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American Journal of Hematology 76:57–60 (2004)

Elevated Plasma sVCAM-1 Levels in Children With

Sickle Cell Disease: Impact of Chronic
Transfusion Therapy
Vishwas S. Sakhalkar,1* Sreedhar P. Rao,2 Jeremy Weedon,3 and Scott T. Miller 2
1
Department of Pediatrics, Children’s Hospital of Shreveport, Shreveport, Louisiana
2
Department of Pediatrics, State University of New York-Downstate Medical Center/Kings County Hospital Center, Brooklyn, New York
3
Scientific Computing Center, State University of New York-Downstate Medical Center/Kings County Hospital Center, Brooklyn, New York

Vascular cell adhesion molecule-1 (VCAM-1) has been implicated as being important in the
pathophysiology of acute pain episodes (APE) and acute chest syndrome (ACS) of sickle
cell disease (SCD). The frequency of these episodes is reduced by chronic transfusion
therapy. The impact of chronic transfusion therapy on VCAM-1 expression is unknown.
Soluble VCAM-1 (sVCAM-1) levels were measured in plasma using an ELISA assay (R&D
Systems) in 61 patients with SCD (age range 1.5–20 years) and 12 normal controls (2.5–14
years). SCD patients included 20 with ACS, 14 with APE, 12 at well-child visits, and 15
receiving chronic transfusion therapy. Asymptomatic SCD patients had higher sVCAM-1
levels compared to normal subjects (P < 0.001). Levels of sVCAM-1 were further elevated
during ACS (P < 0.001) and APE (P = 0.072) and returned to the asymptomatic range on
resolution. Levels were significantly lower in transfused patients (P = 0.003) compared to
asymptomatic SCD patients. Our findings of increased VCAM-1 expression during ACS and
perhaps APE offer a rationale for therapeutic use of cytokine and other VCAM-1 modula-
tors. The reduction of sVCAM-1 levels observed in our transfused SCD patients offers
insight into the mechanism of the protective effect of transfusion against ACS and APE
and possibly stroke. Am. J. Hematol. 76:57–60, 2004. ª 2004 Wiley-Liss, Inc.
Key words: vascular cell adhesion molecule-1; sickle cell anemia; acute pain episode;
acute chest syndrome; chronic transfusion therapy; white blood cell count

INTRODUCTION vation antigen-4 (VLA-4) on erythrocyte and leuko-

cyte membranes and VCAM-1 on the endothelium
The episodic and unpredictable acute pain episode
[3,6,7]. This increases capillary transit time, worsens
(APE) is the hallmark and the most common cause for
hypoxia, and finally leads to endothelial damage and
hospitalization among patients with sickle cell disease
vaso-occlusion [8]. Repeated endothelial injury and the
(SCD) [1]. Acute chest syndrome (ACS) is the second
erythrocyte/leukocyte/endothelial interaction mediated
most common cause of hospitalization and a leading
by VCAM-1 may play a role in the smooth muscle
cause of mortality and morbidity in SCD [2]. It is
hypertrophy and other microscopic vascular changes
believed that the manifestations of APE and ACS are
seen in SCD [8].
mediated in part by vascular cell adhesion molecule-1
(VCAM-1) upregulation in pulmonary and other vas-
cular endothelial cells [3,4]. This process is not counter- *Correspondence to: Vishwas S. Sakhalkar, M.D., Assistant
balanced by the release of cytoprotective mediators Professor of Pediatrics, LSUHSC, P.O. Box 33932, 1501 Kings
that normally inhibit this endothelial VCAM-1 up- Highway, Shreveport, LA 71130-3932. E-mail: vishwas@pol.net
regulation [5]. Hypoxia, among other factors, mark-
Received for publication 1 April 2003; Accepted 17 October 2003
edly enhances the adherence of sickle erythrocytes
and leukocytes to vascular endothelium by increased Published online in Wiley InterScience (www.interscience.wiley.com).
expression and interaction between the very late acti- DOI: 10.1002/ajh.20016
ª 2004 Wiley-Liss, Inc.
 10968652, 2004, 1, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/ajh.20016 by Readcube (Labtiva Inc.), Wiley Online Library on [22/10/2022]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
58 Case Report: Sakhalkar et al.

In vitro studies on vascular endothelium have con- Patients in the transfusion group were receiving
vincingly demonstrated up-regulation of VCAM-1 on chronic blood transfusion to prevent cerebrovascular
the endothelial surface to be associated with increased accident (CVA) or splenic sequestration (one patient)
production and secretion of sVCAM-1 [4]. Plasma for more than three months (median ¼ 29 months,
sVCAM-1 levels are thus indicators of endothelial mean ¼ 29.3 months, and range ¼ 4–60 months).
upregulation of VCAM-1 receptors. They all had sickle hemoglobin of less than 50% at
Chronic transfusion therapy, proven beneficial in the time of blood collection.
preventing cerebrovascular accidents [9], also decreases This study was reviewed and approved by the Insti-
the incidence of APE and ACS in these patients [10,11]. tutional Review Board for the protection of human
The purpose of this study was to further elucidate the subjects at University Hospital of Brooklyn and
role of VCAM-1 in SCD, especially regarding APE and Kings County Hospital Center. Blood was drawn
ACS, and also to examine the effect of chronic transfu- from patients and control subjects during venepunc-
sion on VCAM-1 expression in sickle cell patients. ture for other investigations after informed consent
was obtained; patient assent was also obtained where
MATERIAL AND METHODS appropriate.
Blood was collected in EDTA tubes and plasma was
Patients
separated within 30 min of collection by centrifugation
Sixty-one patients with SCD (age range 1.5–20 at 1,000g for 10 min at 4
C. All samples were stored at
years) and 12 normal controls (age range 2.5–14 80
C. A complete blood count was also checked from
years) were studied between October 1999 and May a simultaneously drawn blood sample.
2000 (Table I). Asymptomatic SCD children and age
matched normal subjects were randomly selected at Measurement of sVCAM-1 Levels in Plasma
routine well visits at the same hospital and specifically
Plasma levels of sVCAM-1 were measured using a
did not have a history of fever or signs or symptoms
commercially available ELISA Kit (R&D Systems,
of infection. Chronic transfusion patients were stud-
Minneapolis, MN) according to the manufacturer’s
ied during routine visits for transfusion; blood was
instructions.
drawn pre-transfusion.
APE was defined as an admission for pain requiring
Data Analysis
administration of parenteral opiates; only afebrile
patients were studied. ACS was defined as the devel- The number of SCD patients with ACS and APE
opment of a new infiltrate on chest radiograph in required to perform this analysis was estimated based
combination with fever, respiratory symptoms, or on numbers presented in previously published studies.
chest pain. Blood was obtained within 48 hr of admis- All 15 patients with SCD receiving chronic transfusion
sion and before therapeutic blood transfusion, if indi- and fit the study criteria were invited to participate, and
cated. Repeat samples were drawn after resolution of all agreed.
illness and at clinic visits 2 weeks to 2 months after One-way analysis of variance followed by Dunnett’s
discharge. None of the patients was studied for both test was used to compare sVCAM-1 levels in patients
APE and ACS concurrently, although one patient with receiving chronic blood transfusions and in normal
ACS was later readmitted with APE during the period controls with asymptomatic SCD patients. Tukey’s
of study; the pain episode was excluded from study. test was used to analyze differences among other groups.
Student’s paired t-test was used to compare sVCAM-1
TABLE I. Comparison of sVCAM-1 Levels in Different Groups of levels separately within ACS and APE groups during
Subjects* illness and on follow-up. Pearson correlation was used
to measure the association between sVCAM-1 levels
Mean ± 1
standard deviation and WBC counts.
No. of of sVCAM-1 P values (versus
Group subjects levels (ng/ml) asymptomatic SCD)
RESULTS
ACS 20 1727.5 ± 493.9 <0.001
APE 14 1383.9 ± 449.5 0.072 Distribution of patients and plasma levels of
Asymptomatic sVCAM-1 in different subject groups are depicted in
SCD 12 970.5 ± 369.8 Table I. Patients from the asymptomatic SCD group
Chronic had significantly higher sVCAM-1 levels compared to
transfusion 15 613.3 ± 244.9 0.003
Normal 12 227.1 ± 195.8 <0.001
normal subjects, and levels of sVCAM-1 were further
increased during ACS and APE. Plasma sVCAM-1
*ACS vs APE, P ¼ 0.087. levels were significantly lower in patients receiving

Case Report: sVCAM-1 Levels in Sickle Cell Disease
TABLE II. Comparison of sVCAM-1 Levels Within the Two
Groups of Patients With Paired Follow-up Samples
Mean ± 1
standard deviation
No. of of sVCAM-1 P values (within
Group subjects levels (ng/ml) subgroups)
ACS 12 1710.7 ± 487.5 <0.001
Resolved ACS 12 893.8 ± 329.3
APE 7 1383.9 ± 449.5 0.095
Resolved APE 7 810.7 ± 211.1
chronic blood transfusions than in the asymptomatic
sickle patients, though they did not reach levels seen
in non-sickle cell controls.
The elevated sVCAM-1 levels among patients with
APE or ACS were in the ‘‘asymptomatic’’ range on
follow-up (Table II). No bacterial etiology was found
as the cause of ACS. Pearson correlation across all
patient groups revealed that sVCAM-1 levels were
directly proportional to total WBC counts (P ¼ 0.016)
from simultaneously drawn blood samples. All patients
had a neutrophil predominance; there was no statistical
correlation of sVCAM level with absolute numbers of
neutrophils, lymphocytes, or monocytes.
DISCUSSION
The role of vascular endothelial cell receptors in the
pathophysiology of APE and ACS of SCD has recently
become a subject of intense interest. It is well known that
hypoxia leads to irreversible polymerization of sickle
hemoglobin [12], and increased capillary transit time
leads to further polymerization and ultimately to APE
and ACS. Hebbel et al. [13] observed that increased
capillary adherence of sickle erythrocytes contributes
to increased capillary transit time. VCAM-1 and CD36
receptors on the endothelial surface and their counter-
parts VLA-4 and CD36, respectively, on the erythrocyte
membrane are thought to be important factors facili-
tating the adhesion process [5,14]; reticulocytes are
particularly adherent [6]. VCAM-1 and intercellular
adhesion molecule-1 (ICAM-1) similarly influence leu-
kocyte adhesion to endothelial cells [4]. In vitro studies
on vascular endothelium have convincingly demon-
strated up-regulation of VCAM-1 on the endothelial
surface with concomitant increased production and
secretion of sVCAM-1 on interaction of endothelium
with sickle erythrocytes under conditions of shear stress
or in the presence of cytokines such as interleukin 1b [4].
There are two clinical studies investigating sVCAM-1
levels in sickle cell patients, with ACS [5] and APE [15],
respectively. We confirm earlier findings that sVCAM-1
levels are elevated in asymptomatic SCD patients
and are further increased in ACS and perhaps in APE
10968652, 2004, 1, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/ajh.20016 by Readcube (Labtiva Inc.), Wiley Online Library on [22/10/2022]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
59
(though P ¼ 0.07), and that these levels return to steady-
state values on resolution of the clinical episode. These
findings affirm the likely role of VCAM-1 in SCD
pathophysiology and provide rationale for use of cyto-
kine and VCAM-1 modulators (e.g. dexamethasone
[16,17] and sulfasalazine [18]) in treatment of SCD
complications.
The impact of transfusion on sVCAM-1 levels has
not been reported previously. Lower sVCAM-1 levels
documented in our SCD patients on chronic blood
transfusions compared to even asymptomatic SCD
patients offers insight into the observed protection
conferred by chronic transfusion against ACS and
APE [10,11]; presumably, reduction in the percentage
of sickle cells with replacement by cells containing
normal hemoglobin leads to diminished expression
of VCAM-1 and reduced tendency toward initiation
of these pathologic events. It also provides a possible
explanation for the stabilization of cerebrovascular
disease in chronically transfused patients [19]. Dilu-
tion of sickle cells by transfusion presumably attenu-
ates abnormal adhesion and thus prevents further
vascular damage.
The observed correlation of raised WBC counts with
higher sVCAM-1 levels in SCD has not been documen-
ted before. It could be due to several factors. The high
WBC counts in our admitted patients could be related
to inflammation or due to simultaneous increase of
cytokines and other markers including sVCAM-1 [5]
or a combination of more than one of these factors.
High steady-state WBC counts in SCD patients are
associated with recurrent pain [20], acute chest syn-
drome [21], stroke [22], early death [23,24], and other
adverse events [25]. It may be that abnormal expression
of adhesion factors such as VCAM-1 are involved in
this negative association.
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