Name: Non-invasive Assessment of Microvascular and Endothelial Function (Video) | JoVE Journal
Uploaded: 2013-01-29T12:00:00.000+00:00
Duration: 5 min 41 s
Description: Learn non-invasive capillaroscopy and plethysmography techniques to visualize microcirculation and assess endothelial function using forearm blood flow measurements. This cost-effective method enables reliable functional capillary recruitment studies.

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5 minutes

This work was supported by Grant Award Numbers R01HL096593 and K23HL72825 from the National Heart, Lung, and Blood Institute. The content is solely the responsibility of Thomas Jefferson University and does not necessarily represent the official views of the National Heart, Lung, and Blood Institute or the National Institutes of Health.

Case Presentation (required, if applicable): N.A.; this is still an experimental research procedure, not yet used clinically.
Diagnosis, Assessment, and Plan (required, if applicable): N.A.; this is still an experimental research procedure, not yet used clinically.
Procedure (required): This part should include a step-by-step description of relevant procedures, meeting the guidelines below.
1. Capillary Microscopy (Figure 1)
<ol>
 <li>Our capillaroscopy technique was adapted from Serne and his colleagues 1. An exclusion criterion for this procedure is collagen vascular disease, since collagen vascular disease produces known capillary changes 2.</li>
 <li>Following a minimum 10-hr overnight fast and 20 min of seated rest, microvascular measurements are conducted for one half-hour between 7 and 11 am, in a quiet, temperature controlled room (maintained between 21.5-22.5 &deg;C), with the subject in the seated position and the left hand at heart level.</li>
 <li>Nailfold capillaries in the dorsal skin of the third finger are visualized using a stereomicroscope (Olympus; Center Valley, PA), linked to a 4 megapixel SPOT Insight monochrome digital camera (Model number IN-1400: Diagnostic Instruments; Sterling Heights, MI), and a laptop computer (Dell Latitude D600: Dell; Austin, TX). To limit movement, the left hand and forearm are loosely covered with a folded blanket, and rested on another folded blanket positioned at the base of the microscope.</li>
 <li>Nailbed illumination is achieved with a 250-W halogen fiber optic lamp (KL 2500LCD:Schott-Fostec; Elmsford, NY); additional illumination from a supplemental 150W fiber optic halogen light source (B&amp;B Microscopes, Ltd., Warrendale, PA) is used in darkly pigmented individuals.</li>
 <li>To visualize the capillaries, the 3.2x objective (Olympus 3.2/0.07) is used with a total system magnification of 38.4x.</li>
 <li>Using SPOT imaging software provided with the camera, light/dark contrast in the capillary photographs is enhanced using the same standard SPOT software function (stretching of bright and dark levels) to maximize visibility of the capillaries in all subjects.</li>
 <li>To quantify capillary density, digital photomicrographs are taken every 3-5 sec during each of three stages, at resting baseline, during postocclusive reactive hyperemia, and during venous occlusion. (a) At resting baseline, photomicrographs are taken over a three-minute period to detect capillaries perfused at rest. (b) During postocclusive reactive hyperemia, photomicrographs are taken to quantify functionally perfused capillaries (baseline plus reserve capillaries), as follows. First, an occlusion cuff on the left upper arm is inflated to 40 mm Hg above systolic pressure for 10 min. Photomicrographs are then taken during the first minute immediately following release of arterial occlusion, visualizing all functionally perfused capillaries. Lower capillary density following reactive hyperemia indicates impaired functional capillary recruitment, and therefore functional rarefaction. (c) During venous occlusion photomicrographs are taken to quantify maximal capillary density, which includes both perfused (with active red blood cell (RBC) motion) and nonperfused (filled with stagnant, non-moving RBCs) capillaries 3 as follows. Following ten minutes of rest after the postocclusive reactive hyperemia procedure, the arm cuff is inflated to 60 mm Hg for 60 sec, passively forcing blood into all patent capillaries present and photomicrographs were taken during this time. Since maximal capillary density includes all capillaries structurally present, a reduction in maximal capillary density indicates structural rarefaction.</li>
 <li>Capillary density is defined as the number of capillaries per square millimeter of nailfold skin, and is computed as the mean of four measurements obtained from the four most clearly focused images, least distorted by movement. In our studies, typical values for capillary counts (capillaries/mm2) have been 55-80 for baseline, 65-90 for post-ischemic, and 90-105 for venous occlusion. Values for percent capillary recruitment are typically between 5% and 25% (mean ~10-15%) and for percent perfused capillaries between 70% and 95% (mean ~80-90%), with values being lower among hypertensives than normotensives. The reproducibility of the counting procedure has been verified with three observers who performed independent manual assessments of photographs of 10 different subjects (Figure 2). The observers were blinded to the identity and blood pressure of these subjects. Following training, subsequent counts performed independently showed a high level of agreement. Average inter-rater and intra-rater discrepancies were of the order of 2-3 capillaries/mm2, and intraclass correlation coefficients were all greater than 0.90. Short-term variation of the capillaroscopy measures were of the same order of magnitude as inter-rater and intra-rater discrepancies (approximately 2 capillaries/mm2), but longer-term variation observed over 2-3 years was an order of magnitude larger (approximately 15 capillaries/mm2), indicating that longitudinal changes can be readily distinguished from rater variation. Reliability of the two capillary function measures was also high (intraclass correlation coefficient = 0.84 for percent capillary recruitment and 0.82 for percent perfused capillaries).</li>
 <li>
 The investigators now utilize a computer-based method for quantifying capillary density using Image-Pro Plus imaging software (Version 6.2, Media Cybernetics, Inc., Bethesda, MD: Figure 3). Pearson correlations between baseline, post-ischemic, and venous congestion counts done with the software and corresponding manual counts in 10 subjects were 0.78, 0.78, and 0.71 respectively (all p &lt; 0.05), indicating reasonable agreement between the two methods. Reliability of the computer-based counts is slightly lower than that of manual counts but still high (intraclass correlation coefficient = 0.91 for baseline, 0.86 for post-ischemic, and 0.84 for venous occlusion). We have unpublished data also demonstrating the association of automated counts with multiple cardiovascular risk factors including hypertension, which we are currently preparing for publication.</li>
 <li>Table 1 summarizes the capillary density measurements and calculations. 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. Lower values for these measures indicate functional capillary rarefaction.</li>
</ol>
2. Endothelial Function Assessment
<ol>
 <li>Endothelial function is assessed before and after postocclusive reactive hyperemia, using non-invasive plethysmography measurements of forearm blood flow, according to the method of Sivertsson,4 which utilizes the endothelium-dependent stimulus of reactive hyperemia to induce vasodilation.</li>
 <li>With the subject in the seated position following 10 min of supine rest, a mercury-in rubber strain gauge stretched to 10% beyond its resting length is looped around the subject's forearm 5 cm below the antecubital fossa.</li>
 <li>The strain-gauge is connected to a plethysmograph (EC-4: DE Hokanson, Inc; Bellevue, WA), which in turn is connected to a Doppler recorder (CW-1; DE Hokanson, Inc; Belleveue, WA).</li>
 <li>An upper arm occlusion cuff is applied, and the arm is suspended comfortably at heart level using a sling bandage connected to an adjustable intravenous pole. Systolic and diastolic blood pressures and heart rate are obtained with a Dinamap ProCare 100 automatic BP cuff (GE Healthcare, Piscataway, NJ) placed on the opposite arm.</li>
 <li>A pediatric cuff around the wrist is inflated to 200 mm Hg to occlude flow to the hand. The upper arm cuff is inflated to 50 mm Hg, deflated for 1.5 sec, and then re-inflated rapidly prior to each forearm blood flow measurement, obtained through expansion of the strain gauge placed around the forearm.</li>
 <li>Forearm blood flow (FBF) is measured at rested baseline (FBFbase) and again at postocclusive hyperemia-induced maximal vasodilation (FBFmax). For baseline blood flow measurements, four consecutive FBF curves are obtained within 30 sec (FBFbase).</li>
 <li>The occlusion cuff is then inflated to 40 mm Hg above systolic pressure for 10 min. Following release of arterial occlusion (postocclusive reactive hyperemia), four consecutive FBF curves are obtained within the first 30 sec of flow (FBFmax).</li>
 <li>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.5 Forearm vascular resistance at maximal vasodilation (FVRmax) is calculated as the mean arterial pressure (MAP) divided by FBFmax. FBFmaxduring reactive hyperemia is directly related to FBF after maximum infusion of intra-arterial acetylcholine, an endothelial-dependent vasodilator.6 Accordingly, FBFmax and the ratio FBFmax/FBFbase are accepted non-invasive measures of endothelial function.6-8 In addition, both FBFmax and FVRmax reflect resistance artery structural changes(increased wall/lumen ratio).9</li>
</ol>

<ol><li>Serne, E. H., Gans, R. O., ter Maaten, J. C., ter Wee, P. M., Donker, A. J., Stehouwer, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Capillary+recruitment+is+impaired+in+essential+hypertension+and+relates+to+insulin%27s+metabolic+and+vascular+actions%5BTitle%5D)+AND+%22Cardiovasc.+Res%22%5BJournal%5D)">Capillary recruitment is impaired in essential hypertension and relates to insulin's metabolic and vascular actions.</a> Cardiovasc. Res. 49, 161-168 (2001).</li>
<li>Bollinger, A., Fagrell, B. <a target="_blank" href="http://scholar.google.com/scholar?hl=en&safe=off&q=author%3aBollinger%2C+A+%22Clinical+Capillaroscopy%3A+A+guide+to+its+use+in+clinical+research+and+practice%22">Clinical Capillaroscopy: A guide to its use in clinical research and practice</a>. , Hogrefe & Huber Publishers. Lewiston, NY. (1990).</li>
<li>Antonios, T. F., Rattray, F. E., Singer, D. R., Markandu, N. D., Mortimer, P. S., MacGregor, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Maximization+of+skin+capillaries+during+intravital+video-microscopy+in+essential+hypertension%3A+comparison+between+venous+congestion%2C+reactive+hyperaemia+and+core+heat+load+tests%5BTitle%5D)+AND+%22Clin.+Sci.+%28Lond%29%22%5BJournal%5D)">Maximization of skin capillaries during intravital video-microscopy in essential hypertension: comparison between venous congestion, reactive hyperaemia and core heat load tests.</a> Clin. Sci. (Lond). 97, 523-528 (1999).</li>
<li>Sivertsson, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+hemodynamic+importance+of+structural+vascular+changes+in+essential+hypertension%5BTitle%5D)+AND+%22Acta.+Physiol.+Scand.+Suppl%22%5BJournal%5D)">The hemodynamic importance of structural vascular changes in essential hypertension.</a> Acta. Physiol. Scand. Suppl. 343, 1-56 (1970).</li>
<li>Raitakari, O. T., Celermajer, D. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Testing+for+endothelial+dysfunction%5BTitle%5D)+AND+%22Ann.+Med%22%5BJournal%5D)">Testing for endothelial dysfunction.</a> Ann. Med. 32, 293-304 (2000).</li>
<li>Tousoulis, D., Antoniades, C., Stefanadis, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Evaluating+endothelial+function+in+humans%3A+a+guide+to+invasive+and+non-invasive+techniques%5BTitle%5D)+AND+%22Heart%22%5BJournal%5D)">Evaluating endothelial function in humans: a guide to invasive and non-invasive techniques.</a> Heart. 91, 553-558 (2005).</li>
<li>Higashi, Y., Sasaki, S., Nakagawa, K., Matsuura, H., Kajiyama, G., Oshima, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+noninvasive+measurement+of+reactive+hyperemia+that+can+be+used+to+assess+resistance+artery+endothelial+function+in+humans%5BTitle%5D)+AND+%22Am.+J.+Cardiol%22%5BJournal%5D)">A noninvasive measurement of reactive hyperemia that can be used to assess resistance artery endothelial function in humans.</a> Am. J. Cardiol. 87, 121-125 (2001).</li>
<li>Higashi, Y., Yoshizumi, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((New+methods+to+evaluate+endothelial+function%3A+method+for+assessingendothelial+function+in+humans+using+a+strain-gauge+plethysmography%3A+nitric+oxide-dependent+and+independent+vasodilation%5BTitle%5D)+AND+%22J.+Pharmacol.+Sci%22%5BJournal%5D)">New methods to evaluate endothelial function: method for assessingendothelial function in humans using a strain-gauge plethysmography: nitric oxide-dependent and independent vasodilation.</a> J. Pharmacol. Sci. 93, 399-404 (2003).</li>
<li>Lind, L., Sarabi, M., Millgard, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Methodological+aspects+of+the+evaluation+of+endothelium+dependent+vasodilatation+in+the+human+forearm%5BTitle%5D)+AND+%22Clin.+Physiol%22%5BJournal%5D)">Methodological aspects of the evaluation of endothelium dependent vasodilatation in the human forearm.</a> Clin. Physiol. 18, 81-87 (1998).</li>
<li>Cheng, C., Diamond, J. J., Falkner, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Functional+capillary+rarefaction+in+mild+blood+pressure+elevation%5BTitle%5D)+AND+%22Clinical+and+Translational+Science%22%5BJournal%5D)">Functional capillary rarefaction in mild blood pressure elevation.</a> Clinical and Translational Science. 1, 75-79 (2008).</li>
<li>Cheng, C., Daskalakis, C., Falkner, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Capillary+rarefaction+in+treated+and+untreated+hypertensive+subjects%5BTitle%5D)+AND+%22Therapeutic+Advances+in+Cardiovascular+Disease%22%5BJournal%5D)">Capillary rarefaction in treated and untreated hypertensive subjects.</a> Therapeutic Advances in Cardiovascular Disease. 2, 79-88 (2008).</li>
<li>Antonios, T. F., Singer, D. R., Markandu, N. D., Mortimer, P. S., MacGregor, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Rarefaction+of+skin+capillaries+in+borderline+essential+hypertension+suggests+an+early+structural+abnormality%5BTitle%5D)+AND+%22Hypertension%22%5BJournal%5D)">Rarefaction of skin capillaries in borderline essential hypertension suggests an early structural abnormality.</a> Hypertension. 34, 655-658 (1999).</li>
<li>Noon, J. P., Walker, B. R., Webb, D. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Impaired+microvascular+dilatation+and+capillary+rarefaction+in+young+adults+with+a+predisposition+to+high+blood+pressure%5BTitle%5D)+AND+%22J.+Clin.+Invest%22%5BJournal%5D)">Impaired microvascular dilatation and capillary rarefaction in young adults with a predisposition to high blood pressure.</a> J. Clin. Invest. 99, 1873-1879 (1997).</li>
<li>Antonios, T. F., Rattray, F. M., Singer, D. R., Markandu, N. D., Mortimer, P. S., MacGregor, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Rarefaction+of+skin+capillaries+in+normotensive+offspring+of+individuals+with+essential+hypertension%5BTitle%5D)+AND+%22Heart%22%5BJournal%5D)">Rarefaction of skin capillaries in normotensive offspring of individuals with essential hypertension.</a> Heart. 89, 175-178 (2003).</li>
<li>de Jongh, R. T., Ijzerman, R. G., Serne, E. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Visceral+and+truncal+subcutaneous+adipose+tissue+are+associated+with+impaired+capillary+recruitment+in+healthy+individuals%5BTitle%5D)+AND+%22J.+Clin.+Endocrinol.+Metab%22%5BJournal%5D)">Visceral and truncal subcutaneous adipose tissue are associated with impaired capillary recruitment in healthy individuals.</a> J. Clin. Endocrinol. Metab. 91, 5100-5106 (2006).</li>
<li>de Jongh, R. T., Serne, E. H., Ijzerman, R. G., de Vries, G., Stehouwer, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Free+fatty+acid+levels+modulate+microvascular+function%3A+relevance+for+obesity-associated+insulin+resistance%2C+hypertension%2C+and+microangiopathy%5BTitle%5D)+AND+%22Diabetes%22%5BJournal%5D)">Free fatty acid levels modulate microvascular function: relevance for obesity-associated insulin resistance, hypertension, and microangiopathy.</a> Diabetes. 53, 2873-2882 (2004).</li>
<li>de Jongh, R. T., Serne, E. H., RG, I. J., Stehouwer, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Microvascular+function%3A+a+potential+link+betweensalt+sensitivity%2C+insulin+resistance+and+hypertension%5BTitle%5D)+AND+%22J.+Hypertens%22%5BJournal%5D)">Microvascular function: a potential link betweensalt sensitivity, insulin resistance and hypertension.</a> J. Hypertens. 25, 1887-1893 (2007).</li>
<li>Ijzerman, R. G., Voordouw, J. J., Van Weissenbruch, M. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((TNF-alpha+levels+are+associated+withskin+capillary+recruitment+in+humans%3A+a+potential+explanation+for+the+relationship+between+TNF-alpha+and+insulin+resistance%5BTitle%5D)+AND+%22Clin.+Sci.+%28Lond%29%22%5BJournal%5D)">TNF-alpha levels are associated withskin capillary recruitment in humans: a potential explanation for the relationship between TNF-alpha and insulin resistance.</a> Clin. Sci. (Lond). 110, 361-368 (2006).</li>
<li>Yvonne-Tee, G. B., Rasool, A. H., Halim, A. S., Rahman, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Noninvasive+assessment+of+cutaneous+vascular+function+in+vivo+using+capillaroscopy%2C+plethysmography+and+laser-Doppler+instruments%3A+its+strengths+and+weaknesses%5BTitle%5D)+AND+%22Clin.+Hemorheol.+Microcirc%22%5BJournal%5D)">Noninvasive assessment of cutaneous vascular function in vivo using capillaroscopy, plethysmography and laser-Doppler instruments: its strengths and weaknesses.</a> Clin. Hemorheol. Microcirc. 34, 457-473 (2006).</li>
<li>Wilkinson, I. B., Webb, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Venous+occlusion+plethysmography+in+cardiovascular+research%3A+methodology+and+clinical+applications%5BTitle%5D)+AND+%22Br.+J.+Clin.+Pharmacol%22%5BJournal%5D)">Venous occlusion plethysmography in cardiovascular research: methodology and clinical applications.</a> Br. J. Clin. Pharmacol. 52, 631-646 (2001).</li>
<li>Thijssen, D. H., Bleeker, M. W., Smits, P., Hopman, M. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Reproducibility+of+blood+flow+and+post-occlusive+reactive+hyperaemia+as+measured+by+venous+occlusion+plethysmography%5BTitle%5D)+AND+%22Clin.+Sci.+%28Lond%29%22%5BJournal%5D)">Reproducibility of blood flow and post-occlusive reactive hyperaemia as measured by venous occlusion plethysmography.</a> Clin. Sci. (Lond). 108, 151-157 (2005).</li>
<li>Leslie, S. J., Attina, T., Hultsch, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Comparison+of+two+plethysmography+systems+in+assessment+of+forearm+blood+flow%5BTitle%5D)+AND+%22J.+Appl.+Physiol%22%5BJournal%5D)">Comparison of two plethysmography systems in assessment of forearm blood flow.</a> J. Appl. Physiol. 96, 1794-179 (2004).</li>
<li>James, M. A., Tullett, J., Hemsley, A. G., Shore, A. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Effects+of+aging+and+hypertension+on+the+microcirculation%5BTitle%5D)+AND+%22Hypertension%22%5BJournal%5D)">Effects of aging and hypertension on the microcirculation.</a> Hypertension. 47, 968-9674 (2006).</li>
<li>Debbabi, H., Uzan, L., Mourad, J. J., Safar, M., Levy, B. I., Tibirica, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Increased+skin+capillary+density+in+treated+essential+hypertensive+patients%5BTitle%5D)+AND+%22Am.+J.+Hypertens%22%5BJournal%5D)">Increased skin capillary density in treated essential hypertensive patients.</a> Am. J. Hypertens. 19, 477-483 (2006).</li>
<li>Serne, E. H., Gans, R. O., ter Maaten, J. C., Tangelder, G. J., Donker, A. J., Stehouwer, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Impaired+skincapillary+recruitment+in+essential+hypertension+is+caused+by+both+functional+and+structural+capillary+rarefaction%5BTitle%5D)+AND+%22Hypertension%22%5BJournal%5D)">Impaired skincapillary recruitment in essential hypertension is caused by both functional and structural capillary rarefaction.</a> Hypertension. 38, 238-242 (2001).</li>
<li>Carpentier, P. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((New+techniques+for+clinical+assessment+of+the+peripheral+microcirculation%5BTitle%5D)+AND+%22Drugs%22%5BJournal%5D)">New techniques for clinical assessment of the peripheral microcirculation.</a> Drugs. 58, 17-22 (2001).</li>
<li>Shore, A. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Capillaroscopy+and+the+measurement+of+capillary+pressure%5BTitle%5D)+AND+%22Br.+J.+Clin.+Pharmacol%22%5BJournal%5D)">Capillaroscopy and the measurement of capillary pressure.</a> Br. J. Clin. Pharmacol. 50, 501-513 (2000).</li>
</ol>

The authors have no financial conflicts of interest related to the conduct or publication of this work to disclose.

Capillaroscopy (capillary microscopy) is a non-standard measure of capillary structure. However, currently, there are no standard methods for direct assessment of capillary structure.Furthermore, capillary microscopy has been widely used for the direct evaluation of capillarydensity in a large and growing body of published work 12, 13, 10, 11, 14-18. Additionally, we have validated the capillary microscopy technique by correlating capillaroscopy findings with forearm blood flow, a well-established measure of v...

non-invasive assessment of microvascular and endothelial function

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.

Differences in the appearance of the microvasculature between normotensive and hypertensive individuals are readily apparent by comparing Figures 4 and 5. Figure 4 shows the typical network of straight capillaries in well organized rows in a normotensive individual. In contrast, Figure 5 demonstrates a more disarranged pattern of shrunken, coiled capillaries.
The authors have an ongoing interest in the role of microvascular dy...

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 ...

The overall goal of these procedures is to non-invasively, visualize the microcirculation using capillary and assess endothelial function with plethysmography using capillary. This is accomplished by first inducing postocclusive reactive hyperemia on the subject's arm. Next images are taken to identify all functionally perfused capillaries.Then using plethysmography, a strain gauge is wrapped around the subject's forearm and plethysmography measurements of forearm blood flow are recorded. Ultimately, results can be obtained non-invasively that show capillary density through direct visualization by capillary and endothelial function through post react hyperemia strain gauge plethysmography. The main advantage of this technique over existing methods like video cap and invasive plethysmography, is that it is less expensive and non-invasive.Assisting me in demonstrating the procedure will be nausea. Fache, my research coordinator from my laboratory Nail fold capillaries in the dorsal skin of the third finger are visualized using a stereo microscope that uses 250 watt halogen fiber optic lamp illumination, a 3.2 x objective and is linked to a four megapixel spot. Insight monochrome digital camera and a laptop computer with a spot imaging software to carry out the capillary technique following a minimum 10 hour overnight fast, and 20 minutes of seated rest seat the subject in a temperature controlled room and have them raise their left hand to heart level to limit movement.Have the patient rest their left hand and forearm on a folded blanket at the base of the microscope, and use a second covered blanket to loosely cover the forearm and hand to detect the capillaries perfused at rest. Take a series of baseline photo micrographs over a period of three minutes. Capillary density is defined as the number of capillaries per square millimeter of nail fold skin and is computed as the mean of poor measurements obtained from the four most clearly focused images least distorted by movement.To quantify functionally perfused capillaries, induce postocclusive reactive hyperemia by inflating an occlusion cuff on the upper left arm to 40 millimeters, mercury above systolic pressure for 10 minutes. Release arterial occlusion and immediately begin taking images for one minute to identify all functionally perfused capillaries. Following 10 minutes of rest to quantify maximum capillary density induce venous occlusion by inflating the arm cuff to 60 millimeters of mercury for 60 seconds.After releasing the cuff, immediately begin taking images as before. Endothelial function is assessed before and after postocclusive reactive hyperemia using non-invasive plethysmography measurements of forearm blood flow according to the method of sivertson. Using an automatic blood pressure cuff placed on the opposite arm measures systolic and diastolic blood pressures and heart rate.Next place an upper arm occlusion cuff on the subject's arm and use a sling bandage connected to an adjustable intravenous pole to comfortably suspend the arm at heart level Following 10 minutes of supine rest, have the subject in a seated position loop a mercury and rubber strain gauge, stretched to 10%beyond its resting length around the subject's forearm, five centimeters below the antecubital fossa. Connect the strain gauge to a plasm graft that's connected to a doppler recorder to occlude blood flow to the hand. Place a pediatric cuff around the subject's wrist and inflate to 200 millimeters of mercury.Then inflate the upper arm cuff on the opposite arm to 50 millimeters of mercury. Deflate it for one and a half seconds, and then rapidly reinflate and then deflate it to induce postocclusive hyperemia induced maximal vasodilation or FBF max. Immediately use the strain gauge to take four consecutive measurements in 30 seconds of forearm blood flow.To compute the ratio FBF max over FBF base as an estimate of vasodilation. Divide the mean of the four fbf max values by the mean of the four FB F base values. Calculate forearm vascular resistance at maximal vasodilation or FVR max as the mean arterial pressure or map divided by FBF max.A comparison of the appearance of the microvasculature between normotensive and hypertensive individuals is shown here. The left panel shows the typical network of straight capillaries in well-organized rows in a normal oten individual. In contrast, the right panel reveals a more disarranged pattern of shrunken coiled capillaries.Lower capillary density following reactive hyperemia indicates impaired functional capillary recruitment and therefore functional rarefaction. After watching this video, you should have a good understanding of how to visualize the microcirculation and assess endothelial function non-invasively.

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Learn non-invasive capillaroscopy and plethysmography techniques to visualize microcirculation and assess endothelial function using forearm blood flow measurements. This cost-effective method enables reliable functional capillary recruitment studies.

Non-invasive Assessment of Microvascular and Endothelial Function

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