Prognostic Value of Coronary Vascular Endothelial Dysfunction
Background— Whether patients at increased risk can be identified from a relatively low-risk population by coronary vascular function testing remains unknown. We investigated the relationship between coronary endothelial function and the occurrence of acute unpredictable cardiovascular events (cardiovascular death, myocardial infarction, stroke, and unstable angina) in patients with and without coronary atherosclerosis (CAD).
Methods and Results— We measured the change in coronary vascular resistance (ΔCVR) and epicardial diameter with intracoronary acetylcholine (ACh, 15 μg/min) to test endothelium-dependent function and sodium nitroprusside (20 μg/min) and adenosine (2.2 mg/min) to test endothelium-independent vascular function in 308 patients undergoing cardiac catheterization (132 with and 176 without CAD). Patients underwent clinical follow-up for a mean of 46±3 months. Acute vascular events occurred in 35 patients. After multivariate analysis that included CAD and conventional risk factors for atherosclerosis, ΔCVR with ACh (P=0.02) and epicardial constriction with ACh (P=0.003), together with increasing age, CAD, and body mass index, were independent predictors of adverse events. Thus, patients in the tertile with the best microvascular responses with ACh and those with epicardial dilation with ACh had improved survival by Kaplan-Meier analyses in the total population, as did those in the subset without CAD. Similar improvement in survival was also observed when all adverse events, including revascularization, were considered. Endothelium-independent responses were not predictive of outcome.
Conclusions— Epicardial and microvascular coronary endothelial dysfunction independently predict acute cardiovascular events in patients with and without CAD, providing both functional and prognostic information that complements angiographic and risk factor assessment.
Received April 15, 2002; revision received May 21, 2002; accepted May 24, 2002.
The development of acute coronary syndromes is often abrupt and dependent on the rupture of a nonobstructive atheromatous plaque. Despite extensive research, a reliable independent predictor of the risk of incurring an acute vascular event has failed to emerge. The endothelium plays a critical role in vascular homeostasis by secreting substances that not only acutely regulate vascular tone, platelet activity, and coagulation factors but also influence vascular inflammation, cell migration, and proliferation over the longer term.1 Endothelial function may be determined in vivo by measuring the vasomotor response to pharmacological and physiological stress. Subjects with coronary artery disease (CAD) and those with risk factors typically demonstrate endothelial dysfunction, and although its extent appears to correlate with the traditional risk factor “burden,” there is considerable heterogeneity in the magnitude of dysfunction observed in individuals with similar risk factor profiles.2–4⇓⇓ Novel risk factors for atherosclerosis and endothelial dysfunction, some as yet undiscovered, presumably account for this observed variability. Therefore, endothelial vasodilator function, a test that reflects the overall impact of these multiple environmental and genetic factors on the vasculature, could potentially serve as a useful diagnostic and prognostic tool.
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In agreement with this expectation, recent studies have demonstrated that coronary vascular dysfunction independently predicts adverse long-term cardiovascular prognosis where the majority of “adverse events” were revascularization procedures.5,6⇓ We therefore undertook the present study to investigate whether the presence of coronary vascular endothelial dysfunction in patients with angiographically normal coronary arteries (NCAs) or those with CAD predicted a worse long-term outcome from unpredictable acute cardiovascular events, including death, myocardial infarction, stroke, and unstable angina.
We studied 308 subjects undergoing cardiac catheterization for investigation of chest pain or abnormal noninvasive cardiac investigations at the National Institutes of Health Clinical Center. The study was approved by the Institutional Review Board, and written, informed consent was obtained from each subject. Subjects with unstable angina, recent myocardial infarction (≤3 months), severe valvular disease, New York Heart Association class III to IV heart failure, or unrevascularized 3-vessel or left main disease were excluded. Subjects who were referred for revascularization after cardiac catheterization were also excluded.
CAD was defined as angiographic evidence of irregularity, plaque formation, or stenosis in any epicardial coronary artery. Patients with angiographically smooth coronary arteries represented a group with NCAs. Cardiac risk factors assessed included age, sex, presence of left ventricular dysfunction, tobacco smoking, diabetes, hypercholesterolemia, hypertension, and body mass index. Smokers included subjects with a current or prior history of tobacco use. Diabetes was defined as a fasting blood glucose level >140 mg/dL or treatment with dietary modification, insulin, or oral hypoglycemic agents at the time of the study. Hypercholesterolemia was defined as a fasting serum total cholesterol >240 mg/dL or if the subject was being treated with lipid-lowering medication or dietary modification. Hypertension was defined as a seated systolic blood pressure ≥140 mm Hg or diastolic pressure ≥90 mm Hg on at least 3 occasions, or if such a diagnosis had been made in the past and the patient was being treated with medications or lifestyle modification. Left ventricular dysfunction was defined as resting left ventricular ejection fraction <40% by radionuclide angiography or echocardiography. Body mass index was calculated as weight (kg) divided by height (m)2.
Coronary Vascular Function
All cardiac medications were withdrawn ≥48 hours and at least 5 half lives before the study. ACE inhibitors and aspirin or other cyclooxygenase inhibitors were discontinued ≥7 days before the study. After diagnostic coronary angiography, a 3F infusion catheter was introduced via a 7F guide catheter into a nonobstructed coronary artery (stenosis severity ≤30%). Blood flow velocity was measured, as previously described, with a 0.014- or 0.018-in Doppler wire (FloWire, EndoSonics).7 To calculate coronary blood flow (CBF), epicardial diameter was measured in a 0.25- to 0.5-cm segment of vessel beginning 0.25 cm beyond the tip of the flow wire. Coronary vascular resistance (CVR) was calculated as mean arterial pressure divided by CBF. Quantitative angiography was performed by an observer blinded to the identity of the study infusion with ARTREK software (Quantim 2001, StatView, ImageComm Systems, Inc) or the PIE medical CAAS system.3 Additionally, mid and distal segments of epicardial coronary arteries were also measured after each intervention, and epicardial coronary responses were determined by assessment of the percent change in diameter (ΔDiam), with each drug compared with baseline, in a segment of the mid or distal portion of the study vessel that was straight and free of overlap or major branch points.
Endothelium-dependent vasodilation was determined during a 2-minute intracoronary infusion of acetylcholine (ACh) at 15 μg/min (estimated concentration 10−6 mol/L). Ten minutes after the ACh infusion, endothelium-independent function was estimated during a 3-minute intracoronary infusion of 20 μg/min sodium nitroprusside (SNP, n=241), followed by measurement of vasodilator reserve with a 2-minute infusion of 2.2 mg/min of adenosine (n=270). Study drug doses were doubled when infused directly into the left main-stem coronary artery.
Long-term follow up was performed either by a questionnaire or by telephone contact. All cardiovascular events were validated by review of medical records, ECG, and cardiac enzyme data. The minimum follow-up period was 6 months. For subjects experiencing more than 1 acute event, only the first event was considered in the analysis.
The combined acute cardiovascular event end points assessed during follow-up included cardiovascular death, acute myocardial infarction, unstable angina pectoris, and acute ischemic stroke. Cardiovascular death was defined as death due to a myocardial or cerebral infarction or documented sudden cardiac death. Myocardial infarction was defined as an elevation of creatine kinase levels >2 times the upper limit with or without new ST-segment elevation (>0.1 mV) in at least 2 contiguous leads. Unstable angina pectoris was defined as hospitalization because of angina pectoris that occurred at rest and that was associated with ECG changes (Braunwald classification IIB or IIIB). Ischemic stroke was defined as clinical and radiological evidence of stroke without intracranial hemorrhage.
Data are expressed as mean±SEM. Differences between means were compared by unpaired Student’s t test. Cumulative acute cardiovascular event rates were estimated by Kaplan-Meier survival curves (Lifetest procedure of SAS8). Probability values for survival curve comparisons were calculated with the log-rank statistic. Cox proportional hazards multivariate stepwise regression analysis (SAS) was used to determine the univariate and multivariate relationships between clinical variables and acute cardiovascular events during the follow-up period. Covariates entered in this regression model were ΔCVR with ACh, SNP, or adenosine; ΔDiam with SNP or epicardial dilation/constriction with ACh; and presence of coronary atherosclerosis, age, sex, presence of left ventricular dysfunction, diabetes, smoking, hypertension, hypercholesterolemia, and family history of CAD. All probability values were 2-tailed, and a value <0.05 was considered statistically significant.
We studied 308 patients, 132 with CAD and 176 with NCAs (Table 1). Mean duration of follow-up was 46.3±2.7 months (median 40.7 months). During this period, 35 subjects (11.4%) experienced an acute cardiovascular event (Table 2). Six of these subjects had 2 or more acute events. Additionally, 21 subjects were electively referred for percutaneous (n=16) or surgical (n=5) coronary revascularization during follow-up (Table 2).
Clinical Determinants of Acute Cardiovascular Events
Increasing age, male sex, CAD, increased body mass index, hypercholesterolemia, diabetes, left ventricular dysfunction, and smoking were univariate predictors of acute cardiovascular events by Cox proportional hazards analysis (Table 3). CAD, increasing age, and increased body mass index remained significant independent predictors of acute events after multivariate analysis adjusted for other covariates in the full model, with a nonsignificant trend observed for smokers (P=0.07; Table 3).
Coronary Vascular Function and Acute Cardiovascular Events
Microvascular dilation with ACh was attenuated in subjects with acute events during follow-up compared with those without events (CBF increase 67±12% and CVR fall of 28±6% compared with 114±6% increase in CBF [P=0.007] and 46±2% fall in CVR [P=0.007] in subjects with and without events, respectively). To study the relationship between coronary microvascular endothelial function and acute cardiovascular events, the study population was divided into tertiles according to the ΔCVR with ACh. By Kaplan-Meier analysis, event-free survival from acute cardiovascular events was significantly associated with the vasodilator response to ACh (P=0.047), with outcome being worse in those with a more depressed vasodilator response (Figure 1). When the 2 tertiles with impaired responses to ACh were compared with the best-responding tertile, the difference in outcome was more significant (P=0.017). In contrast, when the population was similarly divided into tertiles according to response to the endothelium-independent vasodilators, no significant association was observed between event-free survival from acute cardiovascular events and ΔCVR with either SNP or adenosine (P=0.11; Figure 1).
After multivariate adjustment for all noted cardiac risk factors, ΔCVR with ACh remained an independent predictor of event-free survival (P=0.019), whereas responses to SNP and adenosine were not (P=0.24 and P=0.08, respectively). To determine the value of coronary microvascular endothelial function in predicting acute cardiovascular events, we compared event rates in subjects in the top versus the lower 2 tertiles of the ACh response (increase in CBF was <130% and fall in CVR was <56% in response to ACh in patients in the lower 2 tertiles). The event rate of patients in the lower 2 tertiles was 14.1% compared with 5.8% for those in the upper tertile.
The frequency of all cardiovascular events that included both elective revascularization procedures and acute events was also investigated in the 3 tertiles according to ΔCVR with ACh. Event-free survival from all cardiovascular events was also significantly worse in the 2 tertiles with relatively impaired coronary microvascular endothelial function than in those in the tertile with preserved function (P=0.035; Figure 2). Finally, when only death, myocardial infarction, or stroke were considered as events, event-free survival was also significantly related to endothelial function (P=0.037 by Kaplan-Meier analysis; Figure 2).
To study the relationship between acute events and epicardial endothelial function, the study population was divided into 2 groups: those who constricted with ACh (ΔDiam <0%, n=153), which denoted endothelial dysfunction, and those who dilated (ΔDiam ≥0%; n=138), which denoted preserved endothelial function.1,3–5⇓⇓⇓ The incidence of acute events was significantly greater in subjects with epicardial constriction than in those who dilated with ACh (P=0.003 by Kaplan-Meier analysis; Figure 3). The event rate in those with a constrictor response was 13% compared with 9.4% in those with dilation in response to ACh. However, when the population was divided into tertiles according to ΔDiam with the endothelium-independent vasodilator SNP, there was no difference in event-free survival from acute coronary vascular events during follow-up (P=0.33; Figure 3). After multivariate adjustment for risk factors, epicardial vasodilation with ACh remained an independent predictor of event-free survival (P=0.019), whereas ΔDiam with SNP was not (P=0.71).
The frequency of all cardiovascular events including revascularization procedures was also investigated according to the epicardial response with ACh. Event-free survival from all events was significantly worse in those exhibiting epicardial vasoconstriction with ACh than in those who had vasodilation in response to ACh (P<0.0001; Figure 2). Additionally, when only death, myocardial infarction, or stroke were considered as events, event-free survival was also significantly worse in those who had vasoconstriction with ACh (P=0.038; Figure 2).
Outcome in CAD and NCA Patients
Because of the observed independent relationship between the presence of CAD and worse prognosis, we investigated whether the relationship between endothelium-dependent vasomotion and prognosis was independent of the presence or absence of CAD by introducing interaction terms (CAD × ΔCVR or CAD × epicardial constriction/dilation with ACh) in the regression analysis. No significant interaction was observed (P=0.87 and P=0.77, respectively), which indicated that endothelial dysfunction predicted outcome independent of the presence of CAD.
To further determine the relationship between coronary vascular endothelial function and prognosis, we investigated whether ΔCVR with ACh also predicted outcome in the subset of 171 subjects with NCAs. When this subgroup was divided into tertiles according to ΔCVR with ACh, the tertile with the best response had a better event-free survival than those in the 2 lower tertiles (Figure 4). When patients with NCAs were divided into 2 groups according to whether they exhibited a dilator or constrictor epicardial response to ACh, a nonsignificant trend toward a better event-free survival from acute coronary events was observed in those who had dilation with ACh (Figure 4).
Because aspirin, ACE inhibitors, and HMG-CoA reductase inhibitors may alter endothelial function and are commonly used to treat subjects with CAD, Kaplan-Meier analyses were performed to assess the influence of these therapies. No differences in the incidence of acute events were observed in patients with CAD who were taking ACE inhibitors (P=0.372), HMG-CoA reductase inhibitors (P=0.275), or aspirin (P=0.296) compared with those not taking these medications. There were no differences in the use of these agents in patients divided into tertiles based on their ACh response or in those with epicardial constriction or dilation in response to ACh.
We have shown that endothelium-dependent coronary vasomotor function independently predicts the long-term risk of spontaneously occurring acute cardiovascular events, including sudden cardiac death, myocardial infarction, and cerebral infarction, after adjustment for presence of CAD and other cardiac risk factors. Not only was an association observed between an increased risk of acute events and endothelial dysfunction of both the epicardial and microvascular coronary circulation, but this relationship remained statistically significant even when the criteria used to define events were made less rigorous by inclusion of either unstable angina or elective revascularization. In contrast, endothelium-independent epicardial or microvascular vasodilator responses with adenosine or SNP were not predictive of cardiovascular events. Thus, clinical assessment of coronary endothelial function provides important independent prognostic information that may have a potential role in the long-term management of subjects with mild to moderate CAD who are managed medically or those with NCAs and risk factors for atherosclerosis.
Based on our findings, within our follow-up period, 14% of patients in the lower 2 tertiles of the microvascular ACh response (CBF increase <130%) had acute events, and 21% had events including revascularization (positive predictive value). Thus, 83% of all acute events and 77% of events including revascularization occurred in patients within these lower 2 tertiles. Moreover, 94% of patients in the top tertile remained free of acute events, and 87% were free of all events, including revascularization. Similar findings were observed when epicardial responses to ACh were considered.
The acute event rate observed in the cohort in the present study was 3% per year, similar to the event rate observed in the HOPE (Heart Outcomes Prevention Evaluation) trial, which recruited a study population with similar clinical characteristics to ours.9 Although an adverse cardiovascular prognosis has been described in subjects with coronary vascular dysfunction in 2 previous studies, both study populations were approximately half the size of the present cohort, subjects with NCAs were underrepresented, a relatively small number of events occurred, and a substantial proportion of the reported events were revascularization procedures.5,6⇓ Suwaidi et al6 presented the prognostic value of coronary microvascular responses to ACh without independently determining the value of endothelium-independent or diameter responses. Furthermore, Schachinger et al5 limited their observations to the epicardial coronary circulation, where the endothelium-independent response to SNP appeared to be a better predictor of prognosis than the ACh response. In contrast, the present study population was considerably larger; we separately analyzed acute unpredictable cardiovascular events; the population with NCAs could be studied independently; and we clearly demonstrated differences in the association between endothelium-dependent and -independent function and long-term cardiovascular prognosis.
The vascular endothelium secretes multiple factors that not only regulate vascular tone but also modulate platelet activity, the endogenous thrombolytic system, vascular inflammation, and cell migration and proliferation.1 Because the coronary vasodilator response to ACh is predominantly caused by release of NO from endothelial cells, and the impaired response to ACh is primarily caused by reduced bioactivity of NO, we used this measure as an index of endothelial health.3,4⇓ We now show that an increased risk of acute lesion progression leading to cardiovascular events may be predicted by the health of the endothelial lining in both the conductance and resistance vessels. This scenario offers new hope for identifying a subgroup of patients at increased risk from within a cohort of low-risk subjects with and without CAD.
Endothelial cells appear to integrate the injury imposed by exposure to the commonly known risk factors of CAD, the more novel factors such as infections, and other as yet undiscovered genetic and environmental risk factors.10,11⇓ Endothelial dysfunction is associated with increased inflammation,12 thrombogenicity,13–16⇓⇓⇓ and enhanced local expression of matrix metalloproteinases,17–19⇓⇓ which together increase the vulnerability of atherosclerotic plaques to rupture and thrombus formation, thus providing a mechanistic link between endothelial dysfunction and adverse cardiovascular outcome. Whether patients with endothelial dysfunction have more rapidly progressive atherosclerosis, as suggested by our data, will require confirmation in a prospective angiographic study. Likewise, these data raise another intriguing question: will interventions such as exercise, diet, and pharmacological therapy that are designed to enhance endothelial function also improve prognosis, as is suggested by outcome trials with aspirin, ACE antagonists, and statin therapy?9,20–22⇓⇓⇓
This study was conducted retrospectively and will therefore require confirmation in a prospective investigation. Our results cannot be extended to all subjects with CAD, because we included only medically managed patients with asymptomatic or mildly symptomatic CAD of mild to moderate severity. Unlike previous studies, patients who were referred for revascularization after angiography were not included, to avoid bias introduced by the revascularization strategy.5 Although we excluded subjects with more severe CAD, the independent association between significant multivessel CAD and an adverse cardiovascular prognosis has been well established in such patients.
We are unable to rule out intimal thickening or eccentric plaque formation in the present NCA cohort. It is likely that a number of these patients who also had exposure to conventional risk factors indeed had early atherosclerosis. However, intimal thickening demonstrated by ultrasound has not been helpful in predicting cardiovascular events in subjects with nonobstructed coronary arteries.6 Nevertheless, the present data provide an important clinical lesson: the presence of angiographically smooth epicardial coronary arteries can be associated with endothelial dysfunction, and the degree of dysfunction is an important predictor of outcome in these patients.
Conclusions and Clinical Implications
We have demonstrated a strong association between coronary vascular endothelial dysfunction and an adverse long-term cardiovascular prognosis. Both conduit arterial and microcirculatory endothelial function are predictive of outcome independent of the presence of CAD and its risk factors. Thus, assessment of endothelial function or its markers may help identify a subgroup of patients at high risk. Whether strategies that improve endothelial dysfunction will uniformly improve prognosis needs to be studied prospectively.
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