Full text of DOI:10.1177/107602960100700304

Original Report
Plasma Markers of Endothelial
in Chronic
Dysfunction
Disease
Obstructive
Pulmonary
*
Giuseppe Cella, M.D., *Alessandra Sbarai, M.D.,
*Gabriella Mazzaro, M.D., *Barbara Vanzo, M.D.,
and †Jawed Fareed, Ph.D.
*
Silvia Romano, M.D., †Debra Hoppensteadt, Ph.D.,
andSurgical Sciences, University of Padova, Medical School, Padova, Italy; and †Department of
Pathology
*
Department of Medical
and
Loyola University ofChicago, Maywood, Illinois, USA
Pharmacology,
Summary: We studied some endothelial cell activity plasma
markers, nitric oxide (NO), thrombomodulin (TM), selectins
decreased
in
patients with
COPD
(p
<
0.001). In contrast, no
differences were seen in sE-selectin. Patients with COPD
(
sP-, sE-,sL-selectin:platelet-P, endothelium-E, leukocyte-L),
showed
a
significantly higher (p
<
0.001) TFPI antigen plasma
and tissue factor
chronic obstructive
inhibitor (TFPI) in 14
with
level (mean 112.28 &plusmn; 6.45
controls 77.68 &plusmn; 0.28
pathway
patients
ng/mL;
disease (COPD), matched with
than controls. Our data
the
of some
abnor-
pulmonary
ng/mL)
support
concept
2
0
normal controls, to evaluate their endothelial cell
form of endothelial cell
and
dysfunc-
dysfunction,
coagulation
tion. Both NO(patients: 23.42 &plusmn;1.67 &mu;g/mL; controls: 40.0 &plusmn;
malities are present in patients with COPD. However, because
3
.38 &mu;g/mL) and TM levels (patients: 5.46 &plusmn; 1.32 ng/mL;
the endothelium seems to produce
a
defense mechanism, the
controls :12.9 &plusmn; 0.51ng/mL) were significantly decreased in
potential usefulness ofan antithrombotic therapy in these pa-
PD0.001).
sP-selectin levels
(patients:
79.42 &plusmn; 18.20
significantly higher
tients needs further
investigation. Key
Words: Chronic ob-
C( p
O
<
controls: 37.5 &plusmn; 2.84
were
structive
disease&mdash;Nitric oxide&mdash;Selectins&mdash;
ng/mL,
mg/mL)
pulmonary
p <
0.02),
whereas sL-selectin levels
584.9 &plusmn; 78.98
factor
pathway inhibitor.
(
(patients:
Thrombomodulin&mdash;Tissue
controls: 1054.02 &plusmn; 61.28
were
ng/mL,
ng/mL)
significantly
Chronic obstructive
disease (COPD) is
airflow
in stable condition
with COPD as
pulmonary
dysfunction
with normal controls.
patients
a
disorder characterized
lung
by progressive
compared
obstruction and chronic inflammation in which throm-
botic
complications may occur
in the
pulmonary circu-
MATERIALSANDMETHODS
lation. However, the mechanism involved in is not
known (1-3).
fully
Westudied 14
men and 4 women, mean
patients (10
affected chronic obstructive
)
Altered endothelial cell and hemostatic markers such
68.3 ±
age
3.9)
(COPD),
by
pulmo-
as nitric oxide, soluble P-selectin, prothrombin Fl+2
disease
in stable
condition,
with no evi-
nary
dence
excretion of 11-deydro-
platelet aggregates are pre-
fragment, fibrinogen, urinary‘
of
by echocardiography
pulmonary hypertension.
or
i3-thromboglobulin,
j TxB2=
The
was established based on clinical
diagnosis
history,
sent in patients with COPD(4-10). These data seem to
physical examination,
chest
X-ray,
were
and
pulmonary
func-
support the concept
in the
that some form of endothelial cell
tion tests. The
with
a
of
patients
taking
variety
dmgs:
dysfunction
pulmonaryand/orsystemic circulation
condition
and ACEinhibit-
theophylline, P2-agonists,
in this
is present
pathologic
(11).
tors (9 patients); antiplatelet drugs (3 patients); andwar-
.
Theaimofthis
wasto evaluatethe
levels
study
plasma
farin
10 men and 10
disease, were considered as
(
2
patients). Twenty subjects,
of some
available markers of endothelial cell
recently
womenwithout
anyapparent
controls.
healthy
Venous blood
were drawn,
a
double
samples
using
into 3.8% sodium citrate
Presented in part at the XVII Congress of the International Society
onThrombosisand Hemostasis; Washington, DC;August14-21, 1999.
syringe butterfly technique,
(9:1, v/v) at baseline. The collected blood
were
samples
Address
and reprint requests to
Giuseppe
Cella,
correspondence
at
for 20 minutes at +4°C, Th~
centrifuged
3,000 g
di ScienzeMedichee Chirurgiche, Universit&agrave;
M.D.,Dipartimento
degli
was divided into
and
kept frozen
at
Studidi Padova, ViaOspedate Civile 105, 35100Padova, Italy; e-mail:
giuseppe.cella@unipd.it.
plasma
aliquots
-
’7~~~ until
analysis.
205
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2
06
Soluble adhesion molecules of the selectin class (sE,
sP and sL), with concentrations
in
expressed
commercial
ng/mL,
color-
were determined
using a
imetric sandwich ELISA
quantitative
Minne-
assay
(R&D
Systems,
Nitric oxide (NO) concentrations,
apolis, MN, USA).
in nmol/mL, were determined
using a
com-
expressed
mercial quantitative colorimetnc assay supplied by Oxis
Int, Portland, OR, USA.
Tissue factor
inhibitor
pathway
andthrombomodulin
were measured
levels,
in rt~ImL,:
antigen
expressed
available ELISA kits
Inc. (Greenwich, CT,
Seine,
by commercially
American
supplied by
Diagnostica
USA) and Diagnostica Stago, Ltd. (Asni6res sur
France) as previously described (12-15).
Statistical
analysis
The results were
as a mean
±
standard error
expressed
SE) of the mean.
The
of the differences was
(
analysis
performed by the
Student t-test.
A
P
value of<0.05
was
taken to be
significant.
RESULTS
‘
The results are summarized in
I, 2, and 3.
Figures
levels were
Both NO and TM
plasma
with COPDas
significantly
with con-
decreased in
patients
compared
trol
<
0.001) (NO
23.42
±
L67
subjects (p
patients:
controls: 40.0
±
338
TM
5.46
~
Lg/mL,
~g/mL:
patients:
±
1.3?
ng/mL, controls:
l?.9
±
0.5
(Fig. I and
ng/mL)
Fig. 2).
The selecting behavior showed a different pattern, as
sP-selectin
(79.42
±
18.20
was significantly higher
controls: 37.5 ± 2.84
ng/mL,
mg/mL)
(p
<
0.05),
controls:
whereas sl-selectin (584.9
054.02 ± f~l.~~
±
78.98
na/mL,
1
was
decreased (p
significantly
ng/mL)
<
.00!) as compared
with the control
3). ’In
group (Fig.
0
contrast, se-selectin (49.23
variation from the control
±
7.22 ng/mL) did not show
± 2.70
any
group
(44.50
ng/mL).
A
0.001) TFPI
significantly higher (~ ~
antigen
(patients: 112.28
ng/mL).
FIG. 2.
plasma
Setectin (sP: p)ate)ets, sL: teukocytes, sE:
endothelium) j;
level was found in our
ng/mL; controls 77.68
plasma
patients
± 0.28
levels (mean
t
se) expressed in ng/mL in patents with
in
j
j
±
6.45
chronic obstructive pulmonary disease (COPD) and
controls.
1
~
DISCUSSION
Chronic obstructive pulmonary disease (COPD) is ’a
!
li
t
pathologic condition
characterized by an endothelial cell
and/or
circu-
dysfunction of pulmonary artery
systemic
lation (4,11). Moreover, in this disorder,
thrombotic 11
complications may occur
in the
pulmonary
circulation
show altered endothelial cell and11
(
1-3). Several reports
hemostatic markers, such as endothelium-dependentpu!-
c
monaryartery relaxation,
nitric oxide, soluble P-selectin,
F I+2
fragment, fibrinogen, urinary
excre-1
prothrombin
FIG. 1, Nitric oxide plasma level (mean ± standard error) ex-
tion of
~i-thramba~~ol~~zlin; or platelet ir
pressed in nmol/mL in patients with
chronic obstructive pulmo-
ll-deydro-TxB2 ,
nary disease (COPD) and
in ccar~trr~l. subjects.
aggregates in patients with COPD(4-1(3~.
~
Downloaded from cat.sagepub.com at East Tennessee State University on May 27, 2015

207
in
with COPD who were in stable condition,
patients
suggesting that it might
considered
a
marker of in vivo
activation
Our
concur with their
platelet
(10).
findings
observation and with the reduced NOlevel, as NOis a
well-known
powerful inhibitor of platelet
activation.
E-selectin is found
on activated endothelium, in
only
contrast to otheradhesion molecules, which have a wider
tissue distribution. It is involved in the adherence of leu-
to the endothelium that have been
to
kocytes
postulated
an
role in atherosclerosis (29-31), and it
play
important
is
in vascular endothelial cells
by injury,
overexpressed
ischemia, cytokines, tissue factor, and thrombin (23).
FIG. 3. Thrombomodulin plasma level
± standard
(
mean
error)
et al. observed that sE-selectin levels were
Kobayashi
in ng/mL in patients with chronic obstructive pulmo-
expressed
.
nary disease (COPD) and in control subjects.
elevated during asthma attacks and decreased in stable
condition
In contrast with Riise et al., we did not
(32).
find
any
variation in sE-selectin
plasma levels in patients
Weusedselect endothelial cell-,
let-derived substances to evaluate their
and
leukocyte-,
plate-
with COPDas
with control
(5). How-
subjects
compared
expression in pa-
reduced
ever, this could be due to the different modes of therapy
used, as the majority their patients, in contrast to our
were not
tients with COPD. We found
significantly
levels ofnitric oxide andthrombomodulin in
/
plasma
pa-
patients,
taking any drugs.
tients with COPD. NO
powerful
produced by the endothelium is a
L-selectin, expressed on a wide variety of leukocytes,
vasodilator and inhibitor of
platelet aggrega-
plays a
role in the
migration
and sites ofchronic inflammation, and
of into acute
of
lymphocytes
into
periph-
tion, and TM is the endothelial surface site for binding
tnd inactivating thrombin with subsequent activation of
eral
lymph nodes
migration
sites (23,24). Our
‘
in the
leukocytes
inflammatory
C
( 6 17). The
of
NOin
protein
patients
though
present
some
production
with COPD is still controversial. In fact, al-
there is evidence that an
NOrelease is
endogenous
with COPDshowed
a
patients
signifi-
reduced level. At the time ofthis
study, patients in
cantly
impaired
vasculature,
our COPD
in stable condition,
anti-
group,
inflammatory drugs that
L-selectin (33). Moreover,
were taking
in the
there are
pulmonary
(4,11)
an increase in reactive
could reduced the
of
expression
reports showing
species production
nitrogen
Noguera et al. found in pa-
and exhaled NO(18-20). Our find-
tients with stable COPDthat L-selectin
was
expression
of
reduced NO levels are in
keeping
ings
significantly
hypothesis
similar to that of control
subjects (34).
with the
that reduced NOrelease can dimin-
Tissue factor
inhibitor
pathway
(TFPI), primarily syn-
cells ofthe microvasculature, is
inhibitor of the factor Vlla/tissue
:
sh vascular
and
and
the
reactivity
promote
perpetuate
so that
thesized
byendothelial
a
of the vascular wall
remodeling process
(11),
Kunitz-type protease
presence
direct inhibitor offactor Xa.
thrombotic
complications may occur in
the
pulmonary
factor
in
offactor Xa(35); it is also
Pulmonary epithelial lining
a
complex
circulation of
with COPD.
patients
Thrombomodulin levels were also found
significantly
cells
to
increased TFPI
appear
synthesize significantly
reduced in our
or
and
further contribute to the
patients,
may
under
conditions
In
with
inflammatory
(36,37).
fibrosis with extensive mes-
patients
of in situ thrombosis. In fact,
high
initiation
worsening
sarcoidosis and
pulmonary
,
levels of thrombomodulin are associated with a de-
we
demonstrated in-
may represent a
enchymal proliferation,
recently
ceased risk of
disease
coronary artery
(2!,22).
creased
levels of TFPI, which
plasma
compensatory
.
Selectins
endothelium-E, and
(
platelet-P,
leukocyte-
a
mechanism to
thrombosis
(38).
prevent
with other adhesion
~
L), together
molecules, play
key
This could also be true for
with COPD.
patients
in the
of inflammation,
immunologic
~
role
p~th~~et~es~s
Our data concur with the evidence of some form of
and of
abnor-
infections in the va~~ul~tur~, ~.nd thrombosis
i(23,24). They
responses,
endothelial cell
dysfunction
coagulation
are
in a
of disease
up-regulated
variety
malities
in
with COPD. However, be-
patients
present
states, and, in some instances, arcreleased into the cir-
cause the endothelium seems to
produce a
defense
where
can be detected in soluble form(sP,
Iculation,
they
mechanism, the potential usefulness ofan antithrombotic
¡
¡
sE, sL-selectin). P-s~l~ctirt, foundin the
a
granules ofthe
therapy in
these
as
suggested by Basili et
al.
patients,
and in the Weibel-Palade bodies ofendothelial
platelets
(
39), needs further investigation.
hasbeen shown to be elevated in thrombogenic
estates such as
infarction,
thrcrtn~c~cyt~peui~ purpura,
deep venous
throm-
myocardial
thrombotic
and
)bosis,
hepa-
This
was
from
A~krtc~w~eclgments:
study
supported bygrant
nn-induced
(25-28). Ferroni et al.
thrombocytopenia
the Ministero dcirUniverstta e della Ricerca Scientifiea
showed
levels of P-selectin
(MURST):
Progetto di
ricerca cofmaziato
immuno-
significantly higher plasma
Risposte
Downloaded from cat.sagepub.com at East Tennessee State University on May 27, 2015

2
08
The au-
20. Clini E, Bianchi L, Vitacca M, et al. Exhaled nitric oxide and
exercise in stable COPDpatients. Chest 2000;117:702.
biologice e fisiopatologiche dell’apparato respiratorio.
thors thank Prof. J. LoscalzoofBoston
ful eritici5m and advice in the
for his
University
help-
21. Salomaa V, Matei C, Aleksic N, et al. Solublethrombomodulin as
of this
preparation
manuscript.
a
incident
heart disease and
coronary
predictor of
symptomless
carotid artery atherosclerosis in the Atherosclerosis Risk in
Com-
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