Coagulation Changes in Individuals With Sickle Cell Trait
Coagulation Changes in Individuals With Sickle Cell Trait
Coagulation Changes in Individuals With Sickle Cell Trait
DOI 10.1002/ajh.10021
Sickle cell disorders, such as Hb SS and Hb SC, are associated with a hypercoagulable
state that may contribute to the vaso-occlusive episodes observed in the disorders. To
what extent increased coagulation activity occurs in individuals with sickle cell trait has
had limited study. Because such information may help clarify clinical and pathologic
®ndings that may occur in these individuals and may be useful in clarifying the hyper-
coagulable state in sickle cell disease, we have examined individuals with Hb AS to
determine the extent that increased coagulation activity does occur. We measured
d-dimers, thrombin±antithrombin (TAT) complexes, prothrombin fragment 1.2 (F1.2),
absolute blood monocyte levels, proteins C and S, and isotypes of antiphospholipid
antibodies in individuals with Hb AS and in matched controls (Hb AA). Results showed
that d-dimers, TAT, and F1.2 were increased signi®cantly above normal levels. Absolute
blood monocyte levels were increased. The d-dimers, TAT, F1.2, and monocyte counts
showed signi®cant increasing trends through groups of increasing severity (Hb AA, Hb
AS, Hb SC, and Hb SS). Our study shows that individuals with Hb AS have increased
coagulation activity, with d-dimers, TAT, and F1.2 being consistent indicators. The
measures of coagulation activity in Hb AS are lower than in patients with Hb SC and Hb
SS disease. These results extend our previous observation that the degree of coagulation
activation parallels the degree of disease severity among sickle cell genotypes. The
®ndings suggest that monocytosis, with the possible expression of monocyte-derived
tissue factor, and the associated hypercoagulable state are driven by disease severity.
Am. J. Hematol. 69:89±94, 2002. ã 2002 Wiley-Liss, Inc.
matched for age, gender, and race, were examined. Protein S activity was measured by a clot-based
The individuals were 20 73 years of age and included analysis of dilute plasma with buer (1:5) and prein-
19 women and 4 men. The controls and patients were cubated with added activators and protein S de®cient
randomly selected and consisted of laboratory and plasma (Bioclot Protein S kits, Biopool, Ventura, CA)
non-laboratory personnel. Technicians performing [6]. Protein S antigen (total, free) was analyzed by
the laboratory assays were blinded as to the origin of EID on antibody-coated agarose plates (Assera-plate
all samples. The study was approved by the Institu- PS antigen kindly supplied by Diagnostica Stago).
tional Review Board of Mount Sinai Hospital, and Total protein S antigen was determined by applying
informed consent was obtained from the participants. native plasma to the plate. Free protein S (unbound
to C4b-BP of complement) was also determined by
EID after separating the free from the bound protein
S. This was accomplished by cold precipitation of the
Assays of Coagulation Activation bound protein S after mixing plasma (13:1) with 25%
Measures of coagulation activation included d-di- PEG (polyethylene glycol), centrifuging and separat-
mers, thrombin antithrombin complexes (TAT), and ing the supernatant as free protein S.
prothrombin fragment 1.2 (F1.2). The d-dimers were Absolute blood monocyte counts were calculated
assayed by ELISA (American Bioproducts Co.; Di- from the percentage of monocytes present in total
agnostica Stago, Asnieres-sur-Seine, France), TAT leukocyte counts. Urine osmolality was determined
was determined by ELISA (Dade Behring, Chicago, by freezing point depression (Advanced Instruments,
IL), and F1.2 was assayed by ELISA (Dade Behring), Norwood, MA) on overnight fasting, ®rst morning
as previously described [3]. specimens.
average trend or the eect of a unit increase in the TABLE I. Clinical Data on Individuals With Sickle Cell Trait
assigned score. Correlations were computed between Hb HbS MCV Urine osmolality
d-dimers and monocyte counts, both unadjusted and Age Gender (g/dl) (%) (¯) (mOsm/kg H2O)
adjusted for group scores; between group scores and
monocyte counts, both unadjusted and adjusted for d- 1. 30 F 13.5 42 88 546
2. 34 F 13.9 42 89.5 592
dimers; and, between group scores and d-dimers, both 3. 31 F 13.3 37 79.4 427
unadjusted and adjusted for monocyte counts [7]. 4. 46 F 13.9 42 93.4 247
5. 46 F 13.3 41 93.9 238
6. 28 M 15.1 42 94.4 801
RESULTS 7. 20 M 14.7 43 91 778
8. 72 M 14.5 40 87.4 469
Clinical data are shown in Table I. All participants 9. 27 F 12.8 26 72 787
10. 43 F 12.4 41 87.5 462
in the study were healthy with no evidence of disease. 11. 56 F 13.2 40 85 467
Two individuals (#13, #22) had a past history of gross 12. 50 M 12.9 41 87.5 439
hematuria on one occasion. None of the individuals 13. 58 F 13.5 40 89.5 457
with sickle cell trait had present or past history of 14. 43 F 10.8 41 92 412
pulmonary embolism or leg ulcers. Ten individuals 15. 37 F 11.1 42 82.7 388
16. 28 F 11.4 43 93 NDa
(#14 #23) had moderately decreased hemoglobin 17. 50 F 11.7 40 94.6 817
levels (10.4 11.8 g/dl). The individuals with de- 18. 47 F 10.8 38 88.6 338
creased hemoglobin levels had no history of preg- 19. 72 F 11.8 40 79 466
nancy for at least 2 years, no clinical or laboratory 20. 25 F 11.7 39 80.9 472
evidence of iron de®ciency, and no reported use of 21. 73 F 11.4 42 83 432
22. 37 F 10.4 31 81 472
oral contraceptives during the period of the study. 23. 30 F 10.5 39 81 674
The decrease in hemoglobin levels in this group may
a
be related to several factors such as co-existent ND, not done.
a-thalassemia [8], aging, race, or gender [9].
Median d-dimer, TAT, and F1.2 levels were sig- Absolute blood monocyte counts were lowest in the
ni®cantly increased in the individuals with sickle cell controls and were progressively higher in individuals
trait (Table II). There was no correlation between d- with Hb AS and in patients with Hb SC and Hb SS
dimer, TAT, and F1.2 and the hemoglobin concen- (controls, 398 [274] ´ 109 cells/l; median [IQR]; Hb
tration, whether normal or decreased. Similarly, the AS, 476 [301]; Hb SC, 613 [ 691]; Hb SS, 949 [867]).
levels of Hb S and urine osmolality did not correlate Pairwise comparisons between groups were not sig-
with the levels of coagulation activity. The d-dimer, ni®cant at P < 0:05. The plots in Fig. 2 show the
TAT, and F1.2 levels in individuals with Hb AS were group median values for monocyte counts plotted
less than those previously described in patients with against the group median values for d-dimers and
Hb SC and Hb SS disease (Table II, Fig. 1) [3]. re¯ect a near-linear relationship across the group
TABLE II. Measures of Coagulation Activation in Individuals With Sickle Cell Trait, Patients With Hb SC Disease, Patients With Hb
SS Disease, and in Controlsa
D-dimer (lg/ml) 0.46 (0.27) [n = 23] 0.30 (0.25) [n = 23] 0.81 (0.70) [n = 16] 1.26 (1.63) [n = 18] 0.2 (0.2) [n = 17]
P 0.0005*, 0.0013**; 0.0792b
0.0002***
TAT (ng/ml) 2.8 (2.5) [n = 23] 1.5 (1.0) [n = 23] 4.1 (3.7) [n = 15] 6.8 (8.1) [n = 18] 2.4 (2.35) [n = 18]
P 0.0072*, 0.0379**; 0.0203b
0.0003***
F1.2 (nm) 0.6 (0.5) [n = 23] 0.5 (0.5) [n = 23] 2.3 (1.8) [n = 15] 2.6 (1.3) [n = 18] 0.76 (0.76) [n = 17]
P 0.0266*, 0.0004**; NSb
0.0002***
a
Values are expressed as median (inter-quartile range); values for Hb SC and Hb SS were obtained from Ref. 3; NS, not signi®cant at <0.05 level;
n = number of subjects.
b
Hb SC v Hb SS.
*Sickle trait v control.
**Sickle trait v Hb SC.
***Sickle trait v Hb SS.
92 Westerman et al.
Fig. 1. Levels of d-dimers, TAT, and F1.2 in patients with Hb SS disease, Hb SC disease, in individuals with sickle cell trait
and in controls. (IÐ-I) Median values for Hb SS, Hb SC, and Hb AS. (OÐ-O) Median values for controls.
means, with the groups ordered in terms of increasing adjusting for monocyte counts, was r 0:78 (P <
disease severity. Plots of TAT and F1.2 did not show 0:0001, n 57). The correlation coecient for the
a similar trend. trend in monocyte counts by disease severity score,
Increasing trends in d-dimers and monocyte counts adjusting for d-dimers, was r 0:33 (P 0:013, n
computed from all disease groups combined showed 57). The association between d-dimers and monocyte
an apparent association between d-dimers and counts was not signi®cant after adjusting for group.
monocyte counts without adjusting for group (r Antiphospholipid antibodies, protein C (activity,
0:27, P 0:04, n 57). The correlation coecient for antigen), and protein S (activity, total, free antigen) in
the trend in d-dimers by disease severity score individuals with Hb AS were in the normal range
(0 Hb AA, 1 Hb AS, 2:5 Hb SC, 3 Hb SS), (results not shown).
Coagulation Changes in Sickle Cell Trait 93