- Email: [email protected]
Frequency of aspirin resistance in patients with congestive heart failure treated with antecedent aspirin
Frequency of Aspirin Resistance in Patients With Congestive Heart Failure Treated With Antecedent Aspirin David C. Sane,
MD,
Scott A. McKee, MD, Alex I. Malinin, Victor L. Serebruany, MD, PhD
hromboembolism is a common complication of congestive heart failure (CHF) with an estimated T frequency of ⬎2%/year. Beta thrombogloublin, 1
platelet factor 4, P-selectin, platelet/endothelial cell adhesion molecule, and other platelet markers are elevated in patients with CHF,2– 6 suggesting that platelet activation contributes to thromboembolic outcomes. Although aspirin is frequently administered to patients with CHF, particularly when the etiology is ischemic, the antiplatelet effect of aspirin in this population has not been carefully examined. We studied platelet function in 88 patients with documented CHF using a variety of techniques including conventional aggregometry (plasma and whole blood), shear stressinduced activation using the platelet function analyzer (PFA-100, Dade Behring, Inc., Miami, Florida), and the expression of major surface receptors using flow cytometry. •••
The study was approved by the Western Investigational Review Board (Olympia, Washington, protocol number 1229 WIRB) and performed in outpatient cardiology clinics. Written informed consent was obtained from all patients. The study population consisted of 88 outpatients with documented evidence of CHF (left ventricular ejection fraction ⬍40% and New York Heart Association class II to IV symptoms) who had been treated with aspirin 325 mg/day for ⱖ1 month. No patient received thienopyridines, glycoprotein IIb/IIIa inhibitors, or anticoagulant drugs. Blood samples were obtained with a 19-gauge needle by venipuncture and drawn into two 7-ml vacutainer tubes at room temperature containing 3.8% trisodium citrate. The vacutainer tube was filled to capacity and gently inverted 3 to 5 times to ensure complete mixing of the anticoagulant. The bloodcitrate mixture was centrifuged at 1,200 g for 2.5 minutes. The resulting platelet-rich plasma was kept at room temperature for use within 1 hour. The platelet count was determined in the plasma sample and adjusted to 3.5 ⫻ 108/ml with homologous platelet-poor plasma. Platelets were stimulated with 5 M of adenosine diphosphate (ADP), 1 g/ml of collagen, or 5 M of epinephrine, and aggregation was assessed From the Wake Forest University School of Medicine, Winston-Salem, North Carolina; and the Sinai Center for Thrombosis Research, Johns Hopkins University, Baltimore, Maryland. Dr. Serebruany’s address is: Center for Thrombosis Research, Sinai Hospital of Baltimore, 2401 West Belvedere Avenue, Schapiro Research Building-R 202, Baltimore, Maryland 21215. E-mail: [email protected]. Manuscript received February 27, 2002; revised manuscript received and accepted June 7, 2002. ©2002 by Excerpta Medica, Inc. All rights reserved. The American Journal of Cardiology Vol. 90 October 15, 2002
MD,
and
using a Chronolog Lumi-Aggregometer (model 560Ca, Chronolog, Inc., Hawerton, Pennsylvania) with the AggroLink software package (Chronolog Corporation, Hawerton, Pennsylvania). Aggregation was expressed as the maximal percent change in light transmittance from baseline, using platelet-poor plasma as a reference. Whole blood aggregation was determined using the Chronolog device. The whole blood-citrate mixture was diluted 1:1 with a 0.5 ml phosphate-buffered saline solution and gently swirled. The sample was warmed to 37°C for 5 minutes and then transferred to the assay well. Next, the electrode was placed in the cuvette and the sample stimulated with 4 g/ml of collagen. The change in electrical impedence was recorded as previously described.7 Platelet receptor expression was assessed by flow cytometry using monoclonal antibodies to the following antigens: CD41 (glycoprotein [GP] IIb/IIIa, ␣ II 3) CD42b (GP Ib), CD62p (P-selectin) (DAKO Corporation, Carpenteria, California), CD51/CD61 (␣v3, or vitronectin receptor), CD31 (platelet/endothelial cell adhesion molecule⫺1), CD107a (lyzosome-associated membrane protein [LAMP]⫺1), CD107b (LAMP⫺2), CD63 (LAMP⫺3), CD151 (platelet-endothelian tetraspan antigen [PETA]-3), and PAC⫺1 (PharMingen, San Diego, California). Platelet-leukocyte aggregate formation was measured by dual labeling with the pan-platelet marker (CD151) and CD14, the macrophage receptor for endotoxin lipopolysaccharides. The blood-citrate mixture (50 l) was diluted with 450 l of tris-buffered saline (10 mmol/L tris, 0.15 mol/L sodium chloride) and mixed by gently inverting an Eppendorf tube 2 times. The corresponding antibody was then added (5 l) and incubated at 4°C for 30 minutes. After incubation, 400 l of 2% buffered paraformaldehyde was added for fixation. The samples were analyzed on a Becton Dickinson FACScan flow cytometer (San Diego, California) set up to measure fluorescent light scatter as previously described.8 All parameters were collected using 4-decade logarithmic amplification. Data were collected in list mode files and then analyzed. P-selectin was expressed as percent positive cells as previously described.9 Other antigens were expressed as log mean fluorescence intensity. Differences between individual flow cytometric histograms were assessed using the Smirnov-Kolmogorov test incorporated in CELLQuest software (San Diego, California). The effect of shear stress on platelet function was analyzed using the platelet function analyzer, PFA100. Closure times were determined using the colla0002-9149/02/$–see front matter PII S0002-9149(02)02718-2
893
TABLE 1 Demographics and Clinical Characteristics of Patients With Heart Failure
Demographics Age (yrs) Men White Black Smoking history Stable angina Unstable angina Previous myocardial infarction Peripheral vascular disease Diabetis mellitus Systemic hypertension Coronary bypass Hypercholesterolemia† Heart failure etiology Ischemic Nonischemic New York Heart Association class II III IV Ejection fraction (%) Medications  blockers ACE inhibitors Calcium-channel blockers Diuretics Glycosides Nitrates Antilipid agents
Aspirin Nonresponders (n ⫽ 50)
Aspirin Responders (n ⫽ 38)
66 ⫾ 12 26 (52%) 11 (22%) 39 (78%) 18 (36%) 24 (48%) 3 (6%) 19 (38%) 2 (4%) 14 (28%) 36 (72%) 10 (20%) 30 (60%)
64 ⫾ 14 18 (50%) 8 (22%) 28 (78%) 12 (33%) 11 (30%)* 3 (8%) 3 (8%)* 2 (5.5%) 5 (14%)* 10 (28%)* 3 (8%)* 13 (36%)*
33 (66%) 17 (34%)
26 (72%) 10 (28%)
16 (32%) 18 (64%) 2 (4%) 33.1 ⫾ 8.8
24 (67%)* 12 (33%)* 0 37.1 ⫾ 7.4
38 40 13 33 8 13 26
(76%) (80%) (26%) (66%) (16%) (26%) (52%)
30 24 14 22 6 11 15
(82%) (71%) (39%)* (60%) (17%) (31%) (41%)
*p ⬍0.05 between groups. † Hypercholesterolemia based on total cholesterol ⬎220 mg/dl and low-density lipoprotein cholesterol ⬎130 mg/dl. ACE ⫽ angiotensin-converting enzyme.
TABLE 2 Platelet Markers Aspirin Nonresponders (n ⫽ 50) ADP aggregation (%) Collagen aggregation (%) Epinephrine aggregation (%) Whole blood aggregation (ohms) Closure time (s) GP IIb/IIIa antigen (MFI) GP IIb/IIIa activity GP Ib (MFI) P-selectin (%⫹) Vitronectin receptor (MFI) CD31 (MFI) CD107a (MFI) CD107b (MFI) CD63 (MFI) CD151 (MFI) CD151 ⫹ CD 14 (MFI)
68 81 28 23 204 553 13 281 11 10 85 10 9 10 136 144
⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾
9 6 8 4 44 52 4 38 4 4 17 3 3 4 27 43
Aspirin Responders (n ⫽ 38) 54 78 28 24 199 488 10 240 8 10 78 9 8 12 111 115
⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾
Data are expressed as mean ⫾ SD. MFI ⫽ mean fluorescence intensity.
gen/epinephrine test cartridges. The closure time is prolonged after therapy with aspirin, and this cartridge has been extensively used to monitor aspirin pharmacodynamics.10 Normal ranges and factors that influ894 THE AMERICAN JOURNAL OF CARDIOLOGY姞
VOL. 90
13 6 7 6 42 85 3 66 3 7 20 3 3 4 47 44
p Value 0.007 NS NS NS NS 0.03 0.04 0.04 0.03 NS NS NS NS NS 0.01 0.02
ence closure time have been well characterized.11 Data are expressed as mean ⫾ SD, and a p value ⬍0.05 was considered significant. We identified persistent platelet activation (aspirin nonresponsiveness) when 4 of the following 5 parameters occurred: collagen-induced aggregation ⬎70%; adenosine diphosphate-induced aggregation ⬎60%, whole blood aggregation ⬎18 ohms; expression of active GP IIb/IIIa ⬎220 log mean fluorescence intensity units; and P-selectin positivity ⬎8%. The demographics and medical characteristics of aspirin responders and nonresponders are listed in Table 1. Groups did not differ in age, race, or gender. However, aspirin nonresponders were more likely to have stable angina, have had a previous myocardial infarction, and have diabetes, hypertension, and hyperlipidemia. These patients were also more likely to have New York Heart Association class III to IV rather than class II CHF symptoms, but less likely to be treated with calcium channel blockers. Overall, clinical characteristics suggest that aspirin nonresponders have more extensive coronary artery disease, greater incidence of risk factors, and more heart failure symptoms than do patients who respond to aspirin therapy. The platelet function studies in the 2 groups are listed in Table 2. Using the criterion of 4 of 5 markers, persistent platelet activation despite aspirin therapy was detected in 50 of 88 patients (56.8%). Patients identified as nonresponsive to aspirin had significantly lower ADP-induced aggregation (p ⫽ 0.007), expression of GP IIb/IIIa (p ⫽ 0.03), activity of GP IIb/IIIa (PAC⫺1; p ⫽ 0.04), expression of GP Ib (p ⫽ 0.04), Pselectin positivity (p ⫽ 0.003), platelet-endothelial cell tetra-span antigen (CD151; p ⫽ 0.01), and platelet-leukocyte aggregates (CD151 ⫹ CD14; p ⫽ 0.04). •••
We have found that patients with CHF are frequently resistant to the antiplatelet activity of aspirin. The prevalence of aspirin resistance is more than double that reported in patients with stable general cardiovascular disease, in whom aspirin resistance or semiresponsiveness was noted in 5.5% to 23.8%.12 In that study, aspirin resistance was defined by aggregation studies as a mean OCTOBER 15, 2002
aggregation ⱖ70% in response to 10 M of ADP, or ⱖ20% with 0.5 mg/ml of arachidonic acid. Semiresponders were defined as patients meeting 1, but not both, of these criteria. Aspirin resistance was also defined as occurring when the closure time was normal (ⱕ193 seconds) using the collagen-epinephrine cartridge in the PFA-100 analyzer.12 There is no standard definition of aspirin resistance, with a variety of criteria having been used in previous publications. If we reanalyze our population using the criterion of closure time with the collagen/epinephrine cartridge ⱕ193 seconds, we find that 49 of 88 patients (55.7%) would be labeled as aspirin resistant. Thus, despite using differing standards of assigning aspirin resistance, it is clear that this condition is common. We postulate that elevated catecholamines,13 angiotensin II,14 or cytosolic calcium levels15 in patients with heart failure may contribute to a high rate of aspirin resistance in this emerging population. The presence of diabetes, hypertension, and hyperlipidemia in patients with CHF may further increase the propensity for platelet activation and aspirin resistance, as was documented in our nonresponders group. These risk factors produce endothelial dysfunction, which is associated with nitric oxide deficiency and platelet activation.16 Patients with acute coronary ischemic syndromes and aspirin resistance have a higher rate of adverse outcomes.17 Similarly, aspirin resistant patients with heart failure may be especially susceptible to thromboembolic events. Studies are warranted to determine whether patients have different risks for thromboembolic events as a function of their sensitivity to aspirin. The major platelet function difference between our 2 groups was in the extent of aggregation to 5 M ADP (68.4 ⫾ 9.4% vs 54.2 ⫾ 13.4%; p ⫽ 0.007). Because clopidogrel is a specific antagonist of the platelet ADP (P2Y12) receptor, is well-tolerated, and has proven benefit in patients with cardiovascular disease,18,19 this drug would appear to be the ideal choice for additional therapy in the population with CHF. The ongoing Warfarin Antiplatelet Therapy in Chronic Heart failure (WATCH) trial comparing aspirin, clopidogrel, and warfarin in patients with CHF will help elucidate the optimum thromboembolism prophylaxis in this population.
Until definitive clinical trials are completed, physicians should be aware of the high rate of aspirin resistance in patients with heart failure and consider clopidogrel for treating high-risk patients or patients in whom thromboembolism occurs despite receiving aspirin. 1. Garg RK, Gheorghiade M, Jafri SM. Antiplatelet and anticoagulant therapy in the prevention of thromboemboli in chronic heart failure. Prog Cardiovasc Dis 1998;41:225–236. 2. Weidinger F, Glogar D, Sochor H, Sinzinger H. Platelet survival in patients with dilated cardiomyopathy. Thromb Haemost 1991;66:400 –405. 3. Jafri SM, Riddle JM, Raman SB, Goldstein S. Altered platelet function in patients with severe congestive heart failure. Henry Ford Hosp Med J 1986;34: 156 –159. 4. Andreassen AK, Nordoy I, Simonsen S, Ueland T, Muller F, Froland SS, Gullestad L, Aukrust P. Levels of circulating adhesion molecules in congestive heart failure and after heart transplantation. Am J Cardiol 1998;81:604 –608. 5. O’Connor CM, Gurbel PA, Serebruany VL. Usefulness of soluble and surfacebound P-selectin in detecting heightened platelet activity in patients with congestive heart failure. Am J Cardiol 1999;83:1345–1349. 6. Serebruany VL, Murugesan SR, Pothula A, Atar D, Lowry D, Gattis WA, O’Connor CM, Gurbel PA. Increased soluble platelet/endothelial cellular adhesion molecule-1, and osteonectin levels in patients with severe congestive heart failure. Independence of disease etiology, and antecedent aspirin therapy. Eur J Heart Fail 1999;1:243–249. 7. Abbate R, Favilla S, Boddi M, Costanzo G, Prisco D. Factors influencing platelet a aggregation in the whole blood. Am J Clin Pathol 1986;86:91–96. 8. Ault KA. Flow cytometric measurement of platelet function and reticulated platelets. Ann New York Acad Sci 1993;677:293–308. 9. Serebruany VL, Kereiakes DJ, Dalesandro MR, Gurbel PA. Model of flow cytometer markedly affects platelet-bound P-selectin expression in patients with chest pain. Are we comparing apples with oranges? Thromb Res 1999;96:51–56. 10. Homoncik M, Jilma B, Hergovich N, Stohlawetz P, Panzer S, Speiser W. Monitoring of aspirin (ASA) pharmacodynamics with the platelet function analyzer PFA-100威. Thromb Haemost 2000;83:316 –321. 11. Jilma B. Platelet function analyzer (PFA-100): a tool to quantify congenital or acquired platelet dysfunction. J Lab Clin Med 2001;138:152–163. 12. Gum PA, Kottke-Marchant K, Poggio ED, Gurm H, Welsh PA, Brooks L, Sapp SK, Topol EJ. Profile and prevalence of aspirin resistance in patients with cardiovascular disease. Am J Cardiol 2001;88:230 –235. 13. Anfossi G, Trovati M. Role of catecholamines in platelet function: pathophysiological and clinical significance. Eur J Clin Invest 1996;26:353–370. 14. Ding YA, MacIntyre DE, Kenyon CJ, Semple PF. Angiotensin II effects on platelet function. J Hypertens 1985;(suppl 3):S251–S253. 15. Negrescu EV, Sazonova LN, Baldenkov GN, Mucharlyamov NM, Mazaev AV, Tkachuk VA. Relationship between the inhibition of receptor-induced increase in cytosolic free calcium concentration and the vasodilator effects of nitrates in patients with congestive heart failure. Int J Cardiol 1990;26:175–181. 16. Loscalzo J. Nitric oxide insufficiency, platelet activation, and arterial thrombosis. Circ Res 2001;88:756 –762. 17. Alexander JH, Harrington RA, Tuttle RH, Berdan LG, Lincoff AM, Deckers JW, Simoons ML, Guerci A, Hochman JS, Wilcox RG, et al. Prior aspirin use predicts worse outcomes in patients with non–ST-elevation acute coronary syndromes. Am J Cardiol 1999;83:1147–1151. 18. CURE Investigators. Effects of clopidogrel in addition to aspirin in patients with acute coronary syndromes without ST-segment elevation. N Engl J Med 2001;345:494 –502. 19. Bhatt DL, Chew DP, Hirsch AT, Ringleb PA, Hacke W, Topol EJ. Superiority of clopidogrel versus aspirin in patients with prior cardiac surgery. Circulation 2001;103:363–368.
BRIEF REPORTS
895
MD,
Scott A. McKee, MD, Alex I. Malinin, Victor L. Serebruany, MD, PhD
hromboembolism is a common complication of congestive heart failure (CHF) with an estimated T frequency of ⬎2%/year. Beta thrombogloublin, 1
platelet factor 4, P-selectin, platelet/endothelial cell adhesion molecule, and other platelet markers are elevated in patients with CHF,2– 6 suggesting that platelet activation contributes to thromboembolic outcomes. Although aspirin is frequently administered to patients with CHF, particularly when the etiology is ischemic, the antiplatelet effect of aspirin in this population has not been carefully examined. We studied platelet function in 88 patients with documented CHF using a variety of techniques including conventional aggregometry (plasma and whole blood), shear stressinduced activation using the platelet function analyzer (PFA-100, Dade Behring, Inc., Miami, Florida), and the expression of major surface receptors using flow cytometry. •••
The study was approved by the Western Investigational Review Board (Olympia, Washington, protocol number 1229 WIRB) and performed in outpatient cardiology clinics. Written informed consent was obtained from all patients. The study population consisted of 88 outpatients with documented evidence of CHF (left ventricular ejection fraction ⬍40% and New York Heart Association class II to IV symptoms) who had been treated with aspirin 325 mg/day for ⱖ1 month. No patient received thienopyridines, glycoprotein IIb/IIIa inhibitors, or anticoagulant drugs. Blood samples were obtained with a 19-gauge needle by venipuncture and drawn into two 7-ml vacutainer tubes at room temperature containing 3.8% trisodium citrate. The vacutainer tube was filled to capacity and gently inverted 3 to 5 times to ensure complete mixing of the anticoagulant. The bloodcitrate mixture was centrifuged at 1,200 g for 2.5 minutes. The resulting platelet-rich plasma was kept at room temperature for use within 1 hour. The platelet count was determined in the plasma sample and adjusted to 3.5 ⫻ 108/ml with homologous platelet-poor plasma. Platelets were stimulated with 5 M of adenosine diphosphate (ADP), 1 g/ml of collagen, or 5 M of epinephrine, and aggregation was assessed From the Wake Forest University School of Medicine, Winston-Salem, North Carolina; and the Sinai Center for Thrombosis Research, Johns Hopkins University, Baltimore, Maryland. Dr. Serebruany’s address is: Center for Thrombosis Research, Sinai Hospital of Baltimore, 2401 West Belvedere Avenue, Schapiro Research Building-R 202, Baltimore, Maryland 21215. E-mail: [email protected]. Manuscript received February 27, 2002; revised manuscript received and accepted June 7, 2002. ©2002 by Excerpta Medica, Inc. All rights reserved. The American Journal of Cardiology Vol. 90 October 15, 2002
MD,
and
using a Chronolog Lumi-Aggregometer (model 560Ca, Chronolog, Inc., Hawerton, Pennsylvania) with the AggroLink software package (Chronolog Corporation, Hawerton, Pennsylvania). Aggregation was expressed as the maximal percent change in light transmittance from baseline, using platelet-poor plasma as a reference. Whole blood aggregation was determined using the Chronolog device. The whole blood-citrate mixture was diluted 1:1 with a 0.5 ml phosphate-buffered saline solution and gently swirled. The sample was warmed to 37°C for 5 minutes and then transferred to the assay well. Next, the electrode was placed in the cuvette and the sample stimulated with 4 g/ml of collagen. The change in electrical impedence was recorded as previously described.7 Platelet receptor expression was assessed by flow cytometry using monoclonal antibodies to the following antigens: CD41 (glycoprotein [GP] IIb/IIIa, ␣ II 3) CD42b (GP Ib), CD62p (P-selectin) (DAKO Corporation, Carpenteria, California), CD51/CD61 (␣v3, or vitronectin receptor), CD31 (platelet/endothelial cell adhesion molecule⫺1), CD107a (lyzosome-associated membrane protein [LAMP]⫺1), CD107b (LAMP⫺2), CD63 (LAMP⫺3), CD151 (platelet-endothelian tetraspan antigen [PETA]-3), and PAC⫺1 (PharMingen, San Diego, California). Platelet-leukocyte aggregate formation was measured by dual labeling with the pan-platelet marker (CD151) and CD14, the macrophage receptor for endotoxin lipopolysaccharides. The blood-citrate mixture (50 l) was diluted with 450 l of tris-buffered saline (10 mmol/L tris, 0.15 mol/L sodium chloride) and mixed by gently inverting an Eppendorf tube 2 times. The corresponding antibody was then added (5 l) and incubated at 4°C for 30 minutes. After incubation, 400 l of 2% buffered paraformaldehyde was added for fixation. The samples were analyzed on a Becton Dickinson FACScan flow cytometer (San Diego, California) set up to measure fluorescent light scatter as previously described.8 All parameters were collected using 4-decade logarithmic amplification. Data were collected in list mode files and then analyzed. P-selectin was expressed as percent positive cells as previously described.9 Other antigens were expressed as log mean fluorescence intensity. Differences between individual flow cytometric histograms were assessed using the Smirnov-Kolmogorov test incorporated in CELLQuest software (San Diego, California). The effect of shear stress on platelet function was analyzed using the platelet function analyzer, PFA100. Closure times were determined using the colla0002-9149/02/$–see front matter PII S0002-9149(02)02718-2
893
TABLE 1 Demographics and Clinical Characteristics of Patients With Heart Failure
Demographics Age (yrs) Men White Black Smoking history Stable angina Unstable angina Previous myocardial infarction Peripheral vascular disease Diabetis mellitus Systemic hypertension Coronary bypass Hypercholesterolemia† Heart failure etiology Ischemic Nonischemic New York Heart Association class II III IV Ejection fraction (%) Medications  blockers ACE inhibitors Calcium-channel blockers Diuretics Glycosides Nitrates Antilipid agents
Aspirin Nonresponders (n ⫽ 50)
Aspirin Responders (n ⫽ 38)
66 ⫾ 12 26 (52%) 11 (22%) 39 (78%) 18 (36%) 24 (48%) 3 (6%) 19 (38%) 2 (4%) 14 (28%) 36 (72%) 10 (20%) 30 (60%)
64 ⫾ 14 18 (50%) 8 (22%) 28 (78%) 12 (33%) 11 (30%)* 3 (8%) 3 (8%)* 2 (5.5%) 5 (14%)* 10 (28%)* 3 (8%)* 13 (36%)*
33 (66%) 17 (34%)
26 (72%) 10 (28%)
16 (32%) 18 (64%) 2 (4%) 33.1 ⫾ 8.8
24 (67%)* 12 (33%)* 0 37.1 ⫾ 7.4
38 40 13 33 8 13 26
(76%) (80%) (26%) (66%) (16%) (26%) (52%)
30 24 14 22 6 11 15
(82%) (71%) (39%)* (60%) (17%) (31%) (41%)
*p ⬍0.05 between groups. † Hypercholesterolemia based on total cholesterol ⬎220 mg/dl and low-density lipoprotein cholesterol ⬎130 mg/dl. ACE ⫽ angiotensin-converting enzyme.
TABLE 2 Platelet Markers Aspirin Nonresponders (n ⫽ 50) ADP aggregation (%) Collagen aggregation (%) Epinephrine aggregation (%) Whole blood aggregation (ohms) Closure time (s) GP IIb/IIIa antigen (MFI) GP IIb/IIIa activity GP Ib (MFI) P-selectin (%⫹) Vitronectin receptor (MFI) CD31 (MFI) CD107a (MFI) CD107b (MFI) CD63 (MFI) CD151 (MFI) CD151 ⫹ CD 14 (MFI)
68 81 28 23 204 553 13 281 11 10 85 10 9 10 136 144
⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾
9 6 8 4 44 52 4 38 4 4 17 3 3 4 27 43
Aspirin Responders (n ⫽ 38) 54 78 28 24 199 488 10 240 8 10 78 9 8 12 111 115
⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾ ⫾
Data are expressed as mean ⫾ SD. MFI ⫽ mean fluorescence intensity.
gen/epinephrine test cartridges. The closure time is prolonged after therapy with aspirin, and this cartridge has been extensively used to monitor aspirin pharmacodynamics.10 Normal ranges and factors that influ894 THE AMERICAN JOURNAL OF CARDIOLOGY姞
VOL. 90
13 6 7 6 42 85 3 66 3 7 20 3 3 4 47 44
p Value 0.007 NS NS NS NS 0.03 0.04 0.04 0.03 NS NS NS NS NS 0.01 0.02
ence closure time have been well characterized.11 Data are expressed as mean ⫾ SD, and a p value ⬍0.05 was considered significant. We identified persistent platelet activation (aspirin nonresponsiveness) when 4 of the following 5 parameters occurred: collagen-induced aggregation ⬎70%; adenosine diphosphate-induced aggregation ⬎60%, whole blood aggregation ⬎18 ohms; expression of active GP IIb/IIIa ⬎220 log mean fluorescence intensity units; and P-selectin positivity ⬎8%. The demographics and medical characteristics of aspirin responders and nonresponders are listed in Table 1. Groups did not differ in age, race, or gender. However, aspirin nonresponders were more likely to have stable angina, have had a previous myocardial infarction, and have diabetes, hypertension, and hyperlipidemia. These patients were also more likely to have New York Heart Association class III to IV rather than class II CHF symptoms, but less likely to be treated with calcium channel blockers. Overall, clinical characteristics suggest that aspirin nonresponders have more extensive coronary artery disease, greater incidence of risk factors, and more heart failure symptoms than do patients who respond to aspirin therapy. The platelet function studies in the 2 groups are listed in Table 2. Using the criterion of 4 of 5 markers, persistent platelet activation despite aspirin therapy was detected in 50 of 88 patients (56.8%). Patients identified as nonresponsive to aspirin had significantly lower ADP-induced aggregation (p ⫽ 0.007), expression of GP IIb/IIIa (p ⫽ 0.03), activity of GP IIb/IIIa (PAC⫺1; p ⫽ 0.04), expression of GP Ib (p ⫽ 0.04), Pselectin positivity (p ⫽ 0.003), platelet-endothelial cell tetra-span antigen (CD151; p ⫽ 0.01), and platelet-leukocyte aggregates (CD151 ⫹ CD14; p ⫽ 0.04). •••
We have found that patients with CHF are frequently resistant to the antiplatelet activity of aspirin. The prevalence of aspirin resistance is more than double that reported in patients with stable general cardiovascular disease, in whom aspirin resistance or semiresponsiveness was noted in 5.5% to 23.8%.12 In that study, aspirin resistance was defined by aggregation studies as a mean OCTOBER 15, 2002
aggregation ⱖ70% in response to 10 M of ADP, or ⱖ20% with 0.5 mg/ml of arachidonic acid. Semiresponders were defined as patients meeting 1, but not both, of these criteria. Aspirin resistance was also defined as occurring when the closure time was normal (ⱕ193 seconds) using the collagen-epinephrine cartridge in the PFA-100 analyzer.12 There is no standard definition of aspirin resistance, with a variety of criteria having been used in previous publications. If we reanalyze our population using the criterion of closure time with the collagen/epinephrine cartridge ⱕ193 seconds, we find that 49 of 88 patients (55.7%) would be labeled as aspirin resistant. Thus, despite using differing standards of assigning aspirin resistance, it is clear that this condition is common. We postulate that elevated catecholamines,13 angiotensin II,14 or cytosolic calcium levels15 in patients with heart failure may contribute to a high rate of aspirin resistance in this emerging population. The presence of diabetes, hypertension, and hyperlipidemia in patients with CHF may further increase the propensity for platelet activation and aspirin resistance, as was documented in our nonresponders group. These risk factors produce endothelial dysfunction, which is associated with nitric oxide deficiency and platelet activation.16 Patients with acute coronary ischemic syndromes and aspirin resistance have a higher rate of adverse outcomes.17 Similarly, aspirin resistant patients with heart failure may be especially susceptible to thromboembolic events. Studies are warranted to determine whether patients have different risks for thromboembolic events as a function of their sensitivity to aspirin. The major platelet function difference between our 2 groups was in the extent of aggregation to 5 M ADP (68.4 ⫾ 9.4% vs 54.2 ⫾ 13.4%; p ⫽ 0.007). Because clopidogrel is a specific antagonist of the platelet ADP (P2Y12) receptor, is well-tolerated, and has proven benefit in patients with cardiovascular disease,18,19 this drug would appear to be the ideal choice for additional therapy in the population with CHF. The ongoing Warfarin Antiplatelet Therapy in Chronic Heart failure (WATCH) trial comparing aspirin, clopidogrel, and warfarin in patients with CHF will help elucidate the optimum thromboembolism prophylaxis in this population.
Until definitive clinical trials are completed, physicians should be aware of the high rate of aspirin resistance in patients with heart failure and consider clopidogrel for treating high-risk patients or patients in whom thromboembolism occurs despite receiving aspirin. 1. Garg RK, Gheorghiade M, Jafri SM. Antiplatelet and anticoagulant therapy in the prevention of thromboemboli in chronic heart failure. Prog Cardiovasc Dis 1998;41:225–236. 2. Weidinger F, Glogar D, Sochor H, Sinzinger H. Platelet survival in patients with dilated cardiomyopathy. Thromb Haemost 1991;66:400 –405. 3. Jafri SM, Riddle JM, Raman SB, Goldstein S. Altered platelet function in patients with severe congestive heart failure. Henry Ford Hosp Med J 1986;34: 156 –159. 4. Andreassen AK, Nordoy I, Simonsen S, Ueland T, Muller F, Froland SS, Gullestad L, Aukrust P. Levels of circulating adhesion molecules in congestive heart failure and after heart transplantation. Am J Cardiol 1998;81:604 –608. 5. O’Connor CM, Gurbel PA, Serebruany VL. Usefulness of soluble and surfacebound P-selectin in detecting heightened platelet activity in patients with congestive heart failure. Am J Cardiol 1999;83:1345–1349. 6. Serebruany VL, Murugesan SR, Pothula A, Atar D, Lowry D, Gattis WA, O’Connor CM, Gurbel PA. Increased soluble platelet/endothelial cellular adhesion molecule-1, and osteonectin levels in patients with severe congestive heart failure. Independence of disease etiology, and antecedent aspirin therapy. Eur J Heart Fail 1999;1:243–249. 7. Abbate R, Favilla S, Boddi M, Costanzo G, Prisco D. Factors influencing platelet a aggregation in the whole blood. Am J Clin Pathol 1986;86:91–96. 8. Ault KA. Flow cytometric measurement of platelet function and reticulated platelets. Ann New York Acad Sci 1993;677:293–308. 9. Serebruany VL, Kereiakes DJ, Dalesandro MR, Gurbel PA. Model of flow cytometer markedly affects platelet-bound P-selectin expression in patients with chest pain. Are we comparing apples with oranges? Thromb Res 1999;96:51–56. 10. Homoncik M, Jilma B, Hergovich N, Stohlawetz P, Panzer S, Speiser W. Monitoring of aspirin (ASA) pharmacodynamics with the platelet function analyzer PFA-100威. Thromb Haemost 2000;83:316 –321. 11. Jilma B. Platelet function analyzer (PFA-100): a tool to quantify congenital or acquired platelet dysfunction. J Lab Clin Med 2001;138:152–163. 12. Gum PA, Kottke-Marchant K, Poggio ED, Gurm H, Welsh PA, Brooks L, Sapp SK, Topol EJ. Profile and prevalence of aspirin resistance in patients with cardiovascular disease. Am J Cardiol 2001;88:230 –235. 13. Anfossi G, Trovati M. Role of catecholamines in platelet function: pathophysiological and clinical significance. Eur J Clin Invest 1996;26:353–370. 14. Ding YA, MacIntyre DE, Kenyon CJ, Semple PF. Angiotensin II effects on platelet function. J Hypertens 1985;(suppl 3):S251–S253. 15. Negrescu EV, Sazonova LN, Baldenkov GN, Mucharlyamov NM, Mazaev AV, Tkachuk VA. Relationship between the inhibition of receptor-induced increase in cytosolic free calcium concentration and the vasodilator effects of nitrates in patients with congestive heart failure. Int J Cardiol 1990;26:175–181. 16. Loscalzo J. Nitric oxide insufficiency, platelet activation, and arterial thrombosis. Circ Res 2001;88:756 –762. 17. Alexander JH, Harrington RA, Tuttle RH, Berdan LG, Lincoff AM, Deckers JW, Simoons ML, Guerci A, Hochman JS, Wilcox RG, et al. Prior aspirin use predicts worse outcomes in patients with non–ST-elevation acute coronary syndromes. Am J Cardiol 1999;83:1147–1151. 18. CURE Investigators. Effects of clopidogrel in addition to aspirin in patients with acute coronary syndromes without ST-segment elevation. N Engl J Med 2001;345:494 –502. 19. Bhatt DL, Chew DP, Hirsch AT, Ringleb PA, Hacke W, Topol EJ. Superiority of clopidogrel versus aspirin in patients with prior cardiac surgery. Circulation 2001;103:363–368.
BRIEF REPORTS
895