Acetylcholine provocation test

The first investigations for CAS included typical clinical history, non-invasive stress tests such as treadmill test, ambulatory electrocardiogram (ECG) and stress echocardiography, with subsequent invasive angiogram performed for confirmation. CAS was induced by intracoronary injection of ACh after diagnostic angiography. Nitrates, calcium channel blockers (CCB), beta blockers (BB), angiotension-converting enzyme inhibitors (ACEI), and other vasodilators or vasoconstrictors were withheld at least 72 h before coronary angiography. ACh was injected by incremental doses of 20 (A1), 50 (A2), and 100 (A3) μg/min into the left coronary artery over a 1 min period with 5-min intervals to the maximum tolerated dose under continuous monitoring of ECG and blood pressure. Angiography was repeated after each ACh dose. Intracoronary infusion with 0.2 mg nitroglycerin was given after completing the ACh provocation test. Angiography was then performed 2 min later. If focal or diffuse significant vasoconstriction of coronary arteries was induced with any dose of ACh, the ACh infusion was stopped. End systolic images for each segment of the left coronary artery were chosen according to the corresponding points on the ECG trace (QRS onset or end of T wave) and analyzed using the proper quantitative laboratory (FD-20, Phillips, Amsterdam, The Netherlands). The coronary artery diameters were measured with quantitative coronary angiography (QCA) before and after administration of ACh at the site that showed the greatest changes following drug administration. Reference diameter was measured at the proximal and distal portion of each artery before intracoronary ACh infusion and the mean reference diameter was used to assess the diameter narrowing. During the coronary angiography and ACh provocation test, significant CAS was defined as focal or diffuse severe transient luminal narrowing (>70%) with or without chest pain or ischemic ECG change such as ST-T segment elevation, depression (≥1 mm), or T wave inversion, and normal coronary appearance was defined as less than 20% luminal narrowing on coronary angiogram that was measured with QCA. Myocardial bridge was considered when the characteristic phasic systolic compression of the coronary artery had a >30% decrease in diameter on the angiogram after intracoronary nitroglycerin infusion. Multivessel spasm (MVS) was defined as CAS of ≥2 major epicardial coronary arteries. The presence of baseline spasm was defined as focal or diffuse narrowing more than 30% in diameter on the angiogram before the ACh provocation test as compared with the final angiogram after intracoronary nitroglycerin infusion. Diffuse CAS was defined as significant CAS site length ≥20 mm.

Study definition

Hypertension was defined as either systolic or diastolic elevation of blood pressure ≥140/90 mmHg or ongoing antihypertensive pharmacological treatment. Dyslipidemia was defined as a total cholesterol level ≥200 mg/dL or current treatment with lipid-lowering drugs. Current smoking was defined as active smoking within the past 12 months. Diabetes mellitus was defined as the fasting blood glucose level ≥126 mg/dL, or use of hypoglycemic agents or insulin. In the present study, if a patient's past history, medical records, present symptoms, or medical examination results accorded with one of the following criteria, the patients were diagnosed with peripheral arterial disease (PAD): (1) Claudication symptoms with ankle brachial indices <0.90; (2) Claudication symptoms with findings of a significant lesion (≥70% diameter stenosis) in peripheral artery on computed tomographic angiography, magnetic resonance angiography, or invasive angiography; (3) symptomatic carotid, subclavian arterial disease (≥70% diameter stenosis) documented by image studies including computed tomographic angiography, magnetic resonance angiography, or invasive angiography. Insignificant coronary artery disease was defined as ≤20% diameter stenosis in coronary arteries documented by image studies including computed tomographic angiography, magnetic resonance angiography, or invasive angiography.

Statistical analysis

All the statistical analyses were performed using SPSS 15.0 (SPSS Inc., Chicago, IL, USA). For continuous variables, differences between two groups were evaluated by unpaired t-test or Mann-Whitney rank test. For discrete variables, differences were expressed as counts and percentages and analyzed with χ² or Fisher's exact test between groups as appropriate. Multivariate logistic regression analysis, which included baseline confounding factors, was used for assessing the independent impact factors. A two-tailed p-value of <0.05 was considered to be statistically significant. Data were expressed as mean ± standard deviations.

Results

The baseline clinical characteristics of patients are shown in Table 1. The prevalence of old age, diabetes mellitus, hypertension, hyperlipidemia, PAD, and cerebral vascular disease (CVD) were higher in the aspirin group. Whereas the level of total cholesterol, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol were lower in the aspirin group, probably due to a higher rate of using lipid-lowering drugs in the aspirin group than in the no aspirin group. Before the ACh provocation test, CCB, BB, and nitrate were more frequently prescribed in the aspirin group, and these results reflect the real world medication treatment of previous LDA users.

The clinical and angiographic parameters of the ACh provocation test are shown in Table 2.

Before the ACh provocation test, at baseline coronary angiogram, the percentage of baseline spasm and myocardial bridge were similar between the two groups. During the ACh provocation test, the incidence of ACh-induced CAS and ischemic chest pain were higher in the aspirin group as compared to no aspirin group. The QCA results are shown in Table 2. Before the ACh test, the baseline epicardial reference diameters after nitroglycerin infusion were similar between the two groups. After ACh infusion, the mean % narrowing by QCA was similar between the two groups. The results for response to the ACh doses and spasm extent are shown in Table 2. The aspirin group required lower ACh dose (50 μg, A2) and the incidence of MVS was higher in the aspirin group.

We evaluated independent predictors for ACh-induced CAS by adjusting possible confounding factors including age, gender, hypertension, diabetes mellitus, hyperlipidemia, current smoking, CVD, PAD, medication history including the previous use of LDA and ACEI, baseline spasm, and myocardial bridge. The results are listed in Table 3. Multivariate analysis showed that the previous use of LDA was an independent predictor of CAS induced by ACh test (adjusted odds ratio, 1.6, 95% confidence interval, 1.0–2.3; p = 0.031). Further, male, old age, lipid-lowering drugs, baseline spasm, and myocardial bridge were also independent predictors of ACh-induced CAS.

Discussion

The main finding of this study is that the patients taking LDA was more frequently associated with ischemic symptoms, CAS, as well as severe and MVS, suggesting this particular subset of patients would require more intensive medical therapies and close follow up.

However, in this study, the prevalence of risk factors for atherosclerosis including old age, diabetes mellitus, hypertension, and hyperlipidemia were higher in the aspirin group. The previous use of LDA was associated with a higher incidence of ACh-induced CAS, frequent ischemic symptoms, MVS, and showed vulnerable response to lower doses of ACh provocation test as compared to the non LDA group. Therefore, it is likely that the association of LDA and CAS that we have observed is not causal but may be hypothesis generating due to significant baseline differences between the two groups. Nevertheless, this study shows that the patients taking the LDA have a higher number of risk factors for atherosclerosis in the real world and would be associated with a higher incidence of clinically significant CAS, frequent ischemic symptoms, MVS, and showed vulnerable response to lower ACh dose. Thus, patients who have received LDA before ACh test would require special attention during ACh provocation test, more intensive medical therapies including antianginal agents, and close clinical follow up. Since the first description of variant angina by Prinzmetal et al. in 1959 [6], the epidemiology of CAS had been studied extensively. Recently, the endothelial dysfunction induced by abnormalities of endothelial nitric oxide synthase (eNOS) and enhanced contractility of vascular smooth muscle in coronary artery have been considered major mechanisms of CAS [7].

Aspirin is known to suppress the synthesis of prostaglandin and thromboxane A2 by blocking cyclooxygenase [8]. Prostaglandins and thromboxane A2 play a role in modulation of coronary artery. Prostaglandins induce vasodilatation of coronary artery and thromboxane A2 invoke vasoconstriction and platelet aggregation [9]. Therefore, unbalance in the levels of thromboxane A2 and prostaglandins would cause angina attacks in patients with coronary artery disease. Numano et al. reported that the patients with variant angina had a lower level of 6-keto prostacycline with statistically significant difference as compared to a control group [10]. But, the level of thromboxane B2 was similar between the two groups. This study suggested that the low level of prostacycline may be more of a contributing factor to CAS compared to high level of thromboxane B2. Miwa et al. reported that large amounts of aspirin (4.0 mg/day) aggravate the occurrence of CAS by the action of blocking the endothelial prostaglandin synthesis of the coronary artery[5]. The use of LDA has widely increased to prevent heart attacks and strokes in high-risk patients [1],[2] and [3]. However, the relationship between LDA and CAS was rarely reported. Robertson et al. reported that LDA has no effect on the frequency and duration of CAS [11]. But in the present study, the previous use of LDA was an independent predictor of CAS induced by intracoronary ACh provocation test. The previous LDA users had higher incidence of ACh-induced CAS, severe spasm extent such as MVS, and more frequent ischemic chest pain as compared to no LDA users.

In the present study, the previous LDA users had higher prevalence of conventional risk factors of atherosclerosis including diabetes mellitus, hypertension, hyperlipidemia, PAD, and CVD than no aspirin users, and this result suggests that in the real world, the previous LDA users have high risk factors of atherosclerosis and have the chance of much more severe endothelial dysfunction than expected. Thus, the incidence and severity of CAS, the degree of spasm extent, and clinical symptoms during the test were more frequent in the LDA users than no aspirin users.

In the present study, old age and myocardial bridge were also predictors of CAS. Old age has been reported to be a significant risk factor of endothelial dysfunction and induce significant CAS during ACh provocation test. In our previous study, myocardial bridge had been reported to be an independent predictor of focal spasm [12]. Myocardial bridge invokes longitudinal compression relaxation stress of coronary artery which induces the endothelial dysfunction and may increase the risk of CAS.

In this study, the presence of baseline spasm before ACh test was an independent predictor of CAS. However, the incidence of baseline spasm was similar between the LDA and no LDA groups. Further, multivariate analysis showed that the LDA was not an independent predictor of baseline spasm (adjusted odds ratio = 1.015, 95% confidence interval, 0.7–1.5; p = 0.943). These results showed that baseline spasm was not associated with LDA and LDA itself did not promote baseline spasm.

Lipid-lowering drugs have been shown to improve endothelial function, and would be thus expected to reduce the likelihood of CAS. However, in the present study, the use of lipid-lowering drugs was an independent predictor of ACh-induced CAS. The most reasonable explanation for this finding is that the lipid-lowering drugs were more commonly prescribed for patients with atherosclerosis, and it is more likely to be associated with higher incidence of risk factors requiring lipid-lowering drugs in the LDA group rather than a causal relationship.

In the present study, the previous LDA users were more frequently prescribed CCB and nitrate as compared to no aspirin users. Nevertheless, the previous LDA users had higher incidence of CAS during ACh provocation test as compared with non aspirin users. According to these results, we assumed that in the real world, the previous LDA users have much more chances of severe endothelial dysfunction than expected and would require more aggressive medical treatment using antianginal drugs than until now [13] and [14].

There are several limitations of this study. In this study, we used a more safe definition of significant CAS as compared to other previous studies related to ACh provocation test. We defined significant CAS as focal or diffuse severe transient luminal narrowing (>70%) in order to prevent the situation of hemodynamic collapse due to severe and intractable CAS. In an earlier period of our registry, we experienced some intractable CAS patients who did not experience quick relief by intracoronary nitroglycerin injection. Few patients experienced cardiogenic shock and cardiac arrest during the ACh provocation test. Our diagnostic criteria were established to ensure the safety of the patients. Our criteria are just for the clinical decision of whether we should treat or not. Thus, in this study, when we observe angiographically significant narrowing (>70%) regardless of the patient's severe chest pain or ST-T change in ECG during ACh provocation test, intracoronary nitroglycerin was injected instantly for the safety of the patients. Therefore, the incidence of ischemic chest pain and ECG change was relatively low. This policy was for the safety of patients and it did not negatively impact on our decision whether to treat or not. If we had ignored the patients’ chest pain or severe narrowing, did not inject nitroglycerin promptly, and increase ACh dose to A3 (100 μg/min), the incidence of ischemic chest pain and ECG change would be drastically increased.

The data for this study are the all comer data of Korean population and are not selectively enrolled data. These results are the observational findings in a real world, all comers-based registry and can be regarded as valuable characteristics found in the Korean population.

In this study, ACh spasm provocation test was performed only in the left coronary artery, not to both the left and right coronary arteries. In Asian countries including Korea, Japan, and China, the diagnostic coronary angiography is now largely being performed by transradial approach instead of transfemoral approach for its safety and shorter stay at the hospital. In this study, most patients in our center underwent coronary angiography and ACh provocation test through 4Fr radial approach for the convenience of study. If we perform the ACh provocation test in dominant left circumflex artery or directly into the right coronary artery, we will need the temporary pacemaker from the femoral vein due to advanced coronary atrioventricular block due to the ACh action. In that case, the patients need additional vascular access in the groin, will cost a lot for the temporary pacing, and further longer hospital stay. Therefore, for the safety of patients and convenience of study, it is difficult to perform both femoral and radial approach, and we performed the provocation test in left coronary artery alone. Further, our preliminary study data showed that the ACh provocation test into left coronary artery alone gives us reliable and definite information to decide the diagnosis and guide for medical therapy.

The data of this study were analyzed retrospectively and comprise the prospective real world spasm registry for all comers in the Korean population. Therefore, the result of this study was a hypothesis-generating conclusion and a larger prospective randomized study will be needed for definite conclusion regarding the impact of LDA on ACh provocation test results in the future.

The focus of this study is regarding the observation of the clinical and angiographic parameters during ACh provocation test, not the major clinical events throughout the longer clinical follow up. Because of the characteristics of the study population without PCI who are medically followed at outpatient department with relatively shorter-term follow up, the extensive information regarding clinical outcomes in the study population is not currently available. The clinical outcome information of CAS patients will be the future challenges of our study. However, at this moment, we focused on the index clinical and procedural parameters at the time of ACh provocation test.

Conclusion

The use of LDA has increased to prevent heart attacks and strokes in high-risk patients. However, it was not well known whether the previous use of LDA has an adverse impact on CAS. Despite the non-randomized nature and significant baseline differences, the previous LDA users were associated with higher incidence of ACh-induced CAS, more ischemic chest pain, MVS, and required lower ACh dose than no aspirin users, suggesting more aggressive medical treatment with special care would be needed.

References

Copyright © 2012 Japanese College of Cardiology. Published by Elsevier Ireland Ltd. All rights reserved.