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Leukocytosis and Ischemic Vascular Disease Morbidity and Mortality

Is It Time to Intervene?
Barry S. Coller From the The Rockefeller University, New York, NY. Correspondence to Barry S. Coller, The Rockefeller University, 1230 York Ave, New York, NY 10021. E-mail collerb@rockefeller.edu

Abstract
The association between leukocytosis and increased morbidity and mortality of ischemic vascular disease has been observed for more than half a century, and recent studies in >350 000 patients conrm the robustness of the association and the dramatically higher relative and absolute acute and chronic mortality rates in patients with high versus low leukocyte counts. Although there is reason to believe that the association is not causal (that is, that leukocytosis is simply a marker of inammation), there is also reason to believe that the leukocytosis directly enhances acute thrombosis and chronic atherosclerosis. Leukocytosis also is associated with poor prognosis and vaso-occlusive events in patients with sickle cell disease, and experimental data suggest a direct role for leukocytes in microvascular obstruction. The only way to test whether leukocytes contribute directly to poor outcome in ischemic cardiovascular disease is to assess the eect of modifying leukocyte function or number. Because selective blockade of leukocyte integrin M2 and P-selectin have thus far been disappointing as therapeutic strategies in human

cardiovascular and cerebrovascular disease, I discuss the potential risks and benets of short-term treatment with hydroxyurea to decrease the leukocyte count in select populations of patients at the highest risk of short-term death. Key Words: leukocytes neutrophils myocardial infarction thrombosis

Introduction
In the 1949 edition of his classic text, Dr Paul Dudley White wrote (without a reference) that severe and sustained leukocytosis after

1 myocardial infarction was associated with poor prognosis, and in 1954, Cole et al reported that >32% of patients with an acute myocardial infarction and a white blood cell (WBC) count of >15 000/L died within the rst 2 months after the infarction, whereas <9% of patients with a WBC count <10 000/L died within this

3 8 years. Comparison of individuals with WBC counts in the highest third with those in the lowest third yielded a coronary heart disease risk ratio of 1.5 (95% CI, 1.4 to 1.6) even though the majority of these studies made adjustments for smoking and other known risk factors (Figure 1). In fact, the risk ratio exceeded 1.0 in each of the 19 studies. Since then, additional studies of WBC count and vascular disease morbidity and mortality involving >350 000 patients treated with more modern therapies have been reported, and in nearly all of these studies, a signicant relationship between leukocyte count and vascular disease morbidity and mortality was observed (Table

2 same period. Subsequent studies of the relationship between leukocytosis and coronary artery disease conducted from 1974 to 1996 were summarized in a meta-analysis of 19 prospective studies involving 7229 patients with a weighted mean follow-up of

1). 427

Figure View larger version: In this window In a new window Download as PowerPoint Slide Figure 1. Prospective studies of leukocyte count and coronary heart disease. Risk ratios compare top and bottom thirds of baseline measurements. Black squares indicate the risk ratio in each study, with the square size proportional to the number of cases and the horizontal lines representing the 99% CIs. The combined risk ratio and its 95% CI are indicated by unshaded

diamonds for subtotals and by shaded diamonds for grand totals. +adjustment for age and sex only; ++for these plus smoking; +++for these plus some other standard vascular risk factors; ++++for these plus markers of social class; and +++++for these plus information on chronic disease at baseline. Reprinted with permission from Danesh et al. View this table: In this window In a new window TABLE 1. Summary of Results of Selected Studies Assessing the Association Between Leukocyte Count and Morbidity and Mortality of Ischemic Vascular Disease 19992004 Strong arguments can be made that despite the robustness of the association, leukocytosis is only a nonspecic marker of other processes that actually cause the increased risk. Thus, it has been known for more than half a century that acute myocardial infarction is commonly accompanied by transient leukocytosis as
3

part of an acute phase reaction. 1 Moreover, there is increasing recognition that atherosclerosis has an important inammatory

28,29 and that cardiovascular risk factors such as component smoking and elevated body mass index (BMI) are associated with 30 Thus, it is reasonable to consider that chronic inammation. the elevated WBC count is just a marker of the chronic inammatory state and that other aspects of inammation may be the direct cause of the vascular disease.

However, there are a number of potential mechanisms by which leukocytes may contribute directly to acute and chronic ischemic vascular disease, and although there is insucient space to discuss these mechanisms in detail, they are listed in Table 2 with select references. This review summarizes the recent epidemiologic evidence of an association of WBC count with acute and chronic vascular disease, with a particular focus on data that may bear on the question of a potential causal role of leukocytes in vascular events, and then addresses the question of whether the existing data justify performing studies to assess the safety and ecacy of reducing the leukocyte count medically as part of the therapy of acute or chronic vascular disease. View this table: In this window

In a new window TABLE 2. Mechanisms by Which Leukocytosis May Predispose to Ischemic Vascular Events

Association of Leukocytosis at the Time of Onset of anis Acute Vascular Event with Short-Term There abundant evidence that relative leukocytosis at the time of admission and for either a cardiovascular or cerebrovascular event Morbidity Mortality

25 and 7.7%, 9 with the higher mortality in the latter group 4.4% most likely reecting the higher age of those enrolled in that study. In sharp contrast, the in-hospital mortalities for those in the highest WBC count groups were 15.9% and 27.3%, respectively, representing strikingly large absolute short-term mortality dierences in these studies of 11.5% and 19.6%. A number of other smaller studies listed in Table 1 reported similar ndings.

correlates with the severity of ischemic damage or subsequent course. For example, in the 2 largest studies of acute myocardial infarction, in which data from a total of 268 486 patients were analyzed, the in-hospital mortality rates for those in the lowest WBC count groups (lowest quartile and quintile, respectively) were

A J-shaped relationship between mortality and leukocyte count was observed in the large registry study by Grzybowski et al
25

9 conducted by Barron et al, patients with leukocyte counts <500/L were excluded, and this may have reduced the likelihood of identifying a J-shaped relationship. It is possible that subgroups with very low WBC counts and higher mortality may have been missed in other studies analyzing only quartiles of leukocyte count, but some studies did not nd this association even when

20 a very low WBC count may indicate general poor health and al, thus confer additional mortality risk independent of the coronary artery disease. Inclusion of such a subgroup in the analysis might dilute the association between the WBC count and cardiovascular risk in the remaining population. In the other large study,

(Figure 2) and in 2 other registry studies, 21,31 wherein patients with the lowest leukocyte counts appeared to be a subgroup with higher mortality than the next highest WBC count group, and the remaining patients demonstrated a nearly linear relationship between leukocyte count and mortality. As suggested by Gurm et

specically looking for it.

20

20 is that the apparent inconsistency suggested by Gurm et al J-shaped curve is more likely to be observed in studies using

One likely explanation for this

registry data of unselected patients clinical trial data

20

21,25,31

rather than in

because patients with low WBC counts and

other illnesses are more likely to be excluded from clinical trials. Figure View larger version: In this window In a new window Download as PowerPoint Slide Figure 2. In-hospital mortality by WBC count in 115 273 patients in the National Registry of Myocardial Infarction 4. Note the J shape of the data with patients with very low WBC counts constituting a subgroup with higher mortality. Modied with permission from Grzybowski et al.
25

Data bearing on the mechanism of the association of leukocyte count and short-term morbidity and mortality were provided by Barron et al, who reported on pooled data on 992 patients from the Thrombolysis in Myocardial Infarction (TIMI) 10A and 10B trials of tenecteplase and reteplase treatment of acute myocardial infarction. WBC counts were obtained at entry, before drug administration. Higher WBC counts were associated with resistance to thrombolysis, increased thrombus burden, and impaired microvascular perfusion. Strikingly, 30-day mortality was 0% in patients with WBC counts 5000/L and 10.4% in patients with WBC counts >15 000/L, and a similar dierence was noted for the development of congestive heart failure or shock (Figure 3). Careful analysis of the totality of the data led the authors to conclude that an elevated leukocyte count was associated with a poorer response to therapy, not just a larger infarct at presentation. In fact, even after adjusting for admission TIMI ow and myocardial perfusion grades, anterior myocardial infarction location, baseline and maximum creatine kinase (CK) levels, smoking status, and a number of other factors, WBC count still demonstrated a strong trend toward an independent association with the development of new congestive heart failure and death (odds ratio, 1.21; P=0.07). Figure View larger version: In this window In a new window Download as PowerPoint Slide Figure 3. Association between WBC count and development
6

within 30 days of death or either congestive heart failure (CHF) or shock in 975 patients in the TIMI 10A and 10B trials.
6

The association of WBC count with response to therapy may, however, be dierent in stroke than in myocardial infarction. Thus, when Kammersgaard et al studied 763 unselected patients with acute stroke, they found that a higher leukocyte count was associated with a larger lesion at presentation (45 versus 34 mm; P<0.001) and more severe presenting symptoms (Figure 4; P<0.00001), but after adjusting for initial stroke severity, a higher leukocyte count was not associated with a poorer outcome. Although more data are required to unequivocally establish whether there is a dierence in the correlation between leukocytosis and response to therapy in myocardial infarction and stroke, and several dierent factors may account for the observed dierence (eg, ecacy of therapy, relative abilities of the heart and brain to withstand infarction), from a practical standpoint, the rationale for intervening to reduce the leukocyte count at presentation appears to be stronger in acute myocardial infarction than stroke. Figure View larger version: In this window In a new window Download as PowerPoint Slide Figure 4. Association of mean WBC count and initial stroke severity in 763 patients stratied into 4 groups according to their initial scores on the Scandinavian Stroke Scale (mild 58 to 44; moderate 43 to 29; severe 28 to 15; and very severe 14 to 0). P<0.0001. Modied with permission from Kammersgaard et al.
5

Eects of Reperfusion Therapy or Revascularization on Association Between When comparative analyses were conducted within a single study, the association between mortality after myocardial infarction and Leukocytosis and Ischemic Vascular Disease leukocyte count was less dramatic in patients who underwent Morbidity and Mortality
reperfusion or early revascularization treatment than in those who did not receive these therapies. Thus, in the National Registry of Myocardial Infarction 4 (n=115 273), the mortality odds ratio (comparing the highest to the lowest quartile of leukocyte count) was 4.0 for those not undergoing reperfusion therapy (n=83 775),

and the absolute in-hospital mortalities were 18.6% and 4.7%, respectively, for the highest and lowest WBC count quartiles, whereas the comparable odds ratio for those undergoing reperfusion therapy with thrombolytic agents or primary percutaneous coronary intervention (PCI; n=30 954) was 2.7 (9.3% versus 3.5%). These data yield absolute dierences in mortality between the highest and lowest WBC count quartiles of 13.9% for those not receiving reperfusion therapy and 5.8% for those

25 In the Platelet Glycoprotein IIb/IIIa in receiving such therapy. Unstable Angina Receptor Suppression Using Integrilin Therapy (PURSUIT) study (n=10 480), which included patients with acute coronary syndromes, the 6-month mortality odds ratio comparing the highest to lowest quartiles of leukocyte count was 2.6 (10.5% versus 4.1% mortality, respectively; P<0.001) in those who did not undergo PCI or surgical revascularization therapy (n=6791) during their index hospitalization, and 1.0 (4% versus 4%) for those undergoing revascularization, yielding absolute mortality 18 (Figure 5) Thus, dierences of 6.4% and 0%, respectively. regardless of the mechanism underlying the association between leukocytosis and mortality after acute myocardial infarction, it appears that modern reperfusion and early revascularization strategies mitigate or eliminate the association between WBC count and mortality during the rst 6 months after a myocardial infarction. These data suggest that patients who are not eligible for, or who do not receive, reperfusion or revascularization therapy may be a more appropriate group in which to test the eect of interventions to lower the WBC count.

Figure View larger version: In this window In a new window Download as PowerPoint Slide Figure 5. Mortality through 6 months as a function of the WBC count by quartile in patients in the PURSUIT trial (n=10 480) who did not undergo in-hospital revascularization (top; 62%) or did undergo in-hospital revascularization (bottom; 38%). Note dierences in ordinate scale in the 2 graphs. Modied with permission from Bhatt et al.
18

Leukocyte Count Association With Perioperative Mortality and Cerebrovascular The mortality associated with coronary Accidents artery bypass surgery as a

function of preoperative leukocyte count was studied in 2057

17 When assessed by increasing quartiles patients by Bagger et al. of leukocyte count, the 30-day mortality was 1.7, 2.7, 3.3, and 7.5% (P<0.0001 for trend). Even after excluding patients with recent myocardial infarction or unstable angina, or patients with conditions that may have predisposed them to elevated leukocyte counts, the association remained signicant. In other surgical studies by Dacey et al (n=11 270) and Albert et al (n=7483), preoperative leukocyte counts correlated with in-hospital mortality or perioperative cerebrovascular accidents.

WBC Count As a Risk Factor for Long-Term Ischemic Cardiovascular Disease In addition to the association of leukocyte count with short-term

morbidity and mortality from vascular disease, there are impressive data on the association of elevated leukocyte count with long-term vascular disease morbidity and mortality. In the Caerphilly and Speedwell population studies, men between the ages of 45 and 63 in the United Kingdom were evaluated and then
27 A total of 4325 completed the followed for the next 10 years. study, of whom 1005 had a history of vascular disease at the time of entry into the study. At the end of the study, 525 (12.2%) had experienced a myocardial infarction (silent, nonfatal, or fatal). After adjusting for age and investigation site, the relative odds of having a myocardial infarction comparing the highest to lowest quintiles of leukocyte count was 2.79 (95% CI, 2.06 to 3.77). The corresponding relative odds values for having an event for values of total cholesterol, triglycerides, and high-density lipoprotein (HDL) cholesterol (comparing lowest to highest quintiles in the latter) were all lower than that for leukocyte count (2.07 [1.55 to 2.78], 2.72 [1.98 to 3.72], and 2.34 [1.72 to 3.19], respectively). By multivariate analysis, the leukocyte count retained its association with adverse events, and the association was strengthened by adjustment for the imprecision of using only a single leukocyte value (regression dilution bias).

One serious limitation of the studies that used the WBC count at the time of admission for an acute myocardial infarction is the short-term impact of the infarction on WBC count. That is why the Multicenter Diltiazem Postinfarction Trial is of particular interest because patients were enrolled into the study and had their WBC counts analyzed 6 months after their myocardial infarction. Data were obtained on 1294 patients followed for a mean duration of 25 months (range 4 to 49 months). The mean leukocyte count at enrollment in those who went on to experience reinfarction or death was 81002400/L versus 76003200/L for patients who did not experience an event (P<0.001).

12 Regression dilution bias was minimized in this study by deaths. using the mean of 7 WBC counts obtained at intervals over 42 months (Table 1). In those with leukocyte counts <6000/L, the mortality was 6.9%, whereas in those with leukocyte counts 9000/L, the mortality was 17.7%, yielding an absolute dierence of 10.8%. Investigators used their data to create an assessment risk score and concluded that the leukocyte count was an independent risk factor for death. Relative to having a leukocyte count of <7000/L, having a count 9000/L was assigned 6 risk score points in men and 8 risk score points in women. For comparison, having left ventricular dysfunction was assigned 7 risk score points in both sexes, having diabetes was assigned 7 points in women and 3 points in men, and having a serum HDL 35 mg/dL compared with >55 mg/dL was assigned 6 points in women and 5 points in men.

In the GISSI-Prevenzione study on myocardial infarction survivors, 11 324 patients with an acute myocardial infarction in the previous 3 months, and with a good short-term prognosis, were studied for a 4-year period, during which there were 1071

Grau et al analyzed leukocyte data on 18 558 patients in the Clopidogrel versus Aspirin in Patients at Risk of Ischemic Events (CAPRIE) study, which enrolled patients with a history of ischemic stroke, myocardial infarction, or peripheral arterial disease. Patients were randomized to clopidogrel or aspirin and followed for 1 to 3 years. Baseline leukocyte counts were used unless they were obtained within 28 days after a myocardial infarction or ischemic stroke, in which case, samples obtained 4 weeks after entry were used. Patients were excluded from the study if they had leukocyte counts <3500/L or >25 000/L. During a mean follow-up period of 1.9 years, 1840 patients (9.9%) experienced ischemic stroke, myocardial infarction, or vascular death. In multivariate analysis, after adjusting for vascular risk factors and diseases, hematocrit, and study treatment, the relative risk of recurrent ischemic events was signicantly higher in patients in the highest quartile of leukocyte counts compared with those in the lowest quartile (1.42; 95% CI, 1.25 to 1.63 overall; 1.30, 1.56, and 1.51 for stroke, myocardial infarction, and vascular death). Thus, in addition to supporting an association of WBC count and myocardial infarction and vascular death in a high-risk population, this study established a relationship between leukocyte count and stroke. Data from the Studies Of Left Ventricular Dysfunction (SOLVD) trials, which enrolled stable patients with left ventricular dysfunction are of interest because the strong association of WBC count with cardiovascular mortality observed in the whole group, even after multivariate analysis (P<0.001), was found to be conned exclusively to the group in which ischemic vascular
24

disease was responsible for the ventricular dysfunction. 4 This observation supports a selective contribution of leukocytosis to the chronic vascular changes or the acute thrombotic phenomena that contribute to ischemic vascular disease.

Acute Changes in Leukocyte Count Before Onset of an Ischemic Separate from the Vascular associationEvent between chronically elevated

24 In fact, leukocyte counts the potential toxicity of clopidogrel. obtained by chance 7 days before the onset of a new vascular event were signicantly higher than the baseline counts in these same patients (n=211; P=0.004), whereas counts obtained anywhere from 8 to 120 days before a new event (and the last counts obtained in patients who did not sustain a recurrent event) did not dier signicantly from the patients baseline values. (Figure 6) Although these intriguing data are consistent with the hypothesis that the rise in leukocyte count contributed to the recurrent event, many other interpretations are also possible. Additional clinical observations are also consistent with an association between transient elevations in leukocyte count and acute vascular events because a number of studies have noted an association between recent infection and the onset of myocardial infarction or acute stroke, even though an association with

leukocyte counts and increased risk of ischemic vascular events is the issue of whether transient elevations in the WBC count are associated with an increased risk of sustaining an acute event. The design of the CAPRIE study allowed this question to be addressed because leukocyte counts were performed frequently to monitor

leukocyte count was not specically studied. Figure View larger version: In this window In a new window Download as PowerPoint Slide

3234

Figure 6. Dierences in leukocyte counts in 18 293 patients in the CAPRIE study between the baseline value and the last measurement before a recurrent event, or at the end of the study for those not having a recurrent event. Modied with permission from Grau et al.
24

Granulocyte colony-stimulating factor (CSF) infusions have been well tolerated by individuals donating peripheral stem cells and in

some stable patients with myocardial infarctions when given alone

3537 However, or as part of peripheral stem cell infusions. granulocyte CSF (G-CSF) and granulocytemacrophage CSF (GM-CSF) treatments have been associated with isolated cases of microvascular and macrovascular venous and arterial thrombosis

in patients with malignancies or hematologic disorders, and G-CSF treatment has been associated with both progression to myocardial infarction in patients with intractable angina and increased risk of restenosis in patients undergoing PCIs. The expanding use of stem cell strategies, including those that involve autologous stem cell mobilization with growth factors that can elevate the WBC count, make it important to obtain additional data on the safety of these agents in patients with cardiovascular disease.
36,37,40

3841

Evaluation of Ischemic Vascular Event Risk by Type of Leukocyte Most studies of the association of leukocytosis and ischemic

24 (Figure 7). In the Evaluation of c7E3 for Prevention of events Ischemic Complications (EPIC) study, the total WBC count correlated best with 3-year mortality, but the neutrophil count

vascular disease recorded a total WBC count without a leukocyte dierential analysis, and thus, there are relatively few data on the specic cell type most associated with the increased risk. In the CAPRIE study, elevations in neutrophils and to a lesser extent monocytes showed signicant associations with the risk of recurrent ischemic events, but elevations in lymphocyte counts showed, if anything, an inverse relationship to the risk of recurrent

20 The monocyte count was not also showed a correlation. correlated with outcome, and as in the CAPRIE study, the lymphocyte count demonstrated an inverse relationship. Similarly, in the SOLVD trials, total WBC and neutrophil counts were signicantly associated with cardiovascular death, even after multivariate adjustments, and once again, a strong trend was observed for an inverse relationship between the lymphocyte

count and cardiovascular mortality. Figure View larger version: In this window In a new window Download as PowerPoint Slide

Figure 7. Univariate association between quartiles of leukocyte count, neutrophil count, monocyte count, and lymphocyte count and risk ratio for annual rate of recurrent ischemic vascular events (ischemic stroke, myocardial infarction, and vascular death) in 18 558 patients in CAPRIE. expressed relative to lowest quartile.
24

Risk ratios are

Monocyte counts were monitored daily for 4 days in a study of

149 patients with Q-wave myocardial infarction. 15 In multivariate analysis, a peak monocyte count 900/L was found to be an independent determinant of cardiac pump failure (relative risk, 9.83; P<0.0001) and cardiac events (death, recurrent myocardial infarction, and readmission for heart failure; relative risk, 6.30; P<0.0001). Thus, among the leukocytes, morbidity and mortality is most closely associated with the neutrophil count, but the association with the monocyte count may be underestimated because of greater regression dilution bias (see below). Monocyte production of tissue factor is well established and can be enhanced by activation, providing a direct link to thrombosis; the importance of

4244 It is neutrophils as a source of tissue factor is less certain. notable that an inverse relationship between mortality and lymphocyte count was observed in all 3 studies, even though the relationship was not as strong as the direct associations involving total WBC count and neutrophils. Because lymphocyte counts are depressed by elevated glucocorticoid levels, even during the

45 it is possible that the lower lymphocyte counts in diurnal cycle, patients with a poorer prognosis reect elevated glucocorticoid levels. In fact, relative lymphocytopenia in patients with chest pain was found to be similar to rapid creatinine kinase-MB in diagnosing a myocardial infarction (sensitivities of 58% and 56% for lymphocytopenia and elevated rapid creatinine kinase-MB, respectively; specicities of 91% and 93%); elevated cortisol levels were, in fact, found in this same group of patients (diagnostic

sensitivity of 68% and specicity of 86%).

46

Eects of Glucocorticoid and Aspirin Therapy on Leukocyte Counts Cardiovascular Disease Pharmacological dosesand of glucocorticoids elevate neutrophil
47

and depress lymphocyte and monocyte counts, with counts a net increase in leukocytes most common. Therefore, it is of interest that in a study of >65 000 patients and 80 000 controls, long-term therapy with doses of glucocorticoids exceeding the physiological range was found to be associated with increased risk

48

of cardiovascular events, even after adjustment for known risk factors.

49

In 4 separate nonrandomized studies of leukocytosis and cardiovascular risk involving a total of >160 000 patients, aspirin P=0.008; 11 P<0.001; 9 and P<0.0001 10 ). Because these studies were not randomized, many confounding factors may have inuenced the apparent association between aspirin use and lower leukocyte counts. However, if aspirin use actually is associated with lower leukocyte counts, at least a portion of the benecial eects of aspirin on cardiovascular events may be related to the reduction in leukocyte counts. use was associated with signicantly lower WBC counts (P=0.04;

Association of Leukocyte Count With Vascular Complications of Diabetes Diabetes, which is known to be associated with increased risk of
9,25,50 Moreover, even within the diabetic population, counts. the extent of leukocytosis is associated with macrovascular and microvascular complications of type 2 diabetes mellitus, as well as

vascular disease, is also associated with elevated leukocyte

26,51,52 Thus, among 3776 subjects endothelial dysfunction. studied in Hong Kong, after adjustment for sex, age, smoking status, disease duration, BMI, mean arterial pressure, waist-to-hip ratio, HbA1c, HDL cholesterol, and low-density lipoprotein cholesterol, patients with higher leukocyte counts remained at higher risk of macrovascular disease (angina, myocardial infarction, stroke, or peripheral vascular disease) and microvascular disease (the combination of retinopathy and albuminuria), with the odds ratio of the patients in the highestto-lowest quintiles of leukocyte count >2 for macrovascular and microvascular disease and with the P value for trend <0.001 for

In another study in type 2 diabetics, leukocyte count was both. associated with endothelial-dependent and endothelialindependent vasodilation, even after adjusting for other known risk factors, suggesting a potential association between leukocytosis and endothelial dysfunction.
52

26

5355 but the of ischemic vascular disease in numerous studies, physiological mechanisms responsible for the protective eects

Relationship Between WBC Count and Fitness, BMI, and Activity Levels Exercise and tness have been demonstrated to decrease the risk

have not been clearly dened. Similarly, there is evidence that BMI is associated with increased risk of cardiovascular disease, but it is less clear whether an elevated BMI is an independent risk Therefore, it is of interest that in a study of 4057 factor. nonsmoking men without a previous history of stroke or heart attack, the mean WBC count showed a signicant inverse correlation with tness (low, moderate, and high tness groups; 6637/L, 6,258/L, and 5,653/L WBC count, respectively;
30

56 Moreover, in accord with previous studies, 57 even P<0.001). after adjustment for age, within each tness group, the WBC count correlated directly with BMI. Further support for an inverse relationship between WBC count and physical activity, even after adjusting for smoking, sex, cardiovascular disease states, age, race, diabetes, BMI, and hypertension (P<0.001), comes from the study by Geken et al, which was conducted in 5868 men and 58 Moreover, in the Atherosclerosis Risk women 65 years of age. in Communities (ARIC) study of 14 679 participants aged 45 to 64 years, leisure activity, sports activity, and work activity indices correlated inversely with WBC count in whites, and the leisure activity and work activity indices correlated inversely with WBC count in blacks, even after excluding smokers and making several 50 Thus, if leukocytosis contributes directly to other adjustments. ischemic vascular disease, it is possible that at least some of the protective eects of exercise may operate through a reduction in WBC count.

Possible Underestimation of the Association of Leukocytosis With Cardiovascular Disease Variabilities in leukocyte determinations and inappropriate multivariate adjustments in the studies reviewed above may have Attributable to Variations in Leukocyte Counts and resulted in their underestimating the true association of Multivariate Adjustments for Smoking and BMI

leukocytosis with increased risk of cardiovascular disease. Thus, because many of the studies used only a single leukocyte determination, short- or medium-term factors such as infection or disease, seasonal or diurnal uctuations, or variability in the

45 may have aected the accuracy of leukocyte determinations results, resulting in mistakenly categorizing some individuals with lower levels as having higher levels and vice versa. When sucient data have been available to calculate the impact of such errors 59,60 ), they have been reported to (termed regression dilution result in underestimations of the true associations with cardiovascular disease of as much as 30% to 35% for leukocyte

and neutrophil counts and 50% for monocyte counts.

24

The strong association between cigarette smoking and elevated leukocyte counts has been documented in many

studies, 6,9,11,13,25,50,57,61 and even regular exposure to secondhand smoke is associated with increased leukocyte As a result, adjusting the association of leukocytosis counts. with ischemic vascular disease for smoking tends to lessen the association between leukocytosis and ischemic vascular events; however, even after this adjustment, the association remains signicant in most studies (Table 1). The implicit rationale for using this adjustment is the unproved assumption that smoking causes its adverse eects on ischemic vascular disease by mechanisms other than elevating the leukocyte count. If, on the other hand, smoking in fact causes its adverse eects through elevating the leukocyte count, then the eect of smoking should be adjusted for the eects of the elevated leukocyte count and not vice versa. When such an adjustment has been made, much of the eect of smoking is, in fact, eliminated (eg, in the Caerphilly collaborative studies, the unadjusted relative odds for cardiovascular events for current cigarette smokers was 2.27 [95% CI, 1.63 to 3.15], whereas after adjusting for WBC count, it was
62

27 Moreover, the rapid reduction in 1.63 [1.15 to 2.31]). cardiovascular risk (stroke and myocardial infarction) that occurs

63 may t well with the hypothesis that at after smoking cessation least some of adverse eects associated with smoking are mediated by leukocytosis.

Experimental Model and Human Blood Leukodepletion Data on the Role of Leukocytes in Given the robust epidemiologic data, it is surprising that there have been relatively few studies to assess the eect of leukocyte Ischemic Vascular Events

depletion on experimental ischemic vascular events. Although there is extensive literature on the importance of leukocytes in ischemia/reperfusion injury in dierent organs, including protection from injury by leukodepletion, adhesion receptor antagonists to M2 and P-selectin that demonstrated ecacy in these models have been disappointing when studied in humans with atherothrombotic ischemic vascular disease. There is intriguing evidence that leukocyte depletion of blood products in humans may be associated with better outcomes in patients undergoing cardiopulmonary bypass surgery, but the studies are not denitive.
71,72 6470

Leukocytes in Sickle Cell Disease

Sickle cell disease is characterized by a severe hemolytic anemia, but much of the morbidity and mortality of the disorder is

attributable to vascular occlusion. Pain crises are thought to be attributable to microvascular obstruction, primarily in
73 Macrovascular occlusion, primarily postcapillary venules. aecting the cerebral circulation in children, also occurs and is attributable to a combination of nonatherosclerotic intimal

74,75 Studies using modern hyperplasia and acute thrombosis. imaging techniques indicate that nearly half of patients with sickle cell disease have MRI evidence of brain injury before age 10, and nearly two thirds have cerebrovascular disease; of those with brain injury, nearly one third have macrovascular cerebrovascular

disease.

76

7780 In addition, count is associated with a worse prognosis. risk of stroke and silent cerebral infarction is greater in at least

The WBC count is elevated in nearly all patients with sickle cell disease, and epidemiologic data indicate that having a higher WBC

some groups of patients with higher leukocyte counts. Acute chest syndrome, one of the most serious complications of sickle cell disease, is accompanied by very high leukocyte and there have been 4 separate reports of patients counts, with sickle hemoglobin sustaining the rapid onset of pain crisis, acute chest syndrome, and even death after receiving a myeloid
83

81,82

8487 Increased expression of CSF (G-CSF or GM-CSF). leukocyte adhesion molecules has also been associated with more

severe disease.

88

89 Mice lacking the endothelial adhesion vessel wall itself. molecules that mediate leukocyte rolling and attachment,

Studies from our laboratory, conducted in collaboration with Dr Paul Frenette and colleagues at Mount Sinai School of Medicine, using intravital microscopy to assess vaso-occlusion in postcapillary venules of mice expressing human sickle hemoglobin, demonstrated that leukocytes play a crucial role in the process, with erythrocytes primarily interacting with the WBCs that are adherent to the wall of the venule rather than with the

P-selectin and E-selectin, were protected from vaso-occlusion. Thus, in this experimental model, leukocytes participate directly in microvascular obstruction. Hydroxyurea was introduced for the therapy of sickle cell disease on the basis of its ability to increase hemoglobin F and ameliorate

89

90 However, hydroxyurea also the frequency of painful crises. decreases the leukocyte count, and there is evidence that some patients respond to hydroxyurea more rapidly than can be accounted for by changes in hemoglobin F but consistent with the

time course of hydroxyurea-induced leukocyte reduction.

80,91

90 It remains possible, correlated with long-term mortality. however, that hydroxyurea-induced reductions in leukocyte counts contributed to the mortality benet in more subtle ways.

However, data from the Multicenter Study of Hydroxyurea in Sickle Cell Anemia did not support a major role for leukocyte reduction as a mechanism of the eects of hydroxyurea because there was no evidence that either pretreatment or post-treatment neutrophil counts above or below either 5000/L or 7000/L

Is it Justied to Try to Intervene to Reduce Leukocyte Counts in Patients With Ischemic Vascular Because evidence of an association between elevated leukocyte counts and ischemic vascular events is compelling, and because it Disease?
is impossible to exclude from current evidence the possibility that leukocytes directly contribute to the risk, it is logical to consider studies to assess the impact of altering leukocyte function or reducing the leukocyte count. Studies targeting leukocyte adhesion via eects on the integrin M2 and P-selectin have not demonstrated positive results in stroke or myocardial

64,70 although it is possible that new agents in this infarction, class will be more eective. However, it is possible that inhibition of any single leukocyte receptor is inadequate to achieve a therapeutic eect, in which case a strategy of lowering the leukocyte count may be more eective. Among the agents available to achieve a reduction in leukocyte count, hydroxyurea has the benets of rapid onset of action, relatively rapid recovery from its eects when the drug is stopped, relatively little organ toxicity, and an extensive history of its use for long periods of time in patients with myeloproliferative disorders and sickle cell 90,92 In general, in sickle cell disease, the toxicity prole disease. has been favorable, with few episodes of symptomatic severe 90,92 The mild reduction in neutropenia or serious anemia. platelet count commonly produced by hydroxyurea may, in fact,

be an added benet in reducing the risk of thrombotic events. However, there is concern that hydroxyurea is leukemogenic. Data from patients with myeloproliferative disorders suggest that hydroxyurea may increase the risk of leukemia, usually after treatment for 8 years, but there is considerable controversy about the extent to which hydroxyurea increases the risk. There is also controversy as to whether these patients underlying myeloproliferative disorders place them at higher risk of developing leukemia from hydroxyurea than individuals without such disorders. Although there were no cases of leukemia in 2 studies of hydroxyurea in patients with sickle cell disease, 1 in 122 children treated for an average of 2.8 years (range 0.5 to 8.4
9397

90

90 and there was no increase in acquired DNA mutations as years, judged by in vitro assays using DNA extracted from peripheral 92 there have been 3 blood mononuclear cells in the latter study, reports of patients with sickle cell disease treated with hydroxyurea developing leukemia: 1 after 2 years, the second

years) 92 and the other in 219 adults treated for an average of 7.6

after 6 years, and the third after 8 years of treatment.

98100

Together, the data summarized in this review make it biologically plausible that reducing the leukocyte count may improve the acute and chronic outcome of ischemic vascular disease. However, because the most attractive drug to accomplish a reduction in leukocyte counts is potentially leukemogenic, it is not certain that treatment would produce more benet than harm. To maximize the benet-to-risk ratio, a number of design features could be considered. Thus, restricting a study to patients who are at high risk for short-term mortality after myocardial infarction or acute coronary syndrome, namely, those who are of older age, have contraindications to reperfusion therapy, and have high leukocyte counts, would increase the potential for any single patient obtaining a benet. Similarly, limiting the therapy to 30 days and again restricting the study to elderly patients would decrease the likelihood that any single patient would experience adverse eects.

Acknowledgments
This work was supported in part by grants 54469 and 19278 from the National Heart, Lung, and Blood Institute. I wish to thank Suzanne Rivera for outstanding secretarial assistance, and Drs Robert Cali, Christopher Cannon, Stephen Fruchtman, John Harlan, and Eric Topol for constructive suggestions.

Received December 1, 2004. Accepted January 10, 2005.

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1. Arteriosclerosis, Thrombosis, and Vascular Biology. 2005; 25: 658-670 Published online before print January 20, 2005, doi: 10.1161/ 01.ATV.0000156877.94472.a5

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