Full text of DOI:10.1002/clc.4960250405

Clin. Cardiol. 25, 154–160 (2002)
Shear-Induced Platelet Aggregation Increases in Patients with Proximal and
Severe Coronary Artery Stenosis
KOICHI KAWANO, M.D., HIDEAKI YOSHINO, M.D., NOBUO AOKI, M.D., FACC, HIROSHI UDAGAWA, M.D., ATSUSHI WATANUKI, M.D.,
YOSHIKO HIOKI, M.D., YUKIHISA HASUMURA, M.D., TOSHIKAKI YASUMURA, M.D., MASASHI HOMORI, M.D., MITSURU MURATA, M.D.,*
YASUO IKEDA, M.D.,* KYOZO ISHIKAWA, M.D., FACC
Second Department of Internal Medicine, Kyorin University School of Medicine, and *Department of Internal Medicine, Keio
University School of Medicine, Tokyo, Japan
Summary
Background: Shear stress generated in stenosed arteries
promotes platelet thrombi formation at the stenosed sites by
accelerating the binding of von Willebrand factor (vWF) to
platelets. Shear-induced platelet aggregation (SIPA) has been
studied in acute coronary syndromes, but not in chronic coro-
nary disease.
Conclusions: Shear-induced platelet aggregation is in-
creased in patients with severe stenosis of the proximal cor-
onary arteries and correlates with plasma concentrations of
vWF larger multimers, suggesting that severe stenosis in the
proximal segments is not only associated with an increased
risk of significant myocardial ischemia, but may also generate
high shear stress in the stenosed artery and increase plasma
vWF larger multimers, thereby promoting the formation of
platelet thrombi.
Hypothesis: We investigated the effect of both the site and
severity of coronary stenosis on SIPA in patients with chronic
coronary artery disease.
Key words: stenosis, coronary disease, von Willebrand factor,
Methods: Shear-induced platelet aggregation was measured
using platelet-rich plasma in 49 patients (41 men and 8 wom-
en; mean age 61 ± 10 years) with coronary artery disease to
evaluate the association between the extent of SIPA and coro-
nary angiographic findings. Stenoses > 75% were considered
severe. In all, 62 healthy individuals (54 men and 18 women;
mean age 45 ± 7 years) served as controls. The correlation be-
tween SIPA and the site and severity of the coronary lesion, and
parameters of coagulation and fibrinolysis were evaluated.
Results: Shear-induced platelet aggregation was increased
in the stenosis group (69.0 ± 10.6%) compared with the con-
trols (57.7 ± 10.3%, p < 0.0001). Patients with severe stenosis
in the proximal segments had significantly increased SIPA (p<
0.0001) and vWF larger multimer concentration (p < 0.0001)
compared with the control group. A significant correlation ex-
isted between SIPA and the vWF larger multimer concentra-
tion in all subjects studied (r = 0.422, p < 0.0001).
platelet
Introduction
Platelet activation and subsequent platelet adhesion and ag-
gregation play a role in the initiation and exacerbation of acute
coronary syndromes, including acute myocardial infarction
(MI).1–6 Shear stress has been considered an important pro-
moter of adhesion and cohesion of platelets to endothelial cells
activated by atherosclerosis.7 The shear stress generated when
blood flows through a vessel increases as blood flows rapidly
through a stenosed artery,8 which accelerates thrombus forma-
tion by activating platelets and inducing platelet adhesion and
aggregation.8–10
Most studies of platelet function, coagulation, and fibri-
nolysis in patients with ischemic heart disease have focused
on acute coronary syndromes.1, 2, 11–15 However, no study has
examined angiographic findings and hematologic parameters
to determine which characteristics place patients with stable
ischemic heart disease at risk of transition to acute coronary
syndrome.
Address for reprints:
We developed a shear-induced platelet aggregometer to
measure shear-induced platelet aggregation (SIPA).16 The de-
vice is useful for examining platelet aggregation after activa-
tion by shear stress in stenosed arteries.
The objective of the present study was to investigate the ef-
fect of the site and severity of coronary stenoses on SIPA in pa-
tients with stable ischemic heart disease who underwent coro-
nary angiography.
Hideaki Yoshino, M.D.
Kyorin University School of Medicine
Second Department of Internal Medicine
6-20-2 Shinkawa, Mitaka
Tokyo 181-8611, Japan
e-mail: yoshino@kyorin-u.ac.jp
Received: September 13, 2000
Accepted with revision: July 3, 2001

K. Kawano et al.: Shear-induced aggregation and coronary stenosis
155
duced by high shear stress that is von Willebrand factor (vWF)
dependent. We compared the maximal value (%) of the high
shear-induced platelet aggregation in this study.
Methods
Study Population
Parameters of Coagulation, Fibrinolysis, Platelet Activity,
and Catecholamines
In all, 49 patients with coronary artery disease (26 with pri-
or MI and 23 with stable angina pectoris), presenting to the
Kyorin University Hospital for routine coronary angiography
from May to December in 1997, were enrolled in the study.
The subjects included 41 men and 8 women (mean age 61.2 ±
0.3 years, range 34–80). Patients with acute MI or unstable
angina pectoris were excluded from the study since elevated
plasma catecholamines in these patients are known to affect
SIPA. Patients were included 3 months after the onset of MI,
and those with unstable angina pectoris were eligible if angina
was stabilized for at least 3 months. Sixty-two healthy subjects
(54 men and 8 women; mean age 44.7 ± 7.0 years, range
34–59) without a history of ischemic heart disease or a positive
treadmill test served as controls. All subjects gave informed
consent prior to participation in this study. The study was ap-
proved by the Institutional Review Committee of Kyorin
University Hospital, and procedures were followed in accor-
dance with institutional guidelines.
Platelet factor 4, ꢁthromboglobulin, and vWF antigen were
measured using an enzyme immunoassay (Asserachrom PF4,
Asserachrom ꢁ-TG and Asserachrom vWF, Diagnostica
Stago, Paris, France). Plasminogen activator inhibitor-1 was
measured via enzyme immunoassay (H-5 PAI-1 ELISA kit,
Monozyme, Hoersholm, Denmark). Thrombin-antithrombin
III complex was assayed using an enzyme immunoassay (TAT
kit, SRL, Tokyo, Japan). Plasmin-antiplasmin complex was
determined by latex agglutination (LPIA ACE PPI, Yatron,
Tokyo, Japan). Antiplasmin was measured by a functional
assay (Testzyme S APL, Daiichi-Kagaku, Tokyo, Japan). Pro-
thrombin fragment F1+2 was determined by enzyme im-
munoassay (Enzygnost F1+2, Dade Behring, Marburg, Ger-
many). Fibrinogen was estimated by the thrombin clotting
time (Fibrinogen a BMY, Diagnostica Stago). Epinephrine
and norepinephrine were measured by high performance liq-
uid chromatography and an assay kit (CA test TOSOH, Tosoh,
Yamaguchi, Japan).
Preparation of Platelet-Rich Plasma
Venous blood (3.8% citrate/blood 1:9) was collected from
all subjects and centrifuged at 100 g for 15 min at 22˚C. Plate-
let-rich plasma was prepared from the supernatant, and the re-
maining blood was centrifuged at 3000 g for 20 min to obtain
platelet-poor plasma. The platelet count in the platelet-rich
plasma was determined using a Coulter counter (S-Plas IV,
Coulter Electronics, Miami, Fla., USA) and the platelet con-
centration was adjusted to 150 ꢀ 109/liter with platelet-poor
plasma. Acetylsalicylic acid (81 mg/day), which reportedly
does not affect SIPA,17 was given to all subjects. Ticlopidine,
which is known to inhibit SIPA, was prohibited during the 2
weeks prior to blood collection.
von Willebrand Factor Larger Multimer
Analysis of the vWF larger multimer was performed using
an SDS-agarose gel electrophoresis. Larger multimers were
defined as bands above the 10th band from the bottom. The
relative amount (%) of each band was measured using a den-
sitometer (Advantec DM303, Nihon Denshi Kagaku, Tokyo,
Japan). The total amount of the larger bands was calculated ac-
cording to the following formula:
The total amount of the larger bands (%) = plasma vWF
antigen (%; the antigen in the pooled plasma of normal sub-
jects defined as 100%) ꢀA/100, where A is the sum of the rel-
ative amounts of larger multimers (%).
Measurement of Shear-Induced Platelet Aggregation
Coronary Angiography
Shear-induced platelet aggregation was measured accord-
ing to a method described previously.16 Briefly, 400 µl of plate-
let-rich plasma was applied to the surface of a polymethyl-
methacrylate plate and exposed to varying degrees of shear
stress. After an initial 15 s at 6 dyn/cm2, the shear stress was
maintained between 6 and 12 dyn/cm2 for 90 s, then between
12 and 108 dyn/cm2 for the next 120 s, and constant at 108
dyn/cm2 for the last 90 s. A laser light with a wavelength of
633 nm was applied to the platelet-rich plasma, and the platelet
aggregation was calculated according to the following formu-
la:18 platelet aggregation (%) = log(Ia/Ib)/(Ipp/Ib) ꢀ 100,
where Ia and Ib indicate transmitted light intensity of the
platelet suspension after and before the application of shear
stress, respectively, and Ipp is the transmitted light intensity of
the platelet-free suspending medium. Platelet aggregation is
biphasic, with the first aggregation induced by low shear stress
that is fibrinogen dependent, and the second aggregation in-
Coronary angiography was performed in multiple views us-
ing the Judkins technique. Serial orthogonal coronary angio-
grams were obtained after maximal coronary vasodilation
with intracoronary injection of isosorbide dinitrate. The loca-
tion of the stenosis was determined according to the Coronary
Artery Surgery Study (CASS) classification system.19 The
severity of coronary artery stenosis was measured by a caliper
method, using a Cardio-500 (Fukuda Denshi Co., Ltd., Tokyo,
Japan), and the severity of each stenosis was assessed accord-
ing to the American Heart Association classification system.20
Subjects were classified into two groups according to the de-
gree of coronary stenosis: mild stenosis group (≤75% steno-
sis), and severe stenosis group (> 75% stenosis). Subjects were
also classified as follows into two groups according to the lo-
cation of the most severe stenosis: proximal stenosis group

156
Clin. Cardiol. Vol. 25, April 2002
(patients with CASS seg. 1, seg. 11, seg. 12, or seg. 18), and
distal stenosis group (patients with the other distal segment le-
sions). Each patient was classified into one of four groups ac-
cording to the location and stenosis of the most severe stenotic
lesion: proximal-severe stenosis group, distal-severe stenosis
group, proximal-mild stenosis group, and distal-mild stenosis
group. Coronary angiograms were assessed independently by
two experienced observers blinded to the clinical findings and
SIPA results. In cases of discordant evaluations, the results
were discussed and the difference reconciled.
(p < 0.0001). Since a significant difference was observed in
the ages of the patient and healthy groups (p< 0.0001), simple
regression analysis was performed using age as the indepen-
dent variable and SIPA as the dependent variable. The regres-
sion was not significant, with a p value of 0.579 for the patient
group and 0.642 for the healthy group. It was concluded that
the two variables were not correlated, and that age does not
affect the extent of SIPA.
Coronary Artery Lesion and Shear-Induced Platelet
Aggregation
Statistical Analysis
There were no significant differences in SIPA between sin-
gle- and multivessel disease (p = 0.77). There were no signif-
icant differences among the four angiographic groups of sin-
gle-, double-, triple-vessel disease and normal coronary artery
(p = 0.80).
Based on the site and severity of coronary stenosis, the sub-
jects were divided into the following groups: mild stenosis in
the distal segments (9 patients), mild stenosis in the proximal
segments (17 patients), severe stenosis in the distal segments
(10 patients), and severe stenosis in the proximal segments (13
patients). No significant difference existed among the four
groups in age, gender, underlying disease (MI or angina pec-
toris), coronary risk factors (hypertension, diabetes mellitus,
hyperlipidemia, and smoking), or medications (beta blockers,
calcium antagonists, angiotensin-converting enzyme inhibi-
tors, or nitrates) (Table I).
Statistical analysis was performed using analysis of vari-
ance for comparison of SIPA and vWF larger multimers. Sta-
tistically significant differences among the five groups were
determined by a one-way analysis of variance (ANOVA), fol-
lowed by a Bonferroni method if the ANOVA probability val-
ue was < 0.05. Association between SIPA and age was eval-
uated using regression analysis. Association between SIPA
and parameters of coagulation, fibrinolysis, and platelet activ-
ity was evaluated with Pearson’s correlation coefficient. The
Kruskal-Wallis test was used to compare clinical characteris-
tics between the subgroups. A p value of < 0.05 was consid-
ered statistically significant.
Results
Comparison of SIPA in the five groups, including the
healthy group, demonstrated that SIPA was greatest in the pa-
tients with severe stenosis in the proximal segments (76.1 ±
9.9%), followed by those with severe stenosis in the distal
segments (67.11 ± 11.6%), mild stenosis in the proximal seg-
ments (66.01 ± 10.7%), mild stenosis in the distal segments
(66.51 ± 6.6%), and the healthy group, which had the lowest
Shear-Induced Platelet Aggregation in Patients with
Ischemic Heart Disease and Healthy Controls
Shear-induced platelet aggregation was increased signif-
icantly in the patients with ischemic heart disease (69.0 ±
10.6%) compared with the healthy controls (57.7 ± 10.3%)
TABLE I Baseline characteristics of the patient groups
Clinical factors
Group 1 (n = 9)
Group 2 (n = 17)
Group 3 (n = 10)
Group 4 (n = 13)
Age, years (mean ± SD)
Male gender, n (%)
61.2 ± 12.1
9 (100)
5 (56)
63.5 ± 7.5
15 (88)
10 (59)
5 (29)
56.5 ± 10.5
9 (90)
6 (60)
61.9 ± 11.8
8 (62)
4 (31)
Myocardial infarction, n (%)
Angina pectoris, n (%)
Coronary risk factors
Hypertension, n (%)
Diabetes mellitus, n (%)
Hyperlipidemia, n (%)
Smoking habit, n (%)
Medications
4 (44)
4 (40)
9 (69)
2 (22)
1 (11)
4 (44)
7 (78)
7 (41)
8 (47)
5 (29)
13 (76)
6 (60)
3 (30)
3 (30)
9 (90)
7 (54)
5 (38)
9 (69)
8 (62)
Beta blocker, n (%)
1 (11)
5 (56)
1 (11)
4 (44)
5 (29)
11 (65)
6 (35)
7 (41)
4 (40)
5 (50)
4 (40)
4 (40)
4 (31)
11 (85)
1 (8)
Calcium antagonist, n (%)
ACE inhibitor, n (%)
Nitrates, n (%)
7 (54)
Group 1: Group with mild stenosis-distal segment, Group 2: Group with mild stenosis-proximal segment; Group 3: Group with severe stenosis-
distal segment; Group 4: Group with severe stenosis-proximal segment.
Abbreviations: ACE = angiotensin-converting enzyme, SD = standard deviation.

K. Kawano et al.: Shear-induced aggregation and coronary stenosis
157
90
80
70
60
50
40
30
20
10
0
proximal segments, and mild stenosis in the distal segments.
Shear-induced platelet aggregation values in the healthy group
were significantly lower than those in all the other groups (vs.
severe stenosis in the proximal segments: p < 0.0001, vs. se-
vere stenosis in the distal segments: p = 0.0081; vs. mild steno-
sis in the proximal segments: p = 0.0037; and vs. mild stenosis
in the distal segments: p = 0.0172).
*
Relationship between Parameters of Coagulation,
Fibrinolysis, and Platelet Activity, and Catecholamines
(Table II)
No significant correlation was observed between SIPA and
parameters of coagulation (vWF, fibrinogen, thrombin-an-
tithrombin III complex, and prothrombin fragment F1 + 2),
fibrinolysis (plasminogen activator inhibitor I, antiplasmin,
and plasmin-antiplasmin complex), or platelet activity (ꢁ-
thromboglobulin and platelet factor 4). Similarly, no correla-
tion existed between epinephrine or norepinephrine, parame-
ters of sympathetic activity, and SIPA.
FIG. 1 Shear-induced platelet aggregation (SIPA) (%) for the pa-
tient and healthy groups: The graph displays the mean (%) and each
bar represents 1 standard deviation. Shear-induced platelet aggrega-
tion was greatest in the patients with severe stenosis in the proximal
segments (76.1 ± 9.9%, *: p<0.0001), followed by those with severe
stenosis in the distal segments (67.1 ± 11.6%), mild stenosis in the
proximal segments (66.0 ± 10.7%), and mild stenosis in the distal
segments (66.5 ± 6.6%). The controls had the lowest value (57.7 ±
10.3%). No significant difference existed between the patients with
mild stenosis in the distal segments, mild stenosis in the proximal
segments, or severe stenosis in the distal segments. The SIPA values
in the healthy group were significantly lower than those in all the
other groups. Control: healthy subjects; mild-distal: patients with
mild stenosis in the distal segments; mild-proximal: patients with
mild stenosis in the proximal segments; severe-distal: patients with
severe stenosis in the distal segments; severe-proximal: patients with
severe stenosis in the proximal segments.
von Willebrand Factor Larger Multimer Analysis (Fig. 2)
The plasma concentration (%) of vWF larger multimers
was greatest in the patients with severe stenosis in the proximal
segments (85.4 ± 43.0%), followed by those with mild steno-
sis in the proximal segments (59.91 ± 33.3%), mild stenosis in
the distal segments (52.8 ± 35.8%), severe stenosis in the distal
segments (46.8 ± 17.2%), and the healthy group, which had
the lowest value (38.4 ± 20.1%) (p < 0.0001). The vWF larger
multimer concentration in the patients with severe stenosis in
the proximal segments was significantly greater than that in
the other four groups (vs. severe stenosis in the distal seg-
ments: p = 0.0016; vs. mild stenosis in the proximal segments:
p = 0.0240; vs. mild stenosis in the distal segments: p =
0.0109; and vs. the healthy group: p < 0.0001). No significant
difference in vWF larger multimer concentration was ob-
value (57.71 ± 10.3%) (p < 0.0001, Fig. 1). The SIPA in the
patients with severe stenosis in the proximal segments was
greater than that in any of the other groups (vs. severe stenosis
in the distal segments: p = 0.0392; vs. mild stenosis in the
proximal segments: p = 0.0088; vs. mild stenosis in the distal
segments: p = 0.0334; and vs. controls: p < 0.0001). No sig-
nificant difference in SIPA was noted between the patients
with severe stenosis in the distal segments, mild stenosis in the
TABLE II Association between shear-induced platelet aggregation and other hematologic parameters
Parameter
n
Mean ± SD
Correlation coefficient
p Value
vWF antigen (%)
Fibrinogen (mg/dl)
49
49
49
49
49
49
49
46
43
49
49
158 ± 55
331 ± 126
7 ± 10
0.159
ꢂ0.137
ꢂ0.063
ꢂ0.028
0.176
ꢂ0.048
0.009
ꢂ0.108
ꢂ0.077
ꢂ0.038
0.007
0.228
0.301
0.648
0.838
0.194
0.726
0.949
0.491
0.623
0.806
0.963
Thrombin-antithrombin III complex (ng/ml)
Prothrombin fragment F1+2 (nmol/l)
Plasminogen activator inhibitor-1(ng/ml)
Antiplasmin (%)
Plasmin-antiplasmin complex (µg/ml)
Beta thromboglobulin (ng/ml)
Platelet factor 4 (ng/ml)
1 ± 0.4
199 ± 93
102 ± 11
1 ± 0.3
116 ± 96
58 ± 63
78 ± 62
418 ± 213
Epinephrine (pg/ml)
Norepinephrine (pg/ml)
Data are displayed as mean ± SD.
Abbreviations: SD = standard deviation, vWF = von Willebrand factor.

158
Clin. Cardiol. Vol. 25, April 2002
Discussion
140
120
100
*
Coronary Angiography Findings in the Chronic Phase and
Shear-Induced Platelet Aggregation
80
60
40
Exposure of tissue factor and collagen on the endotheli-
um following plaque rupture results in platelet activation and
the formation of platelet thrombi in acute coronary syn-
drome.1, 2, 21, 22 In contrast, the role of platelets in the chronic
phase of ischemic heart disease is not yet understood. The re-
sults of the present study demonstrate that SIPA increases
appreciably in patients with chronic ischemic heart disease
compared with healthy individuals. Shear-induced platelet
aggregation was greatest in patients with severe stenosis in
the proximal segments. Although a previous study observed
an increase in SIPA in patients with acute MI,23 ours is the
first study to demonstrate a relationship between coronary
angiography findings and SIPA in ischemic heart disease in
the chronic phase.
An important finding of the present study is that vWF-de-
pendent platelet aggregation is increased in chronic ischemic
heart disease as well as in acute coronary syndromes. Our re-
sults also suggest that patients with severe stenosis in the prox-
imal segments are at increased risk of exacerbation of stenotic
lesions due to the formation of platelet thrombi.
20
0
FIG. 2 von Willebrand factor (vWF) larger multimer concentration
(%) for the patient and healthy groups: The graph displays the mean
(%) and each bar represents 1 standard deviation. The vWF larger
multimer concentration was greatest in patients with severe stenosis
in the proximal segments (85.4 ± 43.0%, *: p< 0.0001), followed by
those with mild stenosis in the proximal segments (59.9 ± 33.3%),
mild stenosis in the distal segments (52.8 ± 35.8%), and severe
stenosis in the distal segments (46.8 ± 17.2%). The controls had the
lowest value (38.4 ± 20.1%). No significant difference existed be-
tween the patients with mild stenosis in the proximal segments, mild
stenosis in the distal segments, and severe stenosis in the distal seg-
ments. Group definitions as in Figure 1.
Shear Stress and Shear-Induced Platelet Aggregation
served among patients with mild stenosis in the proximal seg-
ments, those with mild stenosis in the distal segments, and
those with severe stenosis in the distal segments. The vWF
larger multimer concentration in the healthy group was signif-
icantly lower than that in the patients with severe stenosis in
the proximal segments or mild stenosis in the proximal seg-
ments (p< 0.0001 and p = 0.0094, respectively).
A significant correlation did exist between SIPA and vWF
larger multimer concentration in the patients and controls as a
whole (r = 0.422, p< 0.0001) (Fig. 3).
The shear-induced platelet aggregometer16 is a modified
cone-and-plate viscometer that induces platelet aggregation by
applying shear stress in the absence of agonists. It is useful not
only for examining SIPA by subjecting resting platelets and
plasma proteins to shear stress, but also for evaluating interac-
tions between plasma proteins and platelets already activated
by shear stress in stenosed arteries in vivo. A previous study
using the device has reported that SIPA was accelerated after
exercise in patients with angina pectoris,24 but did not evaluate
the patients on the basis of coronary angiography findings.
The shear stress generated in an artery is proportional to
blood flow, and inversely proportional to the third power of
the internal diameter of the artery.25 Severe stenosis in the
proximal segments should therefore result in extremely high
shear stress. The increase in SIPA observed ex vivo may be at-
tributable to the application of shear stress to platelets, plasma
proteins, or endothelial cells due to severe stenosis in the
proximal segments in vivo.
High SIPA is dependent on vWF and platelet glycoproteins
GPIb-IX and GPIIb-IIIa.16 Previous reports have demon-
strated that elevated plasma vWF concentration is an indepen-
dent coronary risk factor,26 and that aurintricarboxylic acid,
which inhibits binding of GPIb-IX and vWF,27 monoclonal
antibodies,28 and vWF fragments,29 and reduces thrombus for-
mation in the coronary arteries of animal models. These exper-
imental and clinical findings indicate that vWF and the platelet
glycoproteins GPIb-IX and GPIIb-IIIa play an important role
in atherosclerosis and arterial thrombus formation in vivo. To
determine the effect of shear stress on platelets, plasma pro-
100
90
80
70
60
50
40
30
SIPA (%) = 64.025 + 0.095 ꢀ vWF larger multimer (%)
20
0
20 40 60 80 100 120 140 160 180 200
vWF larger multimer (%)
FIG. 3 Correlation between shear-induced platelet aggregation
(SIPA) and von Willebrand factor (vWF) larger multimer concentra-
tion in the patient and healthy groups as a whole. A positive correla-
tion was observed (r = 0.422, p< 0.0001).

K. Kawano et al.: Shear-induced aggregation and coronary stenosis
159
teins, and endothelial cells, we examined the correlation be-
tween SIPA and parameters of plasma activity, plasma fib-
rinogen concentration, plasma vWF concentration, the results
of vWF larger multimer analysis, and parameters of coagula-
tion and fibrinolysis.
known. A study using atomic force microscopy demonstrated
that vWF underwent a shear stress-induced conformational
transition from a globular state to an extended chain conforma-
tion.32 This conformational change may increase the number
of sites available to interact with platelets. The number of the
interaction sites should increase with the multimer size, thus
making larger multimers more potent to support higher SIPA.
The Effect of Shear Stress on Platelet Activity
Although it has been reported that shear stress causes acti-
vation of the platelet glycoprotein GPIIb-IIIa,7 the effect of
shear stress on GPIIb-IIIa could not be evaluated in the present
study, since we did not measure GPIIb-IIIa activity. Instead,
we measured ꢁ-thromboglobulin and platelet factor 4, which
are constituents of the ꢃ-granule, as markers of platelet secre-
tion. However, no correlation existed between these param-
eters and SIPA. This supports a previous suggestion30 that
platelet activation and aggregation caused by shear stress are
not accompanied by the release of ꢃ-granules.
Shear-Induced Platelet Aggregation and Coagulation and
Fibrinolysis
In an acute coronary syndrome, the progression of the ex-
trinsic coagulation pathway in the coronary endothelium stim-
ulates platelet activation through thrombin formation.21, 22
In our patients, thrombin-antithrombin III complex and pro-
thrombin fragment F1 + 2 did not correlate with SIPA, sug-
gesting that thrombin generation is not involved in the acceler-
ation of SIPA in the chronic phase of ischemic heart disease.
Similarly, SIPA did not correlate with antiplasmin, plasmin-
antiplasmin complex, or fibrin degradation product, thus ex-
cluding the involvement of fibrinolysis.
The Effect of Shear Stress on Plasma Proteins (Coronary
Angiography Findings and von Willebrand Factor Larger
Multimer Concentration)
Epinephrine has also been reported to stimulate SIPA;33
however, we found no correlation between SIPA and plasma
epinephrine concentration, removing the possibility that
platelet aggregability was stimulated by increased sympathet-
ic activity.
Low SIPA is fibrinogen dependent and high SIPA is vWF
dependent.16 In the present study, fibrinogen concentration did
not correlate with high SIPA. A previous study showed that the
vWF concentration was elevated in acute MI and correlated
with SIPA.23 In the present study, no correlation was observed
between SIPA and vWF concentration in the patient and
healthy groups overall, but a correlation was present between
SIPA and vWF larger multimer concentration (Fig. 3). Our
study also showed that the vWF larger multimer concentration
was greatest in the patients with severe stenosis in the proximal
segments, the group in which SIPA was increased the most.
A positive correlation between SIPA and vWF larger mul-
timer concentration has also been reported in patients with sta-
ble angina pectoris after treadmill exercise test,24 and in cere-
bral ischemia.17 This study is the first to correlate SIPA, vWF
larger multimer concentration, and angiography findings,
demonstrating that both SIPA and the vWF larger multimer
concentration are elevated in patients with severe stenosis in
the proximal segments. This suggests that plasma vWF larger
multimer concentration increases as a result of an unknown
mechanism and may stimulate SIPA.
Clinical Implications
Coronary angioplasty ameliorates coronary stenosis, re-
stores coronary blood flow, and decreases the risk of coronary
ischemia. However, angioplasty appears to do more than re-
store coronary flow: our results suggest that angioplasty
might decrease shear stress generated in stenosed arteries, re-
duce the release of vWF larger multimers from endothelial
cells, and suppress the formation of platelet thrombi. The re-
duction of coronary stenosis by coronary intervention should
therefore be considered for patients with severe stenosis in the
proximal segments who are stable with medical treatment.
Further prospective studies of the effects of coronary inter-
vention on SIPA and vWF larger multimer concentration are
therefore needed.
Study Limitations
The Effect of Shear Stress on Vascular Endothelial Cells
This was a study of a relatively small number of patients. A
significant correlation existed between SIPA and the vWF
larger multimer concentration, though relatively weak. A
larger study would be needed to confirm the relation between
SIPA and coronary stenosis, and between SIPA and vWF
larger multimers. The present study focused on platelet ag-
gregation but did not investigate platelet adhesion, another
important function of platelets. The vWF has also been re-
ported to play a role in platelet adhesion under high shear
stress,34 emphasizing the need to study shear-induced platelet
adhesion in ischemic heart disease.
Although the mechanism responsible for the increase in
vWF larger multimer concentration seen in the present study is
unclear, shear stress on stenosed arteries may act on the endo-
thelial cells to promote the release of vWF larger multimers.31
Shear stress generated in stenosed arteries may act on endo-
thelial cells to stimulate vWF larger multimer secretion and in-
crease the plasma vWF larger multimer concentration, thereby
further precipitating platelet aggregation.
How vWF larger multimers induce greater platelet aggrega-
tion than smaller multimers under high shear stress is un-

160
Clin. Cardiol. Vol. 25, April 2002
The role of von Willebrand factor and fibrinogen in platelet aggre-
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