# Characterization of Arterial Wave Reflection in Healthy Bonnet Macaques:  Feasibility  of Applanation Tonometry

Authors: Jason Lazar; Ghazanfar Qureshi; Haroon Kamran; Leonard A. Rosenblum; John G. Kral; Louis Salciccioli
Journal: Journal of Biomedicine and Biotechnology
 (2009)

---

## Abstract

Nonhuman primates are commonly used in cardiovascular  research. Increased arterial stiffness is a
marker of subclinical atherosclerosis and  higher CV risk. We determined the augmentation index (AI) using applanation tonometry
in 61  healthy monkeys (59% female, age 1–25 years). Technically adequate studies were obtained in all subjects and
required1.5±1.3 minutes. The brachial artery provided the highest yield (95%). AI was correlated with heart rate (HR) (r=−0.65,P<.001), crown rump length (CRL) (r = 0.42, P = .001), and left ventricular (LV) mass determined using
echocardiography (r=0.52,P<.001). On multivariate analysis, HR (P<.001) and CRL (P = .005) were independent predictors of AI (R2=0.46,P<.001). Body Mass Index (BMI) and AI were independent predictors of higher LV mass on multivariate analysis (P<.001 and
P=.03). In conclusion, applanation tonometry is feasible for determining AI. Reference values are
provided for AI in bonnet macaques, in whom higher AI is related to HR and CRL, and in turn contributes to higher LV mass.


---

## Body

## 1. Background

Nonhuman primates are commonly used in biomedical research. Macaques and
baboons are the most widely studied species [1–8].
Macaques are anatomically similar to humans and exhibit similar cardiovascular
(CV) physiology and metabolism [9–16]. Increased arterial stiffness
is a marker of subclinical atherosclerosis and a predictor of higher CV risk.
Higher stiffness also contributes to isolated systolic hypertension [17]. Human studies have shown an association
between increased arterial stiffness and left ventricular hypertrophy, carotid, and coronary artery
disease [18–20]. Recent
advances in applanation tonometry have enhanced assessment of arterial
stiffness/wave reflection and have lead to its frequent use in human
cardiovascular research [17, 18].
However, there are few data using these techniques in nonhuman primates
as only one prior study has utilized applanation tonometry in 6 rhesus monkeys [6].
The objective of this study was to determine the feasibility of performing
applanation tonometry in apparently healthy bonnet macaques. Bonnet macaques (Macaca radiata) are
similar to rhesus in size and appearance and have anecdotally been observed to
develop cardiovascular events.


## 2. Methods

The characteristics
of the SUNY Downstate Medical Center primate colony have been described
previously [16]. In summary, there are 250 laboratory-born and raised
bonnet macaques living in social groups of 6 to 10 and maintained on commercial
monkey chow. The SUNY Downstate Medical Center Division of Laboratory Animal
Research approved this prospective study. All procedures were performed in
careful conformance with the Guide for the Care and Use of Laboratory Animals
published by the National Institutes of Health. We studied 61 bonnet macaques
without recent or ongoing participation in physiological or pharmacological
studies.


## 3. Laboratory Methods

Anesthesia was administered using
ketamine (15 mg/kg) via intramuscular injection as clinically indicated to
achieve sedation throughout the procedure. Immediately after sedation, each
monkey was weighed, crown rump length (CRL) was measured, and blood pressures
were recorded by sphygmomanometry of the right lower extremity. Monkey body
mass index (BMI) was calculated by dividing weight in kilograms by
CRL2 in
meters [21]. Monkey body surface area (BSA) was calculated using the
formula of Liu and Higbee [22].We used a pulse wave analysis
system (Sphygmocor applanation tonometer interfaced with SphygmoCor software,
version 8.0 (AtCor Medical, New South Wales, Australia)). The tonometry
transducer was applied to the brachial artery in all subjects and to the
axillary (AxA) and carotid artery (CA) in some of the monkeys. After BA and AxA,
arterial waveforms were recorded and the central aortic pressure waveform and
augmentation index (AI) were derived from the pressure waveform by means of a
generalized transfer function, which has been validated in humans [18].
As performed in humans, carotid waveforms were not transformed. AI was defined
as the proportional increase in systolic pressure due to the reflected wave and
was expressed as a percentage of the pulse pressure (PP) [17].
Arterial waveforms were considered technically adequate if all of the following
criteria were met: large amplitude and consistency of pulse height, diastolic
decay, and morphology maintained for 10 seconds. Repeat BA studies were
performed in 8 randomly selected monkeys.Two experienced echocardiographers
performed transthoracic echocardiographic studies (JL, LS). Each study was
carefully inspected to assure adequate endocardial definition. Left ventricular
(LV) dimensions were measured from M-mode images according to American Society
of Echocardiography standards [23]. Two dimensional images were used
when the scanning axis was not perpendicular to the axis of the heart. LV mass
and ejection fraction (EF) were calculated by the American Society of
Echocardiography-corrected cube formula and indexed by CRL [23].


## 4. Statistics

Data were expressed as mean ±
standard deviation. Univariate associations between variables were analyzed by
using Spearman’s correlation coefficients. Partial correlation controlling for
age was used to assess the relationship between LV mass and AI. Multiple linear
regression analysis was performed to determine independent predictors of AI and
of LV mass. 95% CI were calculated for indices. Repeatability was expressed as
the percentage of the coefficient of variation (SD of the paired
differences/the overall mean)/√2 × 100. All statistical analyses were achieved
using the Statistical Package for Social Sciences (SPSS) 15.0 software (SPSS
Inc., Chicago, Ill, USA).  A P
<.05 was considered to be statistically significant.


## 5. Results

Clinical
and echocardiographic values including mean values, 95% confidence intervals
(CIs), and minimum to maximum ranges are reported for all 61 macaques in Table1. There were 61 macaques, 36 females, and 25 males with mean age 10 ± 5 years.
Mean BMI was 33.2±11.8 kg/m2 and CRL was 0.46±0.05 meters. Mean HR
was 167 ± 33 beats/min. Technically adequate arterial waveforms were obtained
from the BA in 58 of 61 (95%), from the AxA in 37 of 50 (74%), and from the CA
in 18 of 36 (50%) monkeys tested. Brachial artery waveform recording required
1.5  ±  0.3 minutes of interrogation time. Figure 1 shows an example of a BA
recording from a 9.5 kg male. Mean
AI values were 8.2 ±  18.2%  at the BA, 9.9 ±  20.0% at the AxA, and 6.1 ±  20.0%
at the CA, P=.21. The
coefficient of variation of AlBA  was 4.0%.Table 1
Clinical and echocardiographic measurements
in 61 bonnet macaques.
Mean ± SD95% CIRangeAge (years)10.3±5.29.0, 11.71.0–25.0CRL (meters)0.46±0.050.44, 0.470.30–0.57Weight (kg)7.5±3.96.5, 8.51.4–19.7Body mass index
(kg/m2)33.2±11.830.1, 36.214.5–70.0Body surface area0.26±0.070.24, 0.280.10–0.45Heart rate (beats/min)167 ± 33158, 17597–244Septal WT (cm)0.39±0.070.37, 0.410.26–0.54Posterior WT (cm)0.38±0.060.36, 0.400.27–0.51LVEDD (cm)1.76±0.291.67, 1.851.1–2.4LVESD (cm)0.89±0.260.82, 0.970.49–1.9Relative wall thickness0.45±0.080.42, 0.470.28–0.74EF (%)72 ± 1369, 7630–92LV mass (gm)10.2±3.99.0, 11.44.4–20.6LVMI (LV mass/BSA)37 ± 934.3, 39.617.5–56.4LVMI (LV mass/CRL)21 ± 6.719.6, 23.78.8–39.5Systolic BP111 ± 17102, 12182–142Diastolic BP (mmHg)68 ± 1161, 7444–91Pulse pressure (mmHg)43 ± 739, 4833–56AI-brachial artery (%)8.2±18.23.4, 13−35–43AI-axillary artery (%)9.9 ± 203.3, 17−41–41AI-carotid artery (%)6.1 ± 21−4.4, 16−55–36Figure 1
Pulse wave recording of a 13-year-old male bonnet macaque weighing 9.5 kg.Brachial artery AI was correlated
with HR (r=−0.65,  P<.001), weight (r=0.36, P=.006), and CRL
(r = 0.42, P = .001). AI was not correlated with age (r = 0.16, P = 0.21).
AI was higher in males (16 ± 19 versus 2.6 ± 15%, P = .004). Males were
similar in age (10.1±5.1 versus 10.50±5.3 years, P = .98),
but were greater in weight (9.9±5.0 versus 5.7±1.5 kg, P<.001),
BMI (39.4±16.0 versus 28.8±6.2, P<.001), and CRL (.48 ± .07
versus .44 ± .031, P = .002).
Males had lower HRs (142 ± 27 versus 183 ± 25 beats/min, P<.001).AI was also correlated with the
following echocardiographic indices: LVEDD (r = 0.56, P<.001), PWT
(r = 0.35, P = .02), LVESD (r = 0.60, P<.001), LV mass (r = 0.52, P<.001)
(Figure 2), and LVMI (CRL) (r = 0.41, P = .007).  LV mass was significantly correlated with age
(r = 0.44, P<.001).  Upon partial
correlation adjusting for age, AI remained significantly correlated with LV
mass (r =  0.45, P = .003) (Figure 2) and with LVMI (CRL) (r = 0.36, P = .01).
On multivariate analysis, using age, HR, systolic BP, gender and CRL, and
weight as independent variables, HR (P<.001) and CRL (P = .005)
were independent predictors of AI with a trend toward age (P = .10)
(R2 = 0.46, P<.001). On multiple linear regression using age, gender,
BMI, systolic BP, and AI as independent variables, BMI (P<.001) and
AI (P =  .03) were independent predictors of LV mass (R2 = 0.46, P<.001).Figure 2
Relation between AI and LV mass.


## 6. Discussion

The present study determined the
feasibility of applanation tonometry for the noninvasive recording of arterial
waveforms and deriving AI from a large cohort of adult bonnet macaques.  The subjects represent a wide range of ages
and weights in animals of both sexes with a mean HR of 167 beats/min, thereby
reflecting differences in access to arteries in relatively small animals.
Interpretable waveforms were obtained from the BA in 95% of cases, from the AxA
in 80%, and from the CA in 50% of macaques studied.The BA provided the highest yield
likely because of its close proximity to the surface of the skin and its
location superficial to humerus bone. This allows the vessel to be flattened
with the transmural arterial forces perpendicular to the tonometry probe. In
humans, the radial artery is easily accessible and is supported by bony tissue;
however, it was too small to evaluate pulse waves in macaques. These findings
are likely applicable to rhesus monkeys, which are the most commonly studied
macaques and are similar in size to bonnets. In support of this, LV dimensions
obtained by echocardiography were similar to those previously reported for rhesus
monkeys [7]. The AxA, which is located several cm from the BA,
appeared to be a suitable alternative in the majority of animals. We were able
to adequately record waveforms from the CA in only half of the animals probably
because of their depth
and overlying redundant skinfold.
Although the femoral artery was not interrogated in this study, our
group has been successful in acquiring femoral artery waveforms using this
technique in other bonnets, vervet, and cynomologous monkeys.Indices of arterial wave reflection
derived by these techniques are frequently used in human cardiovascular research,
but have not been well studied in primates [17]. Although several
studies have evaluated arterial stiffness in primates [24, 25], only
one prior study utilized applanation tonometry to assess arterial stiffness in
monkeys [6]. Vaitkevicius measured the AI in 6 older Rhesus monkeys in order
to determine the effects of a synthesized thiazolium compound on arterial
properties in old, healthy Macaca mulatta primates. This prior study
underscores the value and utility of a noninvasive measure of cardiovascular
function. Of note, AI values obtained in the present study are similar to those
obtained without a transfer function from the carotid artery in the prior study
(6.1–8.1% versus 8.8%)
which used telazol anesthesia [6].The AI is a measure of the magnitude
of arterial wave reflection, which is
highly correlated well with invasive and noninvasive measures of arterial
stiffness in humans [17]. In the present study, AI was easily
determined in the vast majority of macaques and was related to HR, CRL, and possibly
age. Age and HR dependency of AI has
been reported in humans [26]. AI was directly related to CRL. This
finding is seemingly contradictory to human data in which AI has been found to
be inversely correlated with height [27]. The reason for this discrepancy is not known, but may relate to
the CRL being used as a measure of length in macaques or to the possibility
that the relation between AI and length differs between anthropoids of shorter
and taller stature. The relation between LV mass and AI is also
consistent with prior human data [28]. In humans, women exhibit
higher AI values than men, even after adjustment for shorter stature [27].  Although males had higher AI than female
bonnets on univariate analysis, gender was not an independent predictor of AI
on multivariate analysis.The strong dependency of AI on HR
was observed at HRs higher than typically observed in humans. These findings
extend the findings of prior studies showing the AI to vary with HRs between 60
and 110 beats/min [26]. The observed slope of the regression line
between AI and HR is quite similar to that reported in humans (−.33 versus
−.39) [26]. In humans, correction of AI to an arbitrary value of  75 beats/min (AI75) has been proposed and reported extensively [27].
The device automatically calculates the AI75 up to an HR of
110 beats/min, the upper level of HR previously studied. Mean HR of the macaques
in the present study was 100 beats/min higher than in humans in whom AI75 was first proposed (167 versus 65 beats/min). Accordingly, correction of AI
to 175 beats/min may be a reasonable convention for use in primates. Correction
of AI to 175 beats/min resulted in mean values of AI175 of BA
4.8 ± 15%, AxA 6.3 ± 17%, and CA 2.9 ± 16%. Although a rapid HR could be expected to
complicate the assessment, reflective wave characteristics were measurable in
the vast majority of subjects. The tonometry device records data points at a
sampling rate of 128 cycles/s and can easily record arterial waveforms
occurring at a rate of 200 beats/min. Higher HRs would limit the sampling to
fewer than 40 points per cardiac cycle.


## 7. Limitations

This is a cross-sectional study of
apparently healthy bonnet
macaques. All measurements were obtained during sedation with ketamine.
Although anesthetic agents are known to influence LV performance, ketamine has
been shown to have minimal effects on cardiac contractility and HR [29].
Mean HR of our macaques was similar to that observed in other monkey
experiments.  Similar LV wall thicknesses
have been obtained in rhesus monkeys sedated with ketamine and with a
combination of Telazol and Isoflurane [7]. LV mass was based on
measurements from the parasternal view, which was easily obtained in all of the
animals. Although pulse wave velocity may also be measured by this technique,
it was not evaluated in the present study. Similarly, other indices derived
from applanation tonometry reflecting cardiac workload were not assessed.  The AI was derived using a general transfer
function, which has been validated in humans but not yet in nonhuman primates.
This merits further study.In conclusion, this study
demonstrates the feasibility and supports the applicability of applanation
tonometry for characterizing arterial wave reflection in bonnet macaques. It
provides preliminary reference values of AI in a group of macaques, which may
be useful for identification of cardiovascular abnormalities in similar
primates. The success of applanation tonometry performed at various arterial
sites might differ slightly among colony breeds.  Validation by comparison of calculated
indices to those obtained invasively merits further study.



---

*Source: 876093-2009-03-10.xml*