450095
2012
VMJ17410.1177/1358863X12450095Parkhurst KL et al.Vascular Medicine
Contribution of blood viscosity in the assessment
of flow-mediated dilation and arterial stiffness
Vascular Medicine
17(4) 231­–234
© The Author(s) 2012
Reprints and permission: sagepub.
co.uk/journalsPermissions.nav
DOI: 10.1177/1358863X12450095
vmj.sagepub.com
Kristin L Parkhurst, Hsin-Fu Lin, Allison E DeVan, Jill N Barnes,
Takashi Tarumi and Hirofumi Tanaka
Abstract
Flow-mediated dilation (FMD) is a non-invasive index of endothelial function. In an attempt to standardize FMD for
shear stimulus, shear rate (velocity/diameter), rather than shear stress (viscosity*velocity/diameter), is commonly used
as a surrogate measure, although it is limited by individual differences in blood viscosity. The purpose of this study was
to determine the contribution of whole blood viscosity to FMD and other key measures of vascular function. Blood
viscosity, FMD, carotid artery compliance, and carotid–femoral pulse wave velocity (cfPWV) were measured in 98
apparently healthy adults varying widely in age (18–63 years). Whole blood viscosity was not significantly correlated with
FMD, cfPWV, or carotid artery compliance. Shear rate was a stronger correlate with FMD than shear stress that takes
blood viscosity into account (r = 0.43 vs 0.28). No significant differences were observed between whole blood viscosity
and traditional risk factors for cardiovascular disease. Age was positively correlated with cfPWV (r = 0.65, p < 0.001)
and negatively correlated with FMD (r = –0.24, p < 0.05) and carotid artery compliance (r = –0.45, p < 0.01). Controlling
for viscosity did not reduce the strength of these relations. These results indicate that whole blood viscosity does not
significantly impact measures of vascular function and suggests that the common practice to use shear rate, rather than
shear stress, in the adjustment of FMD is valid.
Keywords
carotid artery compliance; pulse wave velocity; shear rate; shear stress
Introduction
Flow-mediated dilation (FMD) is a non-invasive index of
endothelial function.1 During the measurement, blood
flow is occluded and then reintroduced in a manner sufficient to elicit a reactive hyperemic response in the conduit
artery of interest. The shear stimulus caused by the sudden
reperfusion stimulates the endothelium to release nitric
oxide causing vasodilation. Investigators have proposed
normalizing FMD by the shear stimulus in an effort to
adjust FMD for a given stimulus.2–4 Common practice in
the literature is to estimate shear rate (velocity/diameter)
rather than shear stress (viscosity*velocity/diameter)
because of the more invasive nature of measuring blood
viscosity and/or a lack of specialized equipment to measure viscosity. In these instances, blood viscosity is
assumed constant across subjects. Although this assumption appears reasonable, emerging evidence linking individual differences in viscosity to cardiovascular disease5–8
casts doubt on such an assumption. Currently, it remains
unclear whether such an assumption is valid and the
degree by which viscosity affects FMD.
Accordingly, the primary purpose of this study was to
determine the contribution of whole blood viscosity to a
measure of FMD. Additionally, as the secondary purpose,
we determined the potential contribution of blood viscosity
on other key vascular measures (pulse wave velocity and
arterial compliance).
Methods
Subjects
A total of 98 adults varying in age (18–63 years) were studied. All subjects were non-smokers and free of overt
Cardiovascular Aging Research Laboratory, Department of Kinesiology
and Health Education, The University of Texas at Austin, Austin, TX,
USA
Corresponding author:
Hirofumi Tanaka
Department of Kinesiology and Health Education
The University of Texas at Austin
1 University Station, D3700
Austin, TX 78712
USA
Email: [email protected]
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232
Vascular Medicine 17(4)
cardiovascular disease as assessed by medical history. None
of the subjects was taking cardiovascular-acting or hematological medications. The study was reviewed and approved
by the Institutional Review Board. All subjects provided
their written informed consent prior to participation.
Measurements
Following a 12-hour overnight fast, a blood sample was collected from the antecubital vein by venipuncture. Plasma
concentrations of total, low-density lipoprotein (LDL)-, and
high-density lipoprotein (HDL)-cholesterol, triglycerides,
and glucose were determined by standard enzymatic methods. Brachial blood pressure was measured in triplicate at
rest in a supine position with an automated oscillometric
device (HEM-907XL; OMRON Healthcare, Vernon Hills,
IL, USA). Whole blood viscosity was measured immediately after blood collection at 60 revolutions/minute (representing a shear rate of 212/s) using a micro cone-plate
viscometer (LVT-I; Brookfield Engineering Laboratories,
Stoughton, MA, USA). The coefficient of variation for
whole blood viscosity is 8% in our laboratory.
All vascular measurements below were performed after
fasting and abstaining from caffeine for ≥ 4 hours. Participants
were asked to avoid strenuous physical activity and alcohol
for ≥ 24 hours before measurement. For premenopausal
women, measures of vascular function were performed during the early follicular phase of their menstrual cycle to control for the effects of estrogen.9 Subjects were studied in a
supine position after ≥ 15 minutes of rest in a quiet temperature-controlled (22–24°C) laboratory setting.
Flow-mediated dilation (FMD)
Brachial FMD measurements were performed as previously described.10 A longitudinal image of the brachial
artery was obtained using an ultrasound machine equipped
with a high-resolution (15 MHz) linear-array transducer (iE
33; Phillips, Bothel, WA, USA). A customized transducerholding device secured the transducer in place 5–10 cm
proximal to the antecubital fossa. Following the baseline
recording, the blood flow occlusion cuff placed on the forearm distal to the elbow was inflated to 100 mmHg above
baseline systolic blood pressure for 5 minutes using a rapid
cuff inflator (E20; Hokanson, Bellevue, WA, USA). All
ultrasound-derived images were transferred and analyzed
using image analysis software (Brachial Analyzer; Medical
Imaging Applications, Coralville, IA, USA). FMD was calculated using the following equation: (maximum diameter
– baseline diameter) / baseline diameter × 100%. Shear rate
was obtained using the equation: blood velocity/brachial
artery diameter. Shear stress was calculated as whole blood
viscosity*blood velocity/brachial artery diameter.
Carotid–femoral pulse wave velocity
(cfPWV)
The cfPWV was determined using an automated previously
validated device (VP-2000; Colin Medical Instruments,
San Antonio, TX, USA), as previously described.11,12
Carotid artery compliance
Carotid artery compliance was measured in a subgroup of
40 subjects with a combination of ultrasound imaging of
the common carotid artery (by B-mode ultrasound) and
recording of contralateral carotid arterial pressure (by
applanation tonometry; VP-2000; Colin Medical
Instruments), as previously described.13,14
Statistical analyses
Pearson product-moment correlation analyses were used to
determine relations among whole blood viscosity and cardiovascular risk factors and measures of vascular function.
Partial correlation analyses were used to control for the
effect of whole blood viscosity on measures of vascular
function. Because both shear rate and shear stress were not
normally distributed based on the Shapiro–Wilk test, the
data were transformed using either the log scale or square
root, and Spearman correlation analyses were then used to
evaluate associations. Significance was set a priori at p <
0.05. All data are expressed as mean ± SEM.
Results
Selected subject characteristics are displayed in Table 1.
Whole blood viscosity displayed a wide range of distribution from 1 to 5.5 cP60. No significant associations were
observed between whole blood viscosity and traditional
risk factors for cardiovascular disease (Pearson-r ranging
from 0.01 to –0.31). Whole blood viscosity was not significantly correlated with FMD (r = –0.11; Figure 1) or any
other measures of vascular function (cfPWV and carotid
artery compliance). Shear rate was found to be a stronger
correlate with FMD than shear stress (r = 0.43 and 0.28;
Figure 2). The results were essentially the same when both
shear rate and shear stress data were normalized and
Spearman correlation analyses were performed. Spearman
correlation coefficients were 0.41 and 0.20. As expected,
age was negatively correlated with FMD (r = –0.24,
Table 1. Selected subject characteristics
Variable
Mean ± SEM
n
Male / female
Age, years
Height, cm
Body mass, kg
Body mass index, kg/m2
Systolic blood pressure, mmHg
Diastolic blood pressure, mmHg
Flow-mediated dilation, %
Carotid compliance, mm2/mmHga
Blood viscosity, cP
Carotid–femoral PWV, cm/s
98
52/46
34 ± 1
168 ± 1
68 ± 1
24.5 ± 0.4
114 ± 1
67 ± 1
6.1 ± 0.3
0.16 ± 0.01
3.2 ± 0.1
881 ± 17
an = 40.
PWV, pulse wave velocity.
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233
FMD (%)
Parkhurst KL et al.
16
14
12
10
8
6
4
2
0
-2
0.0
1.0
2.0
3.0
4.0
5.0
6.0
Whole Blood Viscosity (cP60)
FMD (%)
Figure 1. Association between flow-mediated dilation (FMD)
and whole blood viscosity.
16
14
12
10
8
6
4
2
0
-2
r = 0.28
p<0.05
-2
3
8
13
18
FMD (%)
Shear Stress (dynes m-1sec-1)
16
14
12
10
8
6
4
2
0
-2
r = 0.43
p<0.001
0.0
0.5
1.0
1.5
2.0
2.5
Shear Rate (sec-1)
Figure 2. Associations between flow-mediated dilation (FMD)
and shear stress and shear rate.
p < 0.05) and carotid artery compliance (r = –0.45,
p < 0.01), and positively correlated with cfPWV (r = 0.65,
p < 0.001). Accounting for blood viscosity did not reduce
the strength of the associations as these relations remained
statistically significant (FMD, r = –0.24; cfPWV, r = 0.65;
carotid artery compliance, r = –0.44).
Discussion
The purpose of this study was to determine the impact of
whole blood viscosity on measures of vascular function.
We found no significant correlations between whole blood
viscosity and FMD or other key measures of vascular function (cfPWV and carotid artery compliance). Shear rate was
a stronger correlate with FMD compared with shear stress
in the present study. It appears that the inclusion of blood
viscosity, which has its own inherent variability, may have
introduced greater variability and reduced the association
of shear stress with FMD. The present findings indicate that
blood viscosity may not be a necessary component when
adjusting FMD for shear stimulus.
It is well established that FMD and carotid artery compliance decreases and cfPWV increases with advancing
age.14–17 Consistent with the previous studies, measures of
vascular function were significantly associated with age in
the present study. To confirm the impact of blood viscosity
on these measures, we performed partial correlation analyses controlling for blood viscosity. The relation between
vascular function and age remained statistically significant
even after the individual differences in whole blood viscosity were incorporated. These findings suggest that blood
viscosity does not appear to modulate the relations between
vascular function and age.
Whole blood viscosity is determined by hematocrit,
plasma viscosity, aggregation of red blood cells, and
deformability of red blood cells.18 It was not until the 1960s
that the ability to measure blood viscosity drastically
improved and became considered a component of blood
flow. Since then, a number of studies have focused on the
importance of this measure with regard to both age and disease status.6,18–22 In our sample of apparently healthy adults,
blood viscosity was not significantly correlated with any of
the risk factors for cardiovascular disease, including age.
While some studies have found a positive correlation
between blood viscosity and certain cardiovascular disease
risk factors,5–8,22 others have not.18,21 Similarly, the relation
between aging and blood viscosity is highly debated within
the literature. Some studies have found an increase in blood
viscosity with age,20,23,24 whereas others, like the present
study, have failed to discover similar findings.25,26 Thus, the
association between blood viscosity and risk factors for
cardiovascular disease remains controversial.
Limitations to the present study include the lack of diseased populations. All of the subjects studied were considered apparently healthy without any overt disease. This
would diminish the generalizability of the present study
findings to other populations. However, the subject selection is also a strength of the present study since the influence of disease states and the use of medications that
potentially confound the data interpretation are minimized
by this approach. Despite the choice of subject selection,
we were able to achieve a wide range of whole blood viscosity values in our sample ranging from 1 to 5.5 cP60. To
the best of our knowledge, this is the first study to determine the impact of blood viscosity on FMD and arterial
stiffness in a healthy population. Previous research in
patients with chronic anemia has also found no correlation
between blood viscosity and FMD.27
In summary, whole blood viscosity was not associated
with FMD and other vascular functional measures. Shear
rate was more strongly correlated with FMD than shear
stress. Taken together, these results are not consistent with
the assertion that measurements of whole blood viscosity
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234
Vascular Medicine 17(4)
should be included when evaluating vascular function. The
next step in this line of investigation is to assess what level
of blood viscosity is necessary in order to register an influence on FMD.
Funding
This research received no specific grant from any funding agency
in the public, commercial, or not-for-profit sectors.
Conflict of interest
None declared.
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