<li>Identification of Brachial Artery:
	<ol>
		<li>After finding the brachial artery, rotate the probe 90&#176; to scan the arm longitudinally. Obtain the image between 2 to 10 cm above the antecubital fossa.</li>
		<li>Identify anatomical landmarks such as veins and fascial planes for multiple assessments in the same subject (Figure 3).</li>
	</ol>
	</li>
	<li>Securing the Probe:
	<ol>
		<li>Secure the probe in the stereotactic probe holder. Confirm the probe is appropriately fixed to avoid excessive movements. With the probe secured in the holder, ensure that the image is as good as the image that was obtained manually without the holder.</li>
	</ol>
	</li>
	<li>Optimizing the Resolution of the Image:
	<ol>
		<li>Optimize the image using the time gain controls (TGC&#39;s) with the probe secured. 
		NOTE: An optimal image is achieved when the clearest B-mode image from the anterior and posterior intimal interfaces between the lumen and vessel wall is obtained.</li>
		<li>Have the technician manually adjust the gain, focal points, dynamic range, and harmonics to get a clear and defined image of the near and far walls of the endothelium.</li>
	</ol>
	</li>
	<li>Duplex Doppler Mode:
	<ol>
		<li>Following B-mode acquisition, proceed to duplex scanning in the pulsed Doppler mode.</li>
		<li>Use a heel to toe approach with the probe inside the holder by rocking the transducer up on one end more than the other to adjust the brachial artery image and obtain an angle of insonation of 60&#176;.</li>
	</ol>
	</li>
	<li>Baseline Acquisition:
	<ol>
		<li>Obtain a satisfactory B-mode image that identifies the endothelial layers with clear intima-intima walls of the artery. Ensure that the Doppler signal appears sharp and clear sound with no muffles.</li>
		<li>Reset the ultrasound CINE loop by freezing and unfreezing the image. Press F1 to begin recording data on image software. Record baseline data for at least 30 sec. Analyze the average diameter and blood velocity for 30 sec to represent baseline values. Note: Different ultrasounds and software set-ups may require different sequences to obtain the required action.</li>
	</ol>
	</li>
</ol>
4. Vascular Occlusion Measurements
<ol>
	<li>Forearm Occlusion:
	<ol>
		<li>Rapidly inflate the forearm occlusion cuff, using compressed air, to supra-systolic pressures (250 mm Hg) for 5 min to induce arterial occlusion.</li>
		<li>After 4 min and 30 sec of forearm occlusion, begin acquiring data. 
		NOTE: Occlusion measurements will be represented by the last 30 sec of occlusion.</li>
	</ol>
	</li>
</ol>
5. Reactive Hyperemia (Post Cuff Release) Measurements
<ol>
	<li>Continuing to Acquire Data from Pre-cuff Release:
	<ol>
		<li>Deflate the cuff at 5 min.</li>
		<li>Maintain recording for two minutes following cuff release.</li>
	</ol>
	</li>
	<li>Following 2 min of post cuff release recording, stop and save the recordings. The highest 5 sec averaged interval throughout the 2 min post-occlusion collection period will be used to represent the peak hyperemic diameter.</li>
</ol>
6. Analysis of the Results: Edge Detection and Wall Tracking
<ol>
	<li>Due to the complexity of FMD analysis, use edge-detection and wall-tracking software throughout FMD testing for higher reproducibility according to manufacturer&#39;s instructions. 
	NOTE: This offline analysis is less operator dependent than the manual assessment and therefore improves the accuracy of the FMD data4,34-36. In addition, this off-line analysis system also permits the synchronization with the ECG for the identification of end-diastolic arterial diameters, avoiding distortion of pulse-related changes in diameter4. It should be noted that, although the use of ECG is endorsed to minimize pulsation variability, it is also possible to perform the FMD protocol without ECG gaiting37. Although not recommended, if edge computer-assisted analysis is unavailable, careful manual assessment of both diameter and velocities should be collected36.</li>
	<li>For the evaluation of vessel diameters, it is necessary to visually inspect each frame to determine the best placement of the ultrasonic calipers along the B-mode image38. 
	NOTE: Regardless of data analysis method, it is recommended to collect diameter and velocity data every 4 sec during the first 20 sec of reactive hyperemia and every 5 sec for the remaining post occlusion period4.</li>
</ol>

The authors have nothing to disclose.

Introduced in 198920, the FMD test has been widely used in humans as a non-invasive measure of endothelial function. FMD has not only been shown to predict future vascular-related disease risk19,52,53, lower FMD values have been show to strongly correlate with cardiovascular impairments24,25,54. Although there are other techniques to assess endothelial function, both invasively (coronary angiography) and non-invasively (venous plethysmography and finger plethysmography), FMD has been the ...

Cardiovascular disease is the leading cause of morbidity and mortality worldwide. Dysfunction of the vascular endothelium represents an initial phase toward the development of multiple vascular-related diseases1. Hence, an accurate assessment of endothelial function in humans represents an important technique that could help in understanding the etiology of multiple cardiovascular pathologies, with the ultimate goal of improving the efficacy of the treatment and prevention of disease.
Endothelium
The endothelium is a monolayer of cells that synthesizes numerous vasoactive substances, such as nitric oxide (NO), prostacyclins, endothelins, endothelial cell growth factor, interleukins, and plasminogen inhibitors2. Such factors contribute to the endothelium&#39;s function to regulate blood fluidity, vascular tone, platelet aggregation, permeability of plasma components and vessel wall inflammation2-4. Additionally, NO plays a key anti-atherogenic role in promoting vasodilation and maintaining endothelial integrity. NO regulates vessel tone and diameter through controlling the equilibrium between the delivery of oxygen to the tissues and their metabolic demand3,5. There are multiple endogenous, exogenous, and mechanical stimulator factors that induce endothelial NO synthase (eNOS) which synthesizes NO from L-arginine6,7. The most notable mechanical stimulus is shear stress. Wall shear stress contributes to greater activation of eNOS, resulting in NO production and subsequent smooth muscle relaxation4. For that reason the decrease in NO bioavailability is often used as a measure of endothelial dysfunction8.
Endothelium dysfunction
The imbalance between vasodilator and vasoconstrictor factors leads to a dysfunctional endothelium2. In addition, the release of inflammatory mediators and altered local shear forces may enhance the synthesis of endothelial derived reactive oxygen species (ROS). This upregulation in redox signaling not only modifies the integrity of the endothelium and reduces the synthesis of NO9, it can uncouple eNOS resulting in direct production of additional free radicals. Ultimately, this amelioration in NO bioavailability promotes vasoconstriction, vascular stiffness, and reduced arterial distensibility4.
The degree of dysfunction of the endothelium has been related with the severity of several pathologies such as hypertension10, atherosclerosis11, ischemic stroke12, diabetes13, preeclampsia14 or kidney diseases15 among others. Hence, there is vast interest to not only evaluate changes in endothelial function over time, but also following therapeutic interventions. Different methods have been used for the clinical assessment of endothelial function both invasively (cardiac catheterization and venous occlusion plethysmography3,16) and non-invasively (flow mediated dilation, radial artery tonometry and pulse contour analysis4,17,18) in coronary and peripheral circulations19.
Flow-mediated dilation
Flow mediated dilation (FMD) is a non-invasive, ultrasonic evaluation of endothelial function and has been correlated with the development of vascular health problems. Since its inception in 198920, FMD has been widely utilized as a reliable, in vivo method to evaluate predominately NO-mediated endothelial function in humans19,21,22. Indeed, the brachial artery FMD test has been associated with other invasive techniques23 and numerous investigations have described a strong inverse relationship between FMD and cardiovascular injury24,25 such that individuals with more vascular pathology exhibit a lower FMD25. Accordingly, these data emphasize the prognostic information that this technique can provide as it relates to future cardiovascular disease in asymptomatic subjects26-30.
During the FMD test, the diameters of the brachial artery are continuously measured at baseline and after the release of a circulatory arrest of the forearm. Upon cuff release, the induced-reactive hyperemia promotes an increase in shear stress mediated NO release and subsequent vasodilation19,31. FMD is expressed as the percent increase in arterial diameter following the release of the cuff compared with the diameter at baseline (FMD%).
Despite the increasing clinical interest in this technique, the FMD test is a physiological assessment and therefore, several variables need to be considered in order to conduct a precise assessment of endothelial function in humans. This article describes a standardized protocol and the recommended methodology to minimize the technical and biological issues to help improve the accuracy, reproducibility and interpretation of the FMD test.

<table><tbody><tr><td>Doppler ultrasound</td><td>GE Medical Systems&amp;nbsp;</td><td>Logiq 7</td><td>Essential to include Duplex mode for simultaneous acquisition of B-mode and Doppler</td></tr><tr><td>Electrocardiographic (ECG) gating&amp;nbsp;</td><td>Accusync Medical Research</td><td>Accusync 72</td><td></td></tr><tr><td>12-MHz Linear array transducer&amp;nbsp;</td><td>GE Medical Systems</td><td>11L-D</td><td>A high-resolution linear array probe is essential</td></tr><tr><td>Forearm occlusion cuff&amp;nbsp;</td><td>D.E. Hokanson</td><td>SC5</td><td>5 cm x 84 cm</td></tr><tr><td>Ultrasound transmission gel&amp;nbsp;</td><td>Parker</td><td>01-08</td><td></td></tr><tr><td>Rapid cuff inflator</td><td>D.E. Hokanson</td><td>E-20 AG101</td><td></td></tr><tr><td>Sterotactic-probe holder</td><td>Flexabar&amp;nbsp;</td><td>18047</td><td>Magnetic base fine adjustor</td></tr><tr><td>Edge detection analysis software</td><td>Medical Imaging Applications</td><td>Brachial Analyzer 5</td><td></td></tr></tbody></table>

ultrasound assessment of endothelial function: a technical guideline of the flow-mediated dilation test

Cardiovascular disease is the primary cause of mortality and a major cause of disability worldwide. The dysfunction of the vascular endothelium is a pathological condition characterized mainly by a disruption in the balance between vasodilator and vasoconstrictor substances and is proposed to play an important role in the development of atherosclerotic cardiovascular disease. Therefore, a precise evaluation of endothelial function in humans represents an important tool that could help better understand the etiology of multiple cardio-centric pathologies.
Over the past twenty-five years, many methodological approaches have been developed to provide an assessment of endothelial function in humans. Introduced in 1989, the FMD test incorporates a forearm occlusion and subsequent reactive hyperemia that promotes nitric oxide production and vasodilation of the brachial artery. The FMD test is now the most widely utilized, non-invasive, ultrasonic assessment of endothelial function in humans and has been associated with future cardiovascular events.
Although the FMD test could have clinical utility, it is a physiological assessment that has inherited several confounding factors that need to be considered. This article describes a standardized protocol for determining FMD including the recommended methodology to help minimize the physiological and technical issues and improve the precision and reproducibility of the assessment.

Baseline characteristics from an apparently healthy cohort group are presented in Table 1. The most common variables of FMD testing conducted in the Laboratory of Integrative Vascular and Exercise Physiology (LIVEP) are presented in Table 2. The following variables are considered the main FMD parameters to analyze by the published FMD tutorial4 and guidelines36.
Baseline an...

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Ultrasound Assessment of Endothelial Function: A Technical Guideline of the Flow-mediated Dilation Test

The flow mediated dilation (FMD) test is the most commonly utilized, non-invasive, ultrasound assessment of endothelial function in humans. Although the FMD test has been related with the prediction of future cardiovascular disease and events, it is a physiological assessment with many inherent confounding factors that need to be considered.

Cardiovascular disease is the primary cause of mortality and a major cause of disability worldwide. The dysfunction of the vascular endothelium is a ...

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17.4K Views.  Georgia Regents University. The flow mediated dilation (FMD) test is the most commonly utilized, non-invasive, ultrasound assessment of endothelial function in humans. Although the FMD test has been related with the prediction of future cardiovascular disease and events, it is a physiological assessment with many inherent confounding factors that need to be considered. 

Video: Ultrasound Assessment of Endothelial Function: A Technical Guideline of the Flow-mediated Dilation Test

Assessing Endothelial Vasodilator Function with the Endo-PAT 2000

The endothelium is a delicate monolayer of cells that lines all blood vessels, and which comprises the systemic and lymphatic capillaries. By virtue of the panoply of paracrine factors that it secretes, the endothelium regulates the contractile and proliferative state of the underlying vascular smooth muscle, as well as the interaction of the vessel wall with circulating blood elements. Because of its central role in mediating vessel tone and growth, its position as gateway to circulating immune cells, and its local regulation of hemostasis and coagulation, the the properly functioning endothelium is the key to cardiovascular health. Conversely, the earliest disorder in most vascular diseases is endothelial dysfunction.
In the arterial circulation, the healthy endothelium generally exerts a vasodilator influence on the vascular smooth muscle. There are a number of methods to assess endothelial vasodilator function. The Endo-PAT 2000 is a new device that is used to assess endothelial vasodilator function in a rapid and non-invasive fashion. Unlike the commonly used technique of duplex ultra-sonography to assess flow-mediated vasodilation, it is totally non-operator-dependent, and the equipment is an order of magnitude less expensive. The device records endothelium-mediated changes in the digital pulse waveform known as the PAT ( peripheral Arterial Tone) signal, measured with a pair of novel modified plethysmographic probes situated on the finger index of each hand. Endothelium-mediated changes in the PAT signal are elicited by creating a downstream hyperemic response. Hyperemia is induced by occluding blood flow through the brachial artery for 5 minutes using an inflatable cuff on one hand. The response to reactive hyperemia is calculated automatically by the system. A PAT ratio is created using the post and pre occlusion values. These values are normalized to measurements from the contra-lateral arm, which serves as control for non-endothelial dependent systemic effects. Most notably, this normalization controls for fluctuations in sympathetic nerve outflow that may induce changes in peripheral arterial tone that are superimposed on the hyperemic response.
In this video we demonstrate how to use the Endo-PAT 2000 to perform a clinically relevant assessment of endothelial vasodilator function.

A noninvasive procedure to assess endothelial function is demonstrated using the Endo-PAT 2000.

The endothelium is a delicate monolayer of cells that lines all blood vessels, and which comprises the systemic and lymphatic capillaries. By virtue of ...

Non-invasive Assessment of Microvascular and Endothelial Function

The authors have utilized capillaroscopy and forearm blood flow techniques to investigate the role of microvascular dysfunction in pathogenesis of cardiovascular disease. Capillaroscopy is a non-invasive, relatively inexpensive methodology for directly visualizing the microcirculation. Percent capillary recruitment is assessed by dividing the increase in capillary density induced by postocclusive reactive hyperemia (postocclusive reactive hyperemia capillary density minus baseline capillary density), by the maximal capillary density (observed during passive venous occlusion). Percent perfused capillaries represents the proportion of all capillaries present that are perfused (functionally active), and is calculated by dividing postocclusive reactive hyperemia capillary density by the maximal capillary density. Both percent capillary recruitment and percent perfused capillaries reflect the number of functional capillaries. The forearm blood flow (FBF) technique provides accepted non-invasive measures of endothelial function: The ratio FBFmax/FBFbase is computed as an estimate of vasodilation, by dividing the mean of the four FBFmax values by the mean of the four FBFbase values. Forearm vascular resistance at maximal vasodilation (FVRmax) is calculated as the mean arterial pressure (MAP) divided by FBFmax. Both the capillaroscopy and forearm techniques are readily acceptable to patients and can be learned quickly.
The microvascular and endothelial function measures obtained using the methodologies described in this paper may have future utility in clinical patient cardiovascular risk-reduction strategies. As we have published reports demonstrating that microvascular and endothelial dysfunction are found in initial stages of hypertension including prehypertension, microvascular and endothelial function measures may eventually aid in early identification, risk-stratification and prevention of end-stage vascular pathology, with its potentially fatal consequences.

Capillaroscopy is a non-invasive, relatively inexpensive methodology for directly visualizing the microcirculation. The forearm blood flow technique provides accepted non-invasive measures of endothelial function.

The authors have utilized capillaroscopy and forearm blood flow techniques to investigate the role of microvascular dysfunction in pathogenesis of ...

Technical Aspects of the Mouse Aortocaval Fistula

Technical aspects of creating an arteriovenous fistula in the mouse are discussed. Under general anesthesia, an abdominal incision is made, and the aorta and inferior vena cava (IVC) are exposed. The proximal infrarenal aorta and the distal aorta are dissected for clamp placement and needle puncture, respectively. Special attention is paid to avoid dissection between the aorta and the IVC. After clamping the aorta, a 25 G needle is used to puncture both walls of the aorta into the IVC. The surrounding connective tissue is used for hemostatic compression. Successful creation of the AVF will show pulsatile arterial blood flow in the IVC. Further confirmation of successful AVF can be achieved by post-operative Doppler ultrasound.

The goal is to produce an arteriovenous fistula that is simple and reproducible. This method does not use sutures or glue adhesive. Therefore the samples can be used with the least amount of foreign materials for analysis.

Technical aspects of creating an arteriovenous fistula in the mouse are discussed. Under general anesthesia, an abdominal incision is made, and the aorta ...

A Novel Application of Musculoskeletal Ultrasound Imaging

Ultrasound is an attractive modality for imaging muscle and tendon motion during dynamic tasks and can provide a complementary methodological approach for biomechanical studies in a clinical or laboratory setting. Towards this goal, methods for quantification of muscle kinematics from ultrasound imagery are being developed based on image processing. The temporal resolution of these methods is typically not sufficient for highly dynamic tasks, such as drop-landing. We propose a new approach that utilizes a Doppler method for quantifying muscle kinematics. We have developed a novel vector tissue Doppler imaging (vTDI) technique that can be used to measure musculoskeletal contraction velocity, strain and strain rate with sub-millisecond temporal resolution during dynamic activities using ultrasound. The goal of this preliminary study was to investigate the repeatability and potential applicability of the vTDI technique in measuring musculoskeletal velocities during a drop-landing task, in healthy subjects. The vTDI measurements can be performed concurrently with other biomechanical techniques, such as 3D motion capture for joint kinematics and kinetics, electromyography for timing of muscle activation and force plates for ground reaction force. Integration of these complementary techniques could lead to a better understanding of dynamic muscle function and dysfunction underlying the pathogenesis and pathophysiology of musculoskeletal disorders.

We describe a new ultrasound-based vector tissue Doppler imaging technique to measure muscle contraction velocity, strain and strain rate with sub-millisecond temporal resolution during dynamic activities. This approach provides complementary measurements of dynamic muscle function and could lead to a better understanding of mechanisms underlying musculoskeletal disorders.

Ultrasound is an attractive modality for imaging muscle and tendon  motion during dynamic tasks and can provide a complementary methodological  approach ...

Diagnostic Ultrasound Imaging of Mouse Diaphragm Function

Function analysis of rodent respiratory skeletal muscles, particularly the diaphragm, is commonly performed by isolating muscle strips using invasive surgical procedures. Although this is an effective method of assessing in vitro diaphragm activity, it involves non-survival surgery. The application of non-invasive ultrasound imaging as an in vivo procedure is beneficial since it not only reduces the number of animals sacrificed, but is also suitable for monitoring disease progression in live mice. Thus, our ultrasound imaging method may likely assist in the development of novel therapies that alleviate muscle injury induced by various respiratory diseases. Particularly, in clinical diagnoses of obstructive lung diseases, ultrasound imaging has the potential to be used in conjunction with other standard tests to detect the early onset of diaphragm muscle fatigue. In the current protocol, we describe how to accurately evaluate diaphragm contractility in a mouse model using a diagnostic ultrasound imaging technique.

Diagnostic ultrasound imaging has proven to be effective in diagnosing various respiratory diseases in human and animal subjects. We demonstrate a comprehensive ultrasound protocol utilized by Dr. Zuo's lab to analyze diaphragm kinetics specifically in mouse models. This is also a non-invasive research technique which can provide quantitative information on mouse respiratory muscle function.

Function analysis of rodent respiratory skeletal muscles, particularly the diaphragm, is commonly performed by isolating muscle strips using invasive ...

Ultrasound Assessment of Endothelial-Dependent Flow-Mediated Vasodilation of the Brachial Artery in Clinical Research

The vascular endothelium is a monolayer of cells that cover the interior of blood vessels and provide both structural and functional roles. The endothelium acts as a barrier, preventing leukocyte adhesion and aggregation, as well as controlling permeability to plasma components. Functionally, the endothelium affects vessel tone.
Endothelial dysfunction is an imbalance between the chemical species which regulate vessel tone, thombroresistance, cellular proliferation and mitosis. It is the first step in atherosclerosis and is associated with coronary artery disease, peripheral artery disease, heart failure, hypertension, and hyperlipidemia.
The first demonstration of endothelial dysfunction involved direct infusion of acetylcholine and quantitative coronary angiography. Acetylcholine binds to muscarinic receptors on the endothelial cell surface, leading to an increase of intracellular calcium and increased nitric oxide (NO) production. In subjects with an intact endothelium, vasodilation was observed while subjects with endothelial damage experienced paradoxical vasoconstriction.
There exists a non-invasive, in vivo method for measuring endothelial function in peripheral arteries using high-resolution B-mode ultrasound. The endothelial function of peripheral arteries is closely related to coronary artery function. This technique measures the percent diameter change in the brachial artery during a period of reactive hyperemia following limb ischemia.
This technique, known as endothelium-dependent, flow-mediated vasodilation (FMD) has value in clinical research settings. However, a number of physiological and technical issues can affect the accuracy of the results and appropriate guidelines for the technique have been published. Despite the guidelines, FMD remains heavily operator dependent and presents a steep learning curve. This article presents a standardized method for measuring FMD in the brachial artery on the upper arm and offers suggestions to reduce intra-operator variability.

Endothelial dysfunction is associated with numerous disease states and is predictive of adverse cardiovascular events in humans. Flow-mediated vasodilation (FMD) is a non-invasive ultrasound method of evaluating endothelial function. Methodological choices and operator experience may affect results. A systematic approach to FMD in human studies is discussed here.

The vascular endothelium is a monolayer of cells that cover the interior of blood vessels and provide both structural and functional roles. The ...

A Methodological Approach to Non-invasive Assessments of Vascular Function and Morphology

The endothelium is the innermost lining of the vasculature and is involved in the maintenance of vascular homeostasis. Damage to the endothelium may predispose the vessel to atherosclerosis and increase the risk for cardiovascular disease. Assessments of peripheral endothelial function are good indicators of early abnormalities in the vascular wall and correlate well with assessments of coronary endothelial function. The present manuscript details the important methodological steps necessary for the assessment of microvascular endothelial function using laser Doppler imaging with iontophoresis, large vessel endothelial function using flow-mediated dilatation, and carotid atherosclerosis using carotid artery ultrasound. A discussion on the methodological considerations for each of the techniques is also presented, and recommendations are made for future research.

The present article describes the methodological considerations for several non-invasive assessments of vascular function and morphology that are commonly used in medical research to assess different stages of atherosclerosis.

The endothelium is the innermost lining of the vasculature and is involved in the maintenance of vascular homeostasis.  Damage to the endothelium may ...

Ultrasound Assessment of Flow-Mediated Dilation of the Brachial and Superficial Femoral Arteries in Rats

Arterial vasodilation to increases in wall shear rate is indicative of vascular endothelial function. In humans, the non-invasive measurement of endothelial function can be achieved by employing the flow-mediated dilation technique, typically performed in the brachial or superficial femoral artery. Briefly, a blood pressure cuff placed distal to an ultrasound probe is inflated to a suprasystolic pressure, which results in limb ischemia. After 5 min of occlusion the cuff is deflated, resulting in reactive hyperemia and increases in wall shear rate that signal vasodilatory molecules to be released from the endothelium eliciting vasodilation. Despite the thousands of studies performing flow-mediated dilation in humans, surprisingly, no studies have performed this technique non-invasively in living rats. Considering the recent shift in focus to translational research, the establishment of guidelines for non-invasive measurement of flow-mediated dilation in rats and other rodents would be extremely valuable. In the following article, a protocol is presented for the non-invasive measurement of flow-mediated dilation in brachial and superficial femoral arteries of rats, as those sites are most commonly measured in humans.

Non-invasive assessment of endothelial function in humans can be determined by the flow-mediated dilation technique. Although thousands of studies have used this technique, no study has performed this technique non-invasively in rats. The following article describes non-invasive measurement of flow-mediated dilation in the brachial and superficial femoral arteries of rats.

Arterial vasodilation to increases in wall shear rate is indicative of vascular endothelial function. In humans, the non-invasive measurement of ...

Assessment of Human Adipose Tissue Microvascular Function Using Videomicroscopy

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. It is hypothesized that pro-atherogenic mediators released from fat tissues particularly in association with central/visceral adiposity may promote pathogenic vascular changes locally and systemically, and the notion that cardiovascular disease may be the consequence of adipose tissue dysfunction continues to evolve. Here, we describe a unique method of videomicroscopy that involves analysis of vasodilator and vasoconstrictor responses of intact small human arterioles removed from the adipose depot of living human subjects. Videomicroscopy is used to examine functional properties of isolated microvessels in response to pharmacological or physiological stimuli using a pressured system that mimics in vivo conditions. The technique is a useful approach to gain understanding of the pathophysiology and molecular mechanisms that contribute to vascular dysfunction locally within the adipose tissue milieu. Moreover, abnormalities in the adipose tissue microvasculature have also been linked with systemic diseases. We applied this technique to examine depot-specific vascular responses in obese subjects. We assessed endothelium-dependent vasodilation to both increased flow and acetylcholine in adipose arterioles (50 - 350 &#181;m internal diameter, 2 - 3 mm in length) isolated from two different adipose depots during bariatric surgery from the same individual. We demonstrated that arterioles from visceral fat exhibit impaired endothelium-dependent vasodilation compared to vessels isolated from the subcutaneous depot. The findings suggest that the visceral microenvironment is associated with vascular endothelial dysfunction which may be relevant to clinical observation linking increased visceral adiposity to systemic disease mechanisms. The videomicroscopy technique can be used to examine vascular phenotypes from different fat depots as well as compare findings across individuals with different degrees of obesity and metabolic dysfunction. The method can also be used to examine vascular responses longitudinally in response to clinical interventions.

Videomicroscopy systems are used to examine functional properties of isolated adipose tissue arterioles in response to physiological and pharmacological stimuli. This technique can be used to examine microvascular phenotypes in different adipose tissue domains in obese humans.

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. ...

Using Arterial Stiffness and Endothelial Function for Cardiovascular Assessment

Pulse-Wave Velocity, Flow-Mediated Dilation, and Carotid Intima-Media Thickness to Assess Cardiovascular Risk in Population with Metabolic Syndrome

Pulse-wave velocity (PWV), flow-mediated dilation (FMD), and carotid intima-media thickness (CIMT) are established methods used in research and clinical settings to assess arterial stiffness, endothelial function, and subclinical atherogenesis. These measurements may reflect vascular disease and atherosclerotic progression, which are major causes of adverse cardiovascular events. These methods are particularly valuable in determining cardiovascular dysfunction among populations with different risk factors, such as diabetes mellitus, hypertension, and other metabolic dysfunction-related conditions. They provide a non-invasive and reliable source of information that complements clinical practice. Early detection, risk assessment, and therapeutic decisions regarding cardiovascular disease can be achieved, ultimately contributing to improved patient outcomes. Traditional tools for evaluating cardiovascular disease do not reveal whether metabolic syndrome affects early subclinical cardiovascular disease in patients with obesity. Recent research has highlighted the importance of including arterial stiffness and endothelial function in a comprehensive cardiovascular assessment. Therefore, the aim of the present study is to describe methods that provide information on early subclinical vascular aging, endothelial dysfunction, and atherogenic disease, enabling vascular-targeted risk stratification among populations with obesity and different metabolic profiles.

Author Spotlight: Assessing the Cardiovascular Profile of Patients with Metabolic Syndrome

Endothelial/vascular aging and atherogenesis are key mechanisms that promote the development of cardiovascular diseases. The present protocol describes methods to evaluate arterial stiffness, endothelial dysfunction, and atherogenesis in patients with related risk factors, which are highly valuable in the cardiovascular research field.

Pulse-wave velocity (PWV), flow-mediated dilation (FMD), and carotid intima-media thickness (CIMT) are established methods used in research and clinical ...

Ultrasound Assessment of Endothelial Function: A Technical Guideline of the Flow-mediated Dilation Test

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