Journal of Atherosclerosis and Thrombosis Vol.16, No. 5

662

Original Article

Evaluation of Plaque Stabilization by Fluvastatin with Carotid IntimaMedial Elasticity Measured by a Transcutaneous Ultrasonic-BasedTissue Characterization System
Toshio Yamagishi 1, Makoto Kato 2, Yoshiro Koiwa 3, Ken Omata 4, Hideyuki Hasegawa 5, 6, and Hiroshi Kanai 6, 5
1

Department of Internal Medicine, Tohoku Kosai Hospital, Sendai, Japan
Panasonic Shikoku Electronics Co., Ltd, Yokohama, Japan
3
Oizumi Memorial Hospital, Shiroishi, Japan
4
Health Administration Center, Miyagi University of Education, Sendai, Japan
5
Department of Biomedical Engineering, Graduate School of Biomedical Engineering, Tohoku University, Sendai, Japan
6
Department of Electronic Engineering, Graduate School of Engineering, Tohoku University, Sendai, Japan
2

Aim: As an approach to tissue characterization, we attempted to classify in vivo carotid plaque tissues
in terms of arterial wall elasticity instead of echogenicity on B-mode scanning and investigatedwhether the effect of fluvastatin on carotid elasticity can be detected in hypercholesterolemic patients.
Methods: In 170 subjects, simultaneous measurements of intima-media thickness (IMT) and elastic
modulus in the circumferential direction (Eθ) were performed using a new transcutaneous high-resolution Doppler technique.
Results: From the observation of various tissues, an elasticity library was obtained as follows: lipid
core, 22±15 kPa; calcification, 674±384 kPa; lipid core- and calcification-free plaques, 173±69 kPa;smooth muscle, 104±32 kPa; blood clot, 85±68 kPa; fibrosis, 273±173 kPa.
The effect of fluvastatin (30 mg/day, n = 62) was assessed over 12 months using the elasticity distribution and serum markers. The statin reduced low-density lipoprotein cholesterol, high-sensitivity
CRP, mean IMT and mean Eθ, and increased nitrite/nitrate. In the max IMT ≥ 1.1 mm group, bothEθ and IMT decreased significantly. On the other hand, in the max IMT ＜ 1.1 mm group, Eθ but
not IMT decreased significantly. The histogram of the subgroups showing increased Eθ with max
IMT ≥ 1.1 mm revealed a decrease in areas corresponding to Eθ of 20−200 kPa (lipid/smooth muscle-rich tissue) and an increase in relatively hardened areas of ＜ 250 kPa (collagen fibers).
Conclusion: Non-invasive echographic carotid arterial elasticity measurement is useful for the classification of atherosclerotic plaques and evaluation of chronological and histopathological changes.
J Atheroscler Thromb, 2009; 16:662-673.
Key words; Elasticity, Statin, Echo, Carotid artery

Introduction
Ultrasonography of the carotid arteries is a common imaging study performed for the diagnosis ofatherosclerosis, not only in carotid but also in systemicarteries. Conventional evaluations are based on the
Address for correspondence: Toshio Yamagishi, Department ofInternal Medicine, Tohoku Kosai Hospital, 2-3-11, Kokubuncho, Aoba-ku, Sendai 980-0803, Japan
E-mail: tyamagishi@tohokukosai.com
Received: January 8, 2009
Accepted for publication: April 16, 2009

measurement of carotid arterial intima-media thickness (IMT) to detect morphological changes andremodeling, such as thickening and calcification 1-5)
and on integrated backscatter (IBS) analysis using
echogenicity on B-mode scanning for the soft/hardcharacterization of plaque composition 6-8); however,these procedures provide no information on themechanical stability of plaque against rupture and fortissue characterization. Recently, a new ultrasonic system that can measure regional elasticity in the arterial
wall utilizing the phased tracking method has beendeveloped 9-15). Measurements of the elasticity distribu-

Carotid Plaque Stabilization by Statin

Table 1. Characteristics of the study population
Healthy
volunteers in
20s−40s

Patients withhyperlipidemia

N
46
72
age
35.4±2.760.4±7.6
mean IMT (mm)0.54±0.08
0.88±0.25
max IMT (mm)0.68±0.08
1.15±0.20
mean Eθ (kPa)118.8±26.8
158.5±62.0
SBP (mmHg)
111.2±7.8135.2±13.8
DBP (mmHg)
70.1±7.481.3±8.5
PP (mmHg)
41.1±9.0
54.0±11.7
PR (bpm)70.8±8.971.3±9.6
TC (mg/dL)
177.6±45.5
255.4±32.8
TG (mg/dL)
119.2±70.7
141.9±96.1
HDL-C (mg/dL)
50.7±10.3
59.8±15.9
LDL-C (mg/dL)
98.1±29.8
154.7±21.8
FPG (mg/dL)
87.0±9.3
98.8±13.1
HbA1c (%)
4.7±0.5
5.2±0.7
baPWV (cm/s)
1,106.4±220.4 1,592.0±306.9

663

Table 2. Characteristics of patients with lipid core or calcification in carotid arteries
p-value＜ 0.0001＜ 0.0001＜ 0.0001

0.0006＜ 0.0001＜ 0.0001

0.0001NS＜ 0.0001
NSNS＜ 0.0001
0.01280.0222＜ 0.0001

Nagemean IMT (mm)max IMT (mm)mean Eθ (kPa)
SBP (mmHg)
DBP (mmHg)
PP (mmHg)
PR (bpm)
TC (mg/dL)
TG (mg/dL)
HDL-C (mg/dL)
LDL-C (mg/dL)
FPG (mg/dL)
HbA1c (%)
baPWV (cm/s)

Patients withlipid core

Patients withcalcification

1856.8±9.1
0.91±0.16
1.21±0.20
145.7±40.6
129.8±20.0
79.8±14.1
50.0±13.5
71.5±12.8
230.4±36.8148.5±82.3
52.6±13.9
140.5±37.5116.9±45.5
5.5±1.3
1,421.1±144.4

3361.8±8.90.93±0.17
1.26±0.27
193.7±55.4
140.1±17.7
81.2±9.958.9±12.672.3±10.6
228.6±34.5
117.5±52.7
56.4±13.4
137.0±30.1
104.1±24.6
5.2±0.6
1,644.4±306.6

SBP: systolic blood pressure, DBP: diastolic blood pressure, PP: pulsepressure, PR: pulse rate, TC: total cholesterol, TG: triglyceride,
HDL-C: high density lipoprotein cholesterol, LCL-C: low density
lipoprotein cholesterol, FPG: fasting plasma glucose, baPWV: brachio-ankle pulse velocity wave.

SBP: systolic blood pressure, DBP: diastolic blood pressure, PP: pulsepressure, PR: pulse rate, TC: total cholesterol, TG: triglyceride,
HDL-C: high density lipoprotein cholesterol, LCL-C: low density
lipoprotein cholesterol, FPG: fasting plasma glucose, baPWV: brachio-ankle pulse velocity wave.

tion of lipids, blood clots, fibrous tissue and calcified
tissue in vitro have been reported previously, and anelasticity image can be classified into tissue components using the reference data obtained by in vitro
experiments 16). The usefulness of measuring the elastic
modulus of the common carotid arteries (CCAs) byultrasound imaging to detect early-stage atherosclerotic lesions caused by risk factors with regard to lifestyle-related disease has also been reported 15).
The aim of the present study was to evaluate andclassify in vivo carotid plaque tissues in terms of arterial wall elasticity using this novel real-time ultrasonicmeasurement system rather than conventional methods. Carotid plaque was evaluated to assess atherosclerotic lesions, such as plaques, that showed an increaseof elasticity with progressive lipid deposition andfibrosis resulting from chronological and histopathological changes. In addition, we assessed whether theeffects of fluvastatin on plaque stabilization can bedetected by carotid elasticity distribution in patientswith hypercholesterolemia.

teers, outpatients with untreated hypercholesterolemia,
patients with lipid core or calcification, and a single
case of acute-onset carotid artery dissection, wereenrolled in this study from 2004 to 2006.
The characteristics of the study population,
except the case of carotid artery dissection, are shownin Table 1, 2. Seventy-two subjects with hypercholesterolemia were defined in accordance with the guidelines of the Japanese Society of Atherosclerosis. Hypercholesterolemia was defined as total cholesterol ≥ 220
mg/dL or low-density lipoprotein cholesterol (LDL-C)≥ 140 mg/dL and triglyceride (TG) ≥ 150 mg/dL.
None of the subjects had hypertension, diabetes, a history of recent cardiovascular events, ischemic heartdisease, atrial fibrillation, arteriosclerosis obliterans(ASO, defined as an ankle/brachial blood pressureindex [ABI] ＜ 0.9), renal insufficiency with serum creatinine greater than 3 mg/dL, or severe hepatic disorder. Patients with congestive heart failure were excluded
from this study.
Some patients with lipid core and/or calcificationon B-mode images included untreated patients withhypertension, diabetes mellitus, hypertension being
defined as systolic blood pressure (BP) ≥ 140 mmHg
and/or diastolic BP ≥ 90 mmHg. Diabetes mellitus
was defined as a fasting plasma glucose (FPG) ≥ 126

Methods
Study Population
A total of 170 subjects, including healthy volun-

664

Yamagishi et al .

mg/dL. They did not have any complications.
None of the patients were taking any drugs, suchas inhibitors of the renin-angiotensin system, Ca channel blockers and anti-diabetic agents.
Similar examinations were conducted in 46
healthy subjects between 20 and 49 years of age, who
served as the control group.
In addition, we assessed the effect of fluvastatin(30 mg/day, n = 62) during a period of 12 months,
evaluating IMT, elasticity distribution, brachial-ankle
pulse wave velocity (baPWV) and serum and markers,
including high-sensitivity CRP (hs-CRP) and nitrite/
nitrate (NOx). In the control group (n = 10), no statin
treatment was performed.
The subjects provided informed consent, and thestudy was approved by the Institutional Ethics Committees of Tohoku Kosai Hospital and Matsushita
Electrical Industrial Co., Ltd.
Carotid IMT and Elasticity Measurements
The subjects were studied after having abstained
from caffeine, alcohol, and smoking during the previous 12 hours. They rested in a supine position for 15
min in a quiet room before the baseline hemodynamic
measurements were obtained. Brachial blood pressureand heart rate (HR) (mean of three readings) were
measured in the right arm with an automated digital
oscillometric sphygmomanometer (EW-3110; Matsushita Electric Works, Osaka, Japan). An imaging studyof the bilateral common carotid arteries was performed
using a newly developed high-resolution ultrasonicmeasurement system (Panasonic, prototype) with acenter frequency of 7.5 MHz. One well-trained sonographer who regularly participates in quality control
measurement sessions performed all ultrasoundscans 13, 15). The sonographer has been trained to obtain
an elastic modulus measurement deviation of only 6%
in the same subject (inter- and intra- variations wereless than 10% at our institution). In order to obtainan accurate measurement of the same sites of carotid
arteries in repeated measurements, the sonographer
performed IMT and elastic modulus measurements bycomparing current B-mode images with past B-mode
images on the PC display and positioned the probe so
as to measure a similar position in the carotid arteries.
IMT can only be determined accurately in the
far wall position because only the far wall IMT isdefined by leading edges, which enables correct ultrasonographic representation; therefore, the far walls of
the following sites were measured: (1) the distal 3 cmof the straight part of both CCAs and (2) the distal 1
cm of both carotid bulbs. As in several previous studies, we defined a plaque as a focal raised lesion, using

an IMT cutoff value of 1.1 mm. Mean IMT and max
IMT were determined to be the mean value of IMT
and the maximum value of IMT, respectively, at six
locations in the bilateral CCAs, including the carotid
bulb.
This system measures the radial strain of the arterial wall during one cardiac cycle and calculates theelasticity from radial strain and pulse pressure. The
distribution of elasticity is displayed as a 2D cross-sectional color-coded image in B-mode imaging, and theimage is updated at every heartbeat. To measure minute changes in thickness during one cardiac cycle, thissystem uses the phased tracking method 5-10). Elasticity
was estimated at intervals of 80 μm along each ultrasonic beam in the depth direction and 400 μm in thearterial longitudinal direction. From the maximum
change in thickness, (hmax−hmin), during one heartbeat, the radial strain εr of each assigned layer in theartery wall was calculated as follows:
εr = (hmax−hmin)/hmax,(1)where hmax and hmin are the maximum and minimum
thickness of an assigned layer in the wall during one
heartbeat, respectively.
The elastic modulus in the radial direction (Er)and that in the circumferential direction (Eθ) were calculated as follows:
Er = Pulse Pressure/εrEθ = (1/2)×(r0/h0＋1)×Er.

(2)
(3)

In these formulae, pulse pressure (PP) is the difference
between systolic and diastolic brachial blood pressure(carotid blood pressure would be more accurate but
is difficult to measure), and h0 and r0 are the initialwall thickness and radius of the vessel in end-diastole,
respectively. The radial strain εr is caused by radial
compression and circumferential stretching due tothe increase of blood pressure Δp, with longitudinalstrain being disregarded. As this circumferential
stretching produces radial strain due to the incompressibility of the arterial wall, both radial and circumferential stresses should be considered when evaluating
elasticity; however, as shown in Eq. (2), Er is definedas the ratio of pulse pressure (which corresponds toradial stress) and radial strain. The circumferential
stress acting on the cylindrical shell is not identical topulse pressure, but also depends on r0/h0. Thus, Er isapparent elasticity in the radial direction and is dependent on r0/h0 as well as on the elasticity of the arterial
wall. Alternatively, Eθ of Eq. (3) was introduced toevaluate the elasticity of the arterial wall by suppressing the influence of r0/h0 7). The elastic modulus ineach instrumentation locus was expressed in a histo-

PWV Measurements
Brachial-ankle PWV (baPWV) was measured
using a volume-plethysmographic apparatus (Form/
ABI; Colin Co., Ltd., Komaki, Aichi, Japan) by simultaneous BP and waveform measurements on all four
limbs, in addition to ECG and phonogram tracings.
PWV was automatically calculated by time-phasic
analysis using the following formula: distance between
two sites divided by pulse wave transit time. Values for
baPWV were measured from the ascending point
shown by the right brachial pulse volume recorder tothe ascending point indicated by each ankle pulse volume recorder. The distance between the right brachium and ankle was estimated as described previously 16).
Blood Examination, NOx and hs-CRP
The patients were requested to fast overnight
without medication until blood samples were drawn.
Venous blood samples were then drawn for analysis
of FPG and serum concentrations of total cholesterol (TC), TG, high-density lipoprotein cholesterol(HDL-C) and HbA1c by standard laboratory methods. Serum LDL-C was calculated by Friedewald’s
equation. None of the patients had a TG ＞ 400 mg/
dL, at which the Friedewald equation is known to yield
unreliable results.
The serum level of NOx (nitrite/nitrate; NO2-/
NO3-), an indicator of NO, was measured by theGriess reaction after nitrate reductase treatment 17) witha flow injection analysis system (Tokyo Chemical
Industry Co., Ltd., Tokyo, Japan). The serum NOxlevel was 10−71 μmol/L and the coefficient of its
variation was 3.0%.
The serum level of hs-CRP was measured by
the BNII High Sensitivity CRP assay using anti-CRP
monoclonal antibody coated with polystyrene particles
and fixed-time kinetic nephelometric measurements(N High Sensitivity CRP; Dadebehring Inc., Marburg, Germany) with a lower detection level of 0.2

mean E� (kPa)mean IMT (mm)gram, and the elasticity distribution of each tissue
composition was determined.
The length and thickness of the ROI (region ofinterest), in which the elasticity distribution and average elasticity were calculated, can be changeable anddepend on the effective size of the interested area,
including IMT. When we obtained the elasticity
library, by referring to B-mode echo images, wechanged the sizes and positions of ROI within plaquesof patients in order to confirm that areas with extreme
differences in brightness and/or echo pattern are notincluded in the region of interest.

Blood pressure(mmHg)

Carotid Plaque Stabilization by Statin

665

Figure 1

120
100
80
60
180

�

150
120
90
60
0.7
0.6
0.5
0.4
0.3

��������

20s

30s

40s

Age

Fig. 1. Effects of aging on IMT and elastic modulus in
healthy volunteers.
IMT increased with greater age, but not with higher blood pressure. Overall, Eθ in healthy volunteers was not affected by aging.

mg/L and a coefficient of variation of 3.0% 18).
Statistical Analysis
Data were expressed as the mean±SD. Statisticalanalysis was performed using a JMP ® software package (JMP 7.0.1; SAS Institute, USA), a value of p ＜0.05 being considered significant. Differences between
groups were analyzed by one-way analysis of variance(ANOVA). A post-hoc test for multiple comparisons
was used when ANOVA showed significance. Linear
regression analysis was performed to evaluate the association between IMT, elasticity or baPWV, and otherclinical variables.
Results
Table 1 shows the characteristics of the study
population of healthy volunteers (n = 46), hypercholesteremia patients (n = 72), patients with a lipid corein carotid artery plaque (n = 18), and patients withcalcification (n = 33). The average blood pressure(systolic/diastolic) was significantly lower in healthy
volunteers than in other subjects (ANOVA, p ＜ 0.0001)
and other risk factors for cardiovascular disease werealso lower in healthy volunteers.
Elasticity Library
The effects of aging on the mean IMT and meanEθ in healthy volunteers in their twenties (n = 15),thirties (n = 12) and forties (n = 19) are shown in
Fig. 1. We were unable to obtain data for healthy volunteers over fifty. Although IMT increased signifi-Yamagishi et al .

666

Figure 2A

B
Frequency

Elasticity modulus(E�)Lipid core 45y.o. male

maxIMT 1.6 mm
minIMT 0.7mm

Lipid core 16 kPa

maxIMT 0.6 mm
minIMT 0.4mm

Frequency

674�384 kPa
Smooth muscle 28y.o. male

Smooth muscle 80 kPa

Calcification 75y.o. male�� �����
1000
500
250
maxIMT 3.2 mm
minIMT 0.7mm

0Calcification 555 kPa

Frequency

B-mode1.0
Lipid core
22�15 kPa

0.5
0.0
0
1.0

200 400 600 800 1000
Smooth muscle
104�32 kPa

0.5
0.0
0
1.0

200 400 600 800 1000
Calcification
674�384 kPa

0.5
0.0

0

500

1000

1500

2000

E� (kPa)

Fig. 2. Elastic modulus of lipid core, smooth muscle and calcification.
The right panel (A) shows the relation between B-mode findings and calculated Eθ. The
value of Eθ was related to tissue composite elements. The left panel (B) shows a histogramfor Eθ (lipid core, smooth muscle and calcification).

cantly by age, there was no significant increase in Eθ
or blood pressure. In healthy volunteers in their thirties and forties, no significant increases in TC,
HDL-C, LDL-C, TG or FPG were found in comparison with levels in healthy volunteers in their twenties(data not shown).
Fig. 2A shows the relationship between B-mode
findings and Eθ in three representative cases. The dark
central region of soft plaques was considered to be thelipid core (16 kPa). The normal carotid intima-media
complex in a 28-year-old man was considered to besmooth muscle (80 kPa). The white regions, whichwere not accompanied by an acoustic shadow, wereconsidered to be calcified regions (555 kPa). All data
are summarized as histograms in Fig. 2B (lipid core,
22±15 kPa (lesions: n = 18); calcification, 674±384kPa (lesions: n = 36)). In the 15 healthy subjects who
were free from vascular risk factors, the normal carotid
intima-media complex was considered to be smoothmuscle (104±32 kPa, sites: n = 71).
Next, in order to obtain the elasticity of bloodclots and fibrosis, the following case was used. A
43-year-old woman came to our department becauseof a pulsatile mass 2.5 cm in size in the right submandibular area. Her blood pressure was 108/80 mmHg,
and ECG showed sinus tachycardia (92/min). Imaging of enhanced CT and ultrasound showed acuteonset carotid artery dissection. The patient had no

cardiovascular risk and no history of stroke, and had
experienced a spontaneous cure from intra-pseudosaccular thrombosis. During B-mode scanning to observe
the healing process at our clinic, with permission fromthe patient and approval from our ethics committee,
elastic modulus was measured several times (Fig. 3).
Elasticity Eθ, as determined from the time course ofchanges, was 85±68 kPa in areas corresponding tothrombi and 273±173 kPa in areas of fibrosis.
From the observation of various tissues, the elasticity library of Eθ by transcutaneous ultrasound was
obtained, as shown in Fig. 4. The data indicate that
the Eθ value rises depending on the tissue constituents. The mean Eθ value of lipid core- and calcification-free plaques was 173±69 kPa (lesions: n = 270),
falling between the Eθ values of fibrosis and bloodclot.
Effect of Fluvastatin On Elastic Modulus and IMT
At baseline, patients with hypercholesterolemia
showed significantly higher LDL-C, baPWV, mean
IMT, max IMT and mean Eθ than seen in healthy
volunteers (Table 1). As shown in Fig. 5, in the control group of hypercholesterolemia (n = 10), no changeof LDL-C was seen. In the group treated with fluvastatin (n = 62), LDL-C was significantly reduced immediately. By fluvastatin treatment, hs-CRP and baPWV
decreased gradually but NO increased over time (p ＜Carotid Plaque Stabilization by Statin

667

Figure 3A

0 weekEnhanced CT

10 week

16 week

1) B-mode �Area of obtaining elastic modulusE� (kPa)
1000
500
250

2) Elasticity modulus of posterior wallA

0Frequency

B

118

202

600

0

400
300
200
100
0
0

FibrosisE� 273+173 kPa

C 100

0 week10 week

500

161

Frequency

Mean E� (kPa)-100

Decrement
Increment

-200
-300

Blood clotE� 85+68 kPa

-400
200

400

600

-500
0

200

E� (kPa)

400

600

E� (kPa)

Fig. 3. Aneurysm of right cervical artery in a 43-year-old woman: time course of changes ofthrombosis in the pseudolumen.
Computed tomography at 0 weeks showed dissection, and the B-mode and elastic modulus of thelesion in the upper panel at 0 week showed relatively soft tissue elements; however, Eθ values roseover time, indicating the disappearance of the blood clot. In the lower panel (B), histograms of Eθ at
0 weeks and 10 weeks are shown.

0.05). No significant changes in blood pressure orpulse rate were seen (data not shown). Fluvastatin
decreased the mean IMT and mean Eθ significantlyin the analysis of all sites of all patients (Fig. 6A,
ANOVA, p ＜ 0.05). Mean Eθ decreased significantly
from 3 to 12 months of administration, while mean
IMT decreased only after 12 months of administration, compared with control values at 0 months. On
the other hand, no change of mean IMT or mean Eθ
between 0 and 12 months was seen in the control
group of hypercholesterolemia (Fig. 6A, n = 10).
Since it has been reported that native Japanesereach an IMT of 1.1 mm at age 70 and subjects withmax IMT ＜ 1.1 mm are often classified as not having
atherosclerosis 5), we divided all analyzed sites (n = 242)
into two groups with a max IMT cut off value of 1.1
mm. In the group with max IMT ≥ 1.1 mm (sites:n = 144), both the mean Eθ and mean IMT decreased
significantly (Fig. 6B, ANOVA). In comparison withthe values at 0 months, the values of mean Eθ at 6 and12 months and mean IMT at 12 months were significantly decreased. In the group with max IMT ＜ 1.1
mm (sites: n = 98), the mean Eθ significantly decreased
and IMT showed a tendency to decrease (Fig. 6C).
Next, we divided all sites into three groups by

Figure 4
Calcification

674�384

Fibrosis273�173

Plaque173�69Blood clot

85�68

Smooth muscle

124�41

Lipid core

22�15

0

200

400

600

800

1000

E� (kPa)

Fig. 4. Elasticity library for Eθ (kPa).
Data of the elasticity library of Eθ by transcutaneous ultrasoundwere obtained from the observation of various tissues: lipid core, 22±15 kPa; calcification, 674±384 kPa; lipid core- and calcificationfree plaques, 173±69 kPa; smooth muscle, 104±32 kPa; bloodclot, 85±68 kPa; fibrosis, 273±173 kPa. The dark central region ofsoft plaques was considered to be the lipid core. In healthy subjects
free from vascular risk factors, we considered the normal carotid
intima-media complex to be smooth muscle. The white regions,which are not accompanied by acoustic shadow, were considered tobe calcification regions. Elastic moduli, equivalent to blood clot andfibrosis, were obtained from the observation of an uncomplicatedhealing process of carotid artery dissection, as shown in Fig. 3.

Yamagishi et al .

668

5FigureFigure
5

0

*

*
3

baPWV (cm/sec)

1800 0

6

3

#

9

1600

2000

1800
1200
3

6
months912p=0.0148

0.9
0.6

*

30.3

6

0.0

12
80
60

100

40
20

0

p=0.0148

*
9

12

*

0

p=0.0195

0

3

6

9

12

�

*

p=0.019580
360 6
months912** p<0.05 vs 0 months#
1400
p<0.05vs Control
Fig. 5. Effectsof fluvastatin on LDL-C,
hs-CRP,
baPWV40
and NOx.
A reduction in LDL-C and decreases in baPWV and hs-CRP were noted, along
1200
with a gradual increase in NOx. No difference of LDL-C 20
was seen between thecontrol (n = 10) and fluvastatin group (n = 62) at 0 months. Fluvastatin significantly
1000
reduced LDL-C in the comparison of the two groups at 12 months. P-values were
0 ANOVA
3 for the6fluvastatin
9 group.12
0 of the3 fluv- 6＊
obtained by
p ＜ 0.05 vs. 0 months
monthsastatin group; ＃p ＜ 0.05 months
vs. 12 months of the control.
* p<0.05 vs 0 months#
p<0.05 vs Control

the mean Eθ values at 0 months (before administration of fluvastatin), Eθ ≤ 100 kPa, 100 kPa ＜ Eθ＜ 200
kPa, and 200 kPa ≤ Eθ. In the three groups, IMT did
not change and showed a tendency to decrease (Fig. 7).
On the other hand, in the group with Eθ ≤ 100 kPa,
the mean Eθ significantly increased from 1 monthafter administration of the drug (Fig. 7A, sites: n = 42).
In the group of 200 ≤ Eθ, the mean Eθ significantly
decreased from 1 month after administration of the
drug (Fig. 7C, sites: n = 46). In the group of 100 kPa＜ Eθ＜ 200 kPa, the mean Eθ significantly decreased(Fig. 7B, sites: n = 154).
Two typical examples are shown in Fig. 8, 9. In
the advanced thick plaque (max IMT ≥ 1.1 mm andEθ ≤ 100 kPa) shown in Fig. 8, fluvastatin decreased
IMT and increased Eθ at month 12, particularly inthe hardened areas in the superficial layer. Histogram
analysis of the data revealed a decrease in areas corresponding to Eθ of 20−200 kPa and an increase in therelatively hardened areas of Eθ ＞ 250 kPa for subgroups showing increased Eθ, thought to represent a
decrease of lipid/smooth muscle-rich tissue and/or
an increase of collagen fibers (based on the elasticity
library and Fig. 8).
In the early lesion (max IMT ＜ 1.1 mm) thatshowed heterogeneous distribution of Eθ (Eθ ≥ 200*�

p=0.0485

1400

1000
1600

0

100

p=0.0485

9

0.3
0.0
*
#

12

6

0.6

NO ��mol/L�

100

*

*

0.9

hsCRP(mg/L)

12080

*

*

NO ��mol/L�

*100

hsCRP(mg/L)

160
140
120
140

1.2

p<0.0001

Fluvastatin

80
2000

baPWV (cm/sec)

1.2

p<0.0001

Control
ControlFluvastatin

180

180
160

LDL-C (mg/dL)

LDL-C (mg/dL)

200
200

9

12

kPa) in the control group, the administration of fluvastatin produced no changes in IMT but resulted indecreased Eθ (Fig. 9). Histogram analysis of the data
revealed an increase in areas corresponding to Eθ of
20−200 kPa, as well as a decrease in relatively hardened areas.
Discussion
We first obtained an in vivo elasticity library withtranscutaneous ultrasound from observation of various
tissues of the carotid arterial wall by the phased tracking method (Fig. 4). In in vitro experiments, asreported previously, the main components of tissue
have their own elasticities 16). By referring to the elasticity library, we can speculate as to the tissue typewhose elastic modulus is close to that of each pixel.
Second, this was a new challenge to show thedetection of the effects of fluvastatin on the elasticityof the carotid artery (mean Eθ). The mean Eθ can bechanged by statin treatment; that is, in the group withmax IMT ≥ 1.1 mm, both the mean Eθ and mean
IMT decreased significantly. On the other hand, inthe max IMT ＜ 1.1 mm group, the mean Eθ but not
the mean IMT decreased significantly.
B-mode ultrasound is widely used to measureCarotid Plaque Stabilization by Statin

669

Figure 6
1.2
1.1
1.0
0.9
0.8
0.7
0.6
0.5

p=0.0373

*#
0

3

6

9

mean E� (kPa)mean IMT (mm)A All sites (242 sites)

12

1.4
1.3
1.2
1.1
1.0
0.9
0.8
0.7
0.6

p=0.049

*
0

3

6

9

mean E� (kPa)mean IMT (mm)

B Max IMT > 1.1 mm �144 sites�

12

C Max IMT < 1.1 mm (98 sites)NS

0.8

mean E� (kPa)mean IMT (mm)

0.9

0.7
0.6
0.5

0

3

6
months

9

12

240
220
200
180
160
140
120
100
80

240
220
200
180
160
140
120
100
80

240
220
200
180
160
140
120
100
80

p<0.0001

ControlFluvastatin

*
0

3

*
6

*#
9

12
p=0.0042

*
0

3

6

*
9

12
p=0.0064

*
0

*
3

6

*
9

12

months* p<0.05
p<0.05Control
Fig. 6. Effectsof fluvastatinon the mean#IMT
and vsmean
Eθ.
vs 0 months(A) At all sites of all patients (242 sites), both the mean IMT and mean Eθ were significantly
decreased (ANOVA). Compared with the values at 0 months, the values of mean Eθ at 3, 6 and 12
months and mean IMT at 12 month were significantly decreased. No difference of the mean IMTor mean Eθ was seen between the control (40 sites in 10) and the fluvastatin group at 0 months.
Fluvastatin significantly reduced both the mean IMT and mean Eθ in the comparison of the twogroups at 12 months. (B) In the group with max IMT ≥ 1.1 mm (144 sites), both mean IMT andmean Eθ were significantly decreased. (C) In the group with max IMT ＜1.1mm (98 sites), mean Eθ
but not mean IMT was significantly decreased. P-values were obtained by ANOVA for the fluvastatin group. ＊p ＜0.05 vs. 0 months of the fluvastatin group; ＃p ＜0.05 vs. 12 months of the control.

carotid IMT, an early change that precedes the development of macroscopic plaques. Carotid IMT isincreasingly being used for cardiovascular risk stratification and as an intermediate end point in clinical trials 19), but it has limited ability to determine the extent
of plaque burden and the nature of plaque composition. IBS analysis has shown associations between
echo-lucent carotid plaques and lower HDL-C cholesterol, elevated triglycerides, elevated blood pressureand elevated blood sugar levels 7, 8); however, the IBS
index simply reflects differences in echogenicity, butdoes not show the true elasticity of the arterial wall.
Carotid stiffness, considered an index of sclerosis, canbe determined by the following equation: β= ln(SBP/
DBP)/[(Ds−Dd)/Dd], where Ds and Dd are the endsystolic and end-diastolic diameters of the common
carotid artery, respectively 20, 21); however, this value
does not indicate the two-dimensional distribution of
carotid arterial wall elasticity. Under the assumptionof a blood vessel is a uniform cylinder pipe, PWV andstiffness β can be used to evaluate average elastic properties of the whole transmission passage and the entire
circumference by measuring pulse wave velocity
between dozens of centimeters and alteration of thediameter by beat, respectively. These methods are notnecessarily suitable for comparing plaques or individuals, or for assessing qualitative changes. In contrast,
our method enables the measurement of elasticity distribution in the intima-medial region by ultrasoundand the evaluation of local (lesion) elastic properties
by measuring mural thickness change. In a previous
study, age, blood pressure and pulse rate were shown
to be independent determinants of the average valuesEθ of elasticity distribution 15).
In the present study, an elasticity library withtranscutaneous ultrasound was obtained by observing
various tissues of the carotid arterial wall (Fig. 4). We
were fortunate to be able to observe an uncomplicatedYamagishi et al .

670

Figure 7
1.2
1.1
1.0
0.9
0.8
0.7
0.6
0.5

NS

0

3

6

9

mean E� (kPa)mean IMT (mm)A E� < 100 (kPa) at 0 months (42 sites)

12

180
160
140
120
100
80
60

p<0.0001

*

*
0

3

*

6

*
9

12

1.2
1.1
1.0
0.9
0.8
0.7
0.6
0.5

NS

mean E� (kPa)mean IMT (mm)

B 100 < E� < 200 (kPa) at 0 months (154 sites)

0

3

6

9

12

1.3
1.2
1.1
1.0
0.9
0.8
0.7
0.6
0.5

NS

0

3

6
months9mean E� (kPa)mean IMT (mm)C E� > 200 (kPa) at 0 months (46 sites)

12

200
180
160
140
120
100
80

p<0.0001

*
0

3

6

350

9

12

p=0.0288

300
250

*

200

*

*

3

6
months

*

150
100

0

9

12

* p<0.05 vs 0 month
Fig. 7. Effects of fluvastatin on the mean IMT and mean Eθ in the three groups according
to the mean Eθ values at 0 months.
(A) In the group with Eθ ≤ 100 (42 sites), the mean Eθ was significantly increased after 1-monthadministration of the drug. (B) In the group with 100 kPa ＜ Eθ＜ 200 kPa (154 sites), mean Eθ
was significantly decreased. (C) In the group with 200 ≤ Eθ (46 sites), the mean Eθ was significantly decreased after 1-month administration of the drug.

healing process following carotid artery dissection,
since such cases usually heal spontaneously 22), and toobtain the elastic modulus, equivalent to blood clot
and fibrosis information by histogram. When the relative interrelationships for elasticity in the corresponding vascular tissues were considered in this study, aremarkable similarity was noted to the elasticity findings for lipids, thrombus, smooth muscle, fibrotic tissue and calcification in isolated atherosclerotic vascular specimens as documented by Inagaki et al. 23).
Many risk factors are associated with endothelial
dysfunction, including hypercholesterolemia, hypertension, cigarette smoking and diabetes mellitus.
Increasing evidence indicates that oxidized low-density
lipoprotein cholesterol (ox-LDL) plays an importantrole in endothelial dysfunction. Ox-LDL induces
endothelial injury, inhibits apoptosis, monocyte adhesion and platelet aggregation, and also inhibits theexpression/activity of endothelial nitric oxide synthase(eNOS) 24). Previous research has shown that 3-hydroxy3-methylglutaryl coenzyme A reductase inhibitors(statins) improve endothelial function throughmechanisms that go beyond their primary therapeutic
effects 25-28). Alteration of the endothelial function
might result from increases in eNOS activity, reduced
production of free radicals, inhibition of ox-LDL
action, or other undefined mechanisms. In addition,
coronary spasm is associated with endothelial dysfunction, and fluvastatin has been reported to reduce thenumber of patients with ACh-induced coronary spasm,
which is associated with endothelial dysfunction, incomparison with conventional calcium channel
blocker therapy 26). In our study, fluvastatin increased
NO and decreased hs-CRP. In the group with max
IMT ＜ 1.1 mm, considered to be in the range of subclinical atherosclerotic findings, the elastic modulus
was significantly reduced. This may be related to fluvastatin-induced improvement in endothelial dysfunction. Additionally, statin treatment is reported todecrease carotid arterial stiffness but not IMT (initial
max IMT 0.95±0.4 mm) in patients with hypercholesterolemia 20). Ultrasonic strain imaging has theCarotid Plaque Stabilization by Statin

671

Figure 8A

B

B-mode (before)

B-mode (after Fluvastatin)

160

30

Before
After

140

10
0

120

-10

E� (before)E� (after Fluvastatin)E� (kPa)
1000-20Frequency

Mean IMT 1.1mm

Frequency

100

Mean IMT 1.4mm

80
60

-40
-50
-70

20

-80
-90

0

250

-30

-60

40

500

Decrement
Increment

20

0

200

400

600

800 1000

-100

0

200

400

E� ave 92

600

800 1000E� (kPa)E� (kPa)0

E� ave 221

Fig. 8. A case of increased mean Eθ after fluvastatin administration (50-year-old man).
Images before and after administration of the drug for 12 months are shown. In the histograms, theincrease in Eθ showed a decrease of lipid/smooth muscle-rich tissue and/or an increase of collagenfibers, deterring B-mode findings.

A

B
B-mode (before)

B-mode (after Fluvastatin)
60

40

Before
After

50

30

E� (before)E� (after Fluvastatin)E� (kPa)
1000
500
250

20

30

Frequency��������������

Frequency

40��������������

20
10

0

200

400

E� (kPa)E� ave 230

E� ave 159

10

0

-10

0
-10

Decrement
Increment

600

800

-20

0

200

400

600

800

E� (kPa)

0

Fig. 9. A case of decrease in mean Eθ after fluvastatin administration (50-year-old man).
Images before and after administration of the drug for 12 months are shown. In the histograms, thedecrease in Eθ shows an increase in areas corresponding to Eθ of 20−200 kPa, as well as a decrease inrelatively hardened areas, indicating improvement of endothelial dysfunction.

potential to become a sensitive tool for detecting theeffects of early statin intervention.
Recent studies, including those on advanced
plaque formation, have also shown benefits from
intensive statin therapy in slowing and even reversing
the progression of atherosclerosis, as assessed by surrogate imaging measures such as IVUS and carotid IMT.
In a study of the effectiveness of rosuvastatin against
increases of intima media thickness, a statistically significant slowing in the progression rate of maximum
carotid IMT (initial max IMT 1.15±0.19 mm) was
observed in the rosuvastatin group relative to the placebo group for 12 carotid sites, as well as for eachindividual segment 27). Subanalysis of this METEOR
study found that in the rosuvastatin group, a greater
trend toward regression was seen in patients with morerisk factors and greater baseline thickness 28). Similarly,
in our study, a greater trend toward regression wasseen in the advanced plaque group (max IMT ≥ 1.1
mm, Fig. 6).
As shown in Fig. 7, changes in mean Eθ may berelated to an incremental beneficial effect on plaquestabilization with statins; for example, in the groupwith Eθ ≤ 100 kPa, the softer areas, mean Eθ significantly increased from 1 month after administration of
the drug. This may be the result of a diminution of

672

Yamagishi et al .

the lipid-rich core, a reduction of inflammatory cells
within the plaques, decreased macrophage activation
as well as foam cell formation, and events related tothickening of the fibrous cap as reported by several
investigators 29-31). On the other hand, in the groupwith 200 kPa ≤ Eθ, the harder areas, mean Eθ was significantly decreased. This may be related to improvement of endothelial dysfunction, such as paradoxic
vasoconstriction of cardiac vessels. In the group with100 kPa ＜ Eθ＜ 200 kPa, the middle range of the elastic modulus, mean Eθ significantly decreased. Thismay be the result of variable changes of plaque components mentioned above.
On the basis of comparing histograms for tissue
characterization (Fig. 4) and fluvastatin-inducedchanges in the intima-media complex (Fig. 6, 7, 8, 9),
it may be possible to perform tissue characterizationby comparing elasticity distribution and chronologicalchanges in the distribution as measured by ultrasound.
Collagen and smooth-muscle content have beenreported to be important factors in determining thestability of the fibrous cap on atherosclerotic plaques;
therefore, correlations between elasticity and tissueelements within the arterial wall may provide useful
information for the noninvasive diagnosis of plaquevulnerability 23).
The present study has the following limitations:
Although plaque can be evaluated three-dimensionallyand relatively more easily with certain methods, suchas computed tomography and magnetic resonanceimaging, conventional IMT measurement is performed
in one cross-section, making it difficult to constructthree-dimensional images, which would be a future
hurdle to overcome. In addition, measurement of one
cross-section influences not only elasticity, but also the
IMT. In the present study, to measure the same sitesof carotid arteries, one well-trained sonographer performed the IMT and elastic modulus measurementsby comparing current B-mode images with past Bmode images on a PC and positioned the probe so as
to measure a similar position. Finally, the spatial averages of the elastic moduli were compared so that themean value would be the same when the same area
was included.
Plaques consist of various tissue components,
and we previously reported the comparison of theelastic modulus with histologically corresponding tissues, such as lipids, blood clots and calcification 16). In
that report, the distributions of lipids and blood clots
were relatively narrow and pure; however, those of collagen, smooth muscle and calcification were wide andrather scattered. The rank of the center of elasticity
distributions obtained in the present study was similar

to that of in vitro experiments, indicating that it is
impossible to obtain completely pure plaque tissue
components, although the major components of theplaque can be detected and a classification can bemade based on the elastic modulus.
Eθ may be an ideal marker to show the elasticityof the arterial wall itself because Er (apparent elasticity) is influenced not only by an increase in elasticity
but also by the increasing thickness of the arterial wallat the same location, especially in areas of relatively
great thickness; however, the influence of blood pressure on the library cannot be neglected. The effects ofaging on the mean IMT and mean Eθ are shown in
Fig. 1. It seems that Eθ does not increase greatly innormotensives, even with advancing age. In hypertensives, however, Eθ was found to increase not only withdecrements of strain (the denominator of Eq. (3) forelastic modulus) via tissue characteristics such as fibrosis, calcification and local contraction of smooth muscle, but also with increments of pulse pressure (thenumerator of Eq. (3) for elastic modulus). A comparison with pathological findings for the elastic modulus
is needed; however, in humans it is ethically unacceptable to compare ultrasonic images with pathologicalfindings in early-stage carotid arterial atherosclerosis.
In addition, an elastic modulus obtained by means oftranscutaneous ultrasound differs from that obtained
from isolated atherosclerotic vascular specimens, since
the former is also affected by subcutaneous tissue,
such as muscle and fat.
In summary, the present non-invasive echographic carotid arterial elasticity data suggest that thismethod is useful for classification and evaluation ofatherosclerotic plaques that increase with progressive
lipid deposition and that the measurement data may
reflect chronological and histopathological changes.
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