Relationship between hemostatic abnormalities and neuroendocrine activity in heart failure

Relationship abnormalities heart failure

between hemostatic and neuroendocrine

activity

in

Thromboembolism is an important complication of heart failure. To test the hypothesis that heart failure may be associated with hemostatic dysfunction, we studied hemostatic function in 21 patients with stable chronic heart failure and related these measures to the severity of heart failure as assessed by clinical evaluation, neuroendocrine activation, radionuclide ventriculography, and cardiopulmonary exercise testing. Plasma and blood viscosity were elevated; all patients showed evidence of platelet activation, and many had elevated plasma concentrations of fibrinopeptide A, D-dimer, and von Willebrand factor. The plasma concentrations of these variables were poorly interrelated and related poorly to the severity of heart failure. Plasma concentrations of angiotensin II and endothelin were correlated, and the latter was also correlated with the plasma concentration of von Willebrand factor. Patients with chronic heart failure have hemostatic abnormalities that may predispose them to thromboembolic events and may be in part due to neuroendocrine activation. (AM HEART J 1994;127:607-12.)

Eftihia Sbarouni, MD, Anne Bradshaw, BSc, Felicita Andreotti, MD, PhD, Edward Tuddenham, MD, FRCP, FRCPath, Celia M. Oakley, MD, FRCP, and John G. F. Cleland, MD, FRCP London, England

Patients with heart failure are at increased risk for thromboembolic events,’ and this has generally been attributed to stasis of blood caused by a dilated and poorly contracting heart. Blood stasis may be of primary importance, but endothelial dysfunction and neuroendocrine activation could also alter hemostatic function, increasing the tendency to thrombosis.2-7 Although angiotensin converting enzyme (ACE) inhibitors exacerbate exertional angina in patients with heart failure,8 a recent study indicated that administration of enalapril reduced the incidence of unstable angina and myocardial infarction in such patientq9 suggesting that enalapril may exert a beneficial effect on hemostatic function. We tested the hypothesis that patients with heart failure are in a hypercoagulable state, which may

predispose them to thromboembolic events, and assessed the possible relationship between hemostatic variables and indices of neuroendocrine activity. Three main pathways of hemostasis were monitored; plasma fibrinopeptide A, an index of fibrin formation by thrombinlO; &thromboglobulin, an index of platelet activationrl; and D-dimer an index of plasmin activity on cross-linked fibrin.12 We also measured fibrinogen, blood and plasma viscosity, factor VII, and von Willebrand factor because they may be risk factors for thrombotic events.13-15 Measures of hemostatic activity were related to plasma concentrations of catecholamines, angiotensin II, and endothelin. The relationships between hemostatic factors and ventricular function, exercise tolerance, and causes of heart failure were also analyzed. METHODS Patients.

From the Department Postgraduate Medical Dr. Andreotti Received

of Medicine (Cardiology) School, Hammersmith

was supported for publication

by a grant March

Copyright ‘2 1994 OOOZ-8703/94/$3.00+

by Mosby-Year 0 4/l/51798

Haematology, London.

from The British

15,1993;

Reprint requests: John G. F. Cleland, (Cardiology), Hammersmith Hospital, England.

and Hospital,

accepted

MD, FRCP, Du Cane Book,

Inc.

July

Heart

Royal

Foundation.

5,1993.

Department Rd. London

of Medicine W12 ONN,

Twenty-one patients with moderate to severe but stable heart failure were studied. Twenty were men, and the meanagewas 58 years (range, 34 to 78 years). The diagnosiswas dilated cardiomyopathy in 13 and ischemic heart diseasein eight. Three were in New York Heart Association (NYHA) classII, 12 in classIII, and six in class IV. All patients were in sinus rhythm, and none had hypertension or diabetes. Patients with a recent myocardial infarction, unstable angina, overt abnormalities of co-

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Table I. Viscosities, coagulation screen, and neuroendocrine profile in 21 patients with moderate to severeheart failure Variable (n = 21)

Mean

Plasmaviscosity (mPas) Bloodviscosity (mPas)NH,LS Bloodviscosity NH,HS BloodviscosityCH,LS Bloodviscosity CH,HS Prothrombintime (set) Activated PTT (set) Thrombintime (set) Fibrinogen(rig/ml) Factor VII activity W/ml) Noradrenaline(pg/ml) Adrenaline(pg/ml) AngiotensinII (pg/ml) Endothelin(pmol/L)

1.5 (0.2)

1.3-2.0

1.2-1.5

51 (14)

36-78

28-56

4.6 (0.7)

3.9-5.9

4-5

51 (12) 4.6 (0.5)

38-60 3.9-5.6

34-56 4-5

15 (1)

13-18

12-15.5

(SD)

Range

Normal range

38 14 2.8 0.7

(3) (1) (0.7) (0.2)

32-43 12-17 2.0-5.2 0.5-1.3

30-46 15-19 2-4 0.7-1.3

821 53 38 2.2

(362) (37) (18) (0.67)

304-1386 18-146 12-73 1.1-3.5

135-406 <73 1.2-10.4 0.44-0.64

NH, Native hematocrit; LS, low shear rate; HS, corrected hematocrit (45% ).

high shear rate; CH,

agulation, or systemic diseasewere excluded. All patients were treated with a diuretic and an ACE enzyme inhibitor for at least 3 months. Twenty-one patients were also receiving other drugs (digoxin, 9; amiodarone, 6; enoximone, 3; other vasodilator agents, 3; xamoterol, 1; and flecainide 1). No patient had taken warfarin, aspirin, or nonsteroidal antiinflammatory agentsin the 2 weeksbefore the study. Blood samples. Patients fasted for 12 hours and discontinued medication, including ACE inhibitors, for 24 hours before sampling. Sampleswere taken at 10:00 AM, becauseplatelet aggregability has a circadian variation,16 after 30 minutesof supinerest and from an antecubital vein without a tourniquet. The first 5 ml samplewasdiscarded, and the next samplewasusedonly if satisfactory flow was achieved. Samples were transferred to appropriate prechilled bottles, immediately placed on ice, then centrifuged at 3000 rpm at 4’ C for 10 minutes. Supernatant plasma was stored at -75O C until assayed. Laboratory investigations Conventional coagulation screen. Thrombin, prothrombin, and activated partial thromboplastin time were measured with standard laboratory techniques (Diagnostic Reagents,Thame, Oxfordshire, U. K.). Viscosity. Blood viscosity wasmeasuredboth at the patient’s own (native) and at a corrected (standardized at 45% ) hematocrit, over a rangeof shearrates; however, only values at the two extremes of shearrate (0.277set-’ [low] and 128.5set-’ [high]) were considered.Adjustment to a standard hematocrit, performed by removal or addition of autologousplasma,allowed assessment of rheologic factors

March 1994 Hem Journal

other than hematocrit. Blood viscosity varies inversely with shear rate; at a low shear rate as red cells aggregateand form rouleaux, blood viscosity is high; whereasat a high shearrate red cellsdeform and viscosity decreases.Plasma viscosity was only measured at one shear rate, because plasmais a newtonian liquid. Individual hemostatic factors. Fibrinogen wasmeasured with the Claussmethod.17Fibrinopeptide A wasmeasured with the competitive enzyme-linked immunoassaytechnique (Asserachrom;Diagnostica-Stago,Asnieres,France) after treatment with bentonite to remove fibrinogen. D-dimer and P-thromboglobulin were measuredwith sensitive enzyme-linked immunosorbent assay techniques (Asserachrom,Diagnostica-Stago).Factor VII activity was measuredwith a calorimetric assay(Stachrom, Diagnostica-Stago). Von Willebrand factor antigen wasmeasuredby an enzyme-linked immunosorbent assaymethodls (Dako Corp., Carpenteria, Calif.). Other assays. Platelet count wasmeasuredwith a TOA Analyzer E-5000 (Sysmex, Kobe, Japan). Erythrocyte sedimentation rate (ESR) was measuredby the Westergren method. C-reactive protein wasmeasuredwith aTechnicon RA-1000 Analyzer (Miles Inc., Diagnostics Div., Tarrytown, N.Y.). Hematocrit, white blood cell count, serum urea, creatinine, and electrolytes were measured with standard laboratory techniques. Neuroendocrine profile. Epinephrine and norepinephrine were measuredby high-performance liquid chromatography.lg Angiotensin II and endothelin I were measured by radioimmunoassay.20, “’ Other investigations. Exercisetesting wasperformed on a bicycle, starting at a workload of 10 W/min and increasing by 10 W/min. The maximal oxygen consumption was measuredby massspectrometry. Multi-gated acquisition scanejection fraction wasderived from the anteroposterior and the 45-degreeleft anterior oblique views. Statistical analysis. Values are expressed as mean, (standard deviation), and range.Values are comparedwith the normal range for our laboratory. Student’s unpaired t test was used to compare data from patients with and without coronary artery disease.Values that were not normally distributed underwent log transformation before analysis.Correlationsbetweenthe various parameterswere performed by linear regressionanalysisor Spearman rank correlation as appropriate. A p value of lessthan 0.05 was consideredto be statistically significant. RESULTS The mean peak oxygen consumption for all patients was 13.3 -t 3.6 ml/kg/min, and the mean ejection fraction was 23% f 7%) which is consistent with the diagnosis of moderate or severe heart failure in the majority of subjects. Hematocrit and white blood cell and platelet counts were normal for all patients. The ESR was distinctly elevated in two patients (55 and 104 mm in the first hour). No evidence of infection, malignancy, or connective tissue diseasecould be discerned in these patients, both

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Fig. 1. Plasma concentrations of fibrinopeptide A, &thromboglobulin, D-dimer, and von Willebrand factor in patients with heart failure, divided according to NYHA class. Shaded areas indicate the mean values + 2 SD for healthy control subjects. Filled squares represent patients with dilated cardiomyopathy, and open circles represent patients with coronary artery disease.

of whom were in NYHA class IV heart failure. Postmortem examination, which was later available in each case, demonstrated clinically undetected pulmonary embolism, despite prophylactic use of subcutaneous heparin, but no other cause for the raised ESR. Four patients had an elevated C-reactive protein level (58,42,41, and 97 mg/L); two of them also had an elevated ESR. Again, no cause other than severe heart failure was clinically apparent. Viscosity (Table I). The patients’ mean plasma viscosity was significantly higher than control values (p < 0.05). Blood viscosity at the native hematocrit and at a low shear rate was also significantly higher compared with control values (p < 0.05). Blood viscosity at the native hematocrit but at a high shear rate and blood viscosity at a corrected hematocrit at both low and high shear rates were not statistically significantly higher than control values. Plasma viscosity correlated with plasma fibrinogen (r = 0.75; p < 0.01) and ESR (r = 0.93; p = 0.001). Blood viscosity at the corrected hematocrit also correlated well with ESR (r = 0.8; p < 0.01) and fibrinogen (r = 0.76; p < 0.01).

Hemostatic variables (Table I and Fig. 1). The conventional coagulation screen revealed no significant differences between values for patients with heart failure and control values. Plasma concentrations of fibrinopeptide A were elevated in seven of 21 patients, the highest level being six times the control value (Fig. 1). Fibrinopeptide A correlated with fibrinogen (r = 0.72; p < 0.05). Differences in fibrinopeptide A concentration were not explained by the severity of heart failure or plasma concentrations of any other measured variable. The plasma concentration of P-thromboglobulin was elevated in all patients (Fig. 1). P-Thromboglobulin was higher in patients with heart failure caused by dilated cardiomyopathy than in those with coronary disease (p = 0.035) but showed no relationship to the severity of heart failure or any other measured variable. Concentrations of D-dimer were elevated above the normal range in 13 of 21 patients (Fig. l), tending to be higher in those with more severe heart failure. D-dimer concentrations were not related to any other measured variable. Plasma concentrations of von Willebrand factor

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DISCUSSION

0

1

2

von Willebrand

3

4

5

Factor (U/ml)

Fig. 2. Relationship between the plasma concentrations

of von Willebrand factor antigen and immunoreactive endothelin in 21 patients with moderate to severeheart failure (r = 0.54; p < 0.01).

antigen were elevated above the control range in seven of 21 patients (Fig. 1). Plasma von Willebrand factor was directly related to plasma concentrations of endothelin (r = 0.54; p < 0.02). Plasma concentrations of von Willebrand factor appeared to be related to NYHA class (Fig. l), but this did not achieve statistical significance. Factor VII activity was within normal limits for all patients. Plasma concentrations of fibrinogen were elevated in one of 21 patients. Neuroendocrine variables (Table I). Plasma concentrations of angiotensin II and endothelin were increased compared with the normal range in all patients, norepinephrine was increased in all but two patients, and epinephrine was within normal limits for most patients. Plasma concentrations of endothelin and angiotensin II were related (r = 0.54; p < 0.02). Other investigations. The mean serum creatinine level was 123 +- 31 pmol/L (range, 78 to 195 pmol/L) and was unrelated to the plasma concentration of P-thromboglobulin. Effects of causes of heart failure. There were no significant differences between patients with dilated cardiomyopathy and those with ischemic heart disease with respect to demographic characteristics or hemostatic or neuroendocrine indices other than ,8-thromboglobulin. Effect of severity of heart failure. Severity of heart failure as determined by peak oxygen consumption, ejection fraction, and plasma norepinephrine was unrelated to hemostatic variables.

This study shows that many patients with moderate to severe heart failure have increased procoagulant activity and/or activation of fibrinolysis and platelets. Our patients had no evidence of a recent clinically apparent thromboembolic event. Although occult thromboembolic events could not be excluded and may have been responsible for altered hemostatic function, this would not detract from the relevance of our findings, because detection and treatment of occult thrombotic events might be of great importance to Clinical practice. The routine coagulation screen, as well as the plasma concentrations of fibrinogen and factor VII activity, did not reveal significant abnormalities, possibly reflecting the low sensitivity of these assessments. Plasma viscosity and blood viscosity at the native hematocrit and low shear rate were significantly higher in patients than in control subjects. This suggeststhat increased viscosity might contribute to the increased thrombotic tendency in the venous circulation and the atria1 appendages of patients with heart failure. The plasma concentration of P-thromboglobulin was consistently increased in our patients (Fig. l), suggesting activation of platelets either caused by endothelial dysfunction, as suggested by elevated levels of endothelin and von Willebrand factor, or thrombin activity, as suggested by the increased levels of fibrinopeptide A in some patients. Renal dysfunction, hyperlipidemia, and acute-phase reaction responses can also increase /3-thromboglobulin levels 22but no correlation between creatinine, ESR, or C-reactive protein and P-thromboglobulin was found in the present study. The plasma concentration of D-dimer was also increased in most patients (Fig. l), reflecting enhanced fibrinolysis. This may be secondary to intravascular fibrin formation and subsequent plasmin activation, but fibrinopeptide A levels were not consistently increased. However, fibrinopeptide A has a short halflife (3 to 5 minutes), compared with D-dimer (2 hours) and p-thromboglobulin (100 minutes); therefore, a possible role of intermittent activation of coagulation cannot be excluded. High D-dimer levels could also represent a primary enhancement of fibrinolysis caused by liver dysfunction. Our findings support those of Jafri et a1.‘3who also found that /3-thromboglobulin and D-Dimer, but not fibrinopeptide A, were elevated in patients with heart failure compared with control subjects. In contrast to our study, they suggested that patients with more severe heart failure also had more disturbed hemostasis, though the relationship appeared weak.

Volume 127, Number 3 American Heart Journal

Plasma concentrations of von Willebrand factor, a mediator of platelet adhesion at sites of vascular injury that is secreted by the endothelium and by activated platelets, were also elevated in some patients (Fig. 1). The correlation between von Willebrand factor and P-thromboglobulin levels was poor, suggesting that platelets were not the main source of the elevated von Willebrand factor level. The association with plasma endothelin suggests that increased plasma concentrations of von Willebrand factor may reflect increased release from the endothelium (Fig. 2), although higher levels of von Willebrand factor could also be due to reduced plasma clearance. When the patients with dilated cardiomyopathy were compared with those with coronary disease, no significant differences in hemostatic variables were found between the two groups, other than values for P-thromboglobulin. This suggests that the observed hemostatic abnormalities are related to the heart failure per se rather than to the underlying cause. Plasma concentrations of von Willebrand factor and endothelin have been shown to correlate with pulmonary vascular resistance in patients with mitral stenosis and stable heart failure, respectively.24, 25 Raised levels of von Willebrand factor and endothelin and the hemodynamic disturbances might all result from the same stimulus; alternatively, increased vascular resistance may lead to greater expression or reduced clearance of von Willebrand factor and endothelin. The significant correlation between von Willebrand factor and endothelin (Fig. 2) does not necessarily establish a causal relationship because both may be independent markers of endothelial damage.26 The reason for the increase in plasma endothelin in heart failurez7* 28 remains uncertain. Angiotensin II stimulates endothelin production and release in cell cultures,2g though administration of pathophysiologic doses of angiotensin II to normal volunteers is only a modest stimulus to endothelin production.30 Angiotensin II in concentrations observed in heart failure can cause cardiac myocyte necrosis31 and it is also possible that it could damage the vascular endothelium. Alternatively, the same stimulus to von Willebrand factor and endothelin production could lead to increased production of angiotensin II by endothelium. A unifying explanation for our findings may reside in the effects of neuroendocrine activation on the hemostatic system. Catecholamines, angiotensin II, and antidiuretic hormone enhance platelet aggregability.“-7 Catecholamines and antidiuretic hormone increase circulating levels of von Willebrand factor and trigger tissue plasminogen activator release.3, 4 Neu-

Sbarouni et al. 611

roendocrine variables have been shown to predict outcome in patients with heart failure.32 Although neuroendocrine activation increases as heart failure becomes more severe, this relationship is not variables strong,33, 34 and therefore neuroendocrine are independent prognostic markers.32, 35 If hemostatic abnormalities in patients with heart failure are related to neuroendocrine activation and are one of the mechanisms leading to an earlier demise, this might be altered by administration of ACE inhibitors. Indeed, two studies have now shown that ACE inhibitors can reduce the risk of myocardial infarction in patients with left ventricular dysfunction, with or without heart failure.g* 36 In conclusion, our data indicate that heart failure is associated with an altered hemostatic state. Whether altered hemostasis reflects activation of coagulation and platelets leading to an increased risk of thromboembolic events, is simply a marker of occult thromboembolic events, or is due to a combination of the two remains to be determined. REFERENCES 1. Fuster

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