Increased Angiotensin-Converting Enzyme Activity in Coronary Artery Specimens From Patients With Acute Coronary Syndrome
Background—Angiotensin-converting enzyme (ACE) inhibitors are effective in the secondary prevention of ischemic heart disease, but they do not reduce the rate of restenosis. Vascular ACE activity in the culprit coronary lesions of these patients, however, has never been quantified.
Methods and Results—We measured the ACE activity of vascular tissue obtained by directional coronary atherectomy in patients with acute coronary syndrome (n=17) and in patients with stable ischemic heart disease (n=36), with and without restenosis. The ACE activity of the culprit coronary lesions was significantly increased in patients with acute coronary syndrome (0.87±0.12 nmol · min–1 · mg protein–1; P<0.01) but not in patients with ischemic heart disease with restenosis (n=11, 0.19±0.05 nmol · min–1 · mg protein–1) when compared with those patients with ischemic heart disease without restenosis (n=25, 0.20±0.05 nmol · min–1 · mg protein–1). There was no difference between the ACE activity of the coronary tissue of the in-stent (n=5) and stent-unrelated (n=6) restenosis patients (0.24±0.10 versus 0.15±0.04 nmol · min–1 · mg protein–1). Serum ACE activity did not differ significantly among the patients.
Conclusions—The present study demonstrates increased ACE activity in culprit lesions in acute coronary syndrome, indicating that enhanced ACE activity is related to the causative mechanism of active coronary lesions.
Angiotensin-converting enzyme (ACE) inhibitors are effective in the secondary prevention of ischemic heart disease,1 but they do not reduce the rate of restenosis.2 3 Recently, increased immunohistochemical staining for ACE has been reported in atherosclerotic coronary tissue4 and in the culprit lesions of restenosis after balloon injury,5 and increased plasma ACE levels have been found in patients with in-stent restenosis.6 However, vascular ACE activity in culprit coronary lesions was never quantified. To test the hypothesis that the ACE activity of culprit coronary lesions is increased in unstable ischemic heart disease (IHD), we examined differences between ACE activity in coronary atherectomy specimens from patients with acute coronary syndrome (ACS) and stable IHD.
To obtain atherosclerotic plaque specimens from patients with symptomatic coronary artery disease, patients undergoing directional coronary atherectomy at Osaka Rosai Hospital who gave informed consent were included in the study. None of the patients had been taking ACE inhibitors. A total of 53 patients were enrolled: 17 patients (63±2 years) with de novo coronary stenosis presented with ACS (non–Q-wave infarction and unstable angina, Canadian Cardiovascular Society class III to IV), and 36 patients had stable IHD. In 25 of these 36 patients (63±1 years), symptoms were caused by de novo coronary lesions and, in the remaining 11 of the 36 patients (63±3 years), symptoms were due to restenotic lesions. Coronary specimens from ACS patients and from patients with IHD and restenosis were obtained within 1 week after admission and 5±1 months after percutaneous coronary intervention, respectively.
All patients were pretreated with aspirin, 81 to 162 mg daily, before atherectomy. Concomitant medical therapy, such as nitrates, calcium channel blockers, and β-adrenergic blocking agents, was continued at the discretion of the attending physician. Atherectomy was performed with the femoral approach using 10F arterial sheaths and guide catheters, and a directional coronary atherectomy catheter (AtheroCath-GTO, Devices for Vascular Intervention, Inc) of appropriate size to produce an approximate device-to-artery ratio of 1.1:1 was prepared. All atherectomy specimens were rapidly frozen in liquid nitrogen. Blood was withdrawn from all patients early in the morning on the day of catheterization.
Measurements of ACE Activity
ACE activity was measured using a previously reported method7 with slight modifications. Briefly, coronary tissue was homogenized on ice and centrifuged for 20 minutes at 1500g (4°C). ACE was assayed in a 250-μL final volume of a mixture of 100 mmol/L borate buffer (pH 8.3), 800 mmol/L NaCl, 50 μL of serum or tissue homogenate, and 3.5 mmol/L substrate (Hip-His-Leu, benzyloxy-carbonyl-glycyl-histidyl-leucine). The mixture was incubated at 37°C for 30 minutes (serum) or 120 minutes (tissue homogenates). The His-Leu–liberated amount was measured fluorometrically, with excitation at 360 nm and emission at 500 nm. ACE activity is expressed as the rate of His-Leu production (nanomoles per minute per milligram of protein). The protein content of the supernatant of the tissue homogenates was determined using the method of Lowry et al,8 with bovine serum albumin as the standard. Hip-His-Leu and His-Leu were obtained from the Peptide Institute.
Values are expressed as means±SEM. The significance of differences in the ACE activity of serum and coronary tissue or in age was determined by ANOVA followed by Scheffe’s test, as appropriate. The significance of differences in the baseline characteristics was determined by χ2 test. P<0.05 was considered statistically significant.
The baseline clinical characteristics of the study population are shown in Table⇓. The groups were generally well matched with respect to age, sex, and coronary risk factors.
ACE activity in coronary tissue was significantly higher in patients with ACS (n=17, 0.87±0.12 nmol · min–1 · mg protein–1, P<0.01) than in those with stable IHD, but it was not increased in coronary restenotic lesions (n=11, 0.19±0.05 nmol · min–1 · mg protein–1) compared with non-restenotic lesions (n=25, 0.20±0.05 nmol · min–1 · mg protein–1; Figure⇓). Different ACE activity was not observed in patients with in-stent (n=5) or stent-unrelated (n=6) restenosis (0.24±0.10 versus 0.15±0.04 nmol · min–1 · mg protein–1, P=0.072). The total protein content of the coronary tissue did not differ significantly between the groups (data not shown). There was no significant difference in serum ACE activity between patients with ACS (n=17, 0.34±0.03 nmol · min–1 · mg protein–1) and stable non-restenotic IHD (n=25, 0.32±0.03 nmol · min–1 · mg protein–1) or stable restenotic IHD (n=11, 0.31±0.03 nmol · min–1 · mg protein–1).
The results of this study demonstrate increased ACE activity in the culprit coronary lesions of ACS patients but not in stable IHD patients. Increased accumulation of ACE has been reported in the atherosclerotic coronary arteries of IHD patients.4 The discrepancy between such reports and our own observations may be attributable to differences in the sites in the vessels selected to evaluate ACE activity: atherectomy samples mainly consist of the thickened intima of atherosclerotic arteries. ACE activity in the media and adventitia may increase in atherosclerotic arteries in stable IHD.
ACE activity in blood vessel specimens obtained from ACS patients was significantly increased, indicating a pathophysiological role of increased ACE activity in the instability of atherosclerotic plaque. This is not unexpected, because ACE is a known local mediator of inflammation, and unstable coronary plaques have a strong inflammatory composition. In particular, macrophages are common components of the unstable plaque, and these cells strongly upregulate ACE activity during differentiation. The increased ACE activity observed in this study, therefore, may be a consequence of the higher prevalence of inflammatory cells in the samples obtained from ACS patients. Colocalization of ACE, angiotensin II, and the angiotensin II type 1 receptor with macrophages has been observed at the shoulder lesion of coronary atherosclerotic plaques and in the atherectomy tissue of patients with unstable angina.9 Angiotensin II–induced cytokines9 and oxidative stress10 11 may play a role in plaque instability.
Increased ACE expression was observed in the thickened neointima of restenotic lesions in an experimental model of endothelial injury.12 In humans, however, the ACE activity of restenotic neointimal lesions in this study had not increased when measured 5±1 months after coronary intervention, although the sample size was small. The increased ACE expression after balloon injury may be time-dependent: Ohishi et al5 reported that ACE expression is increased during the first 2 months after coronary intervention. However, with in-stent restenosis, increased ACE expression has been shown to persist for as long as 6 months after intervention.13 In our study, the ACE activity did not differ significantly between in-stent and stent-unrelated restenosis, although the former value exhibited a tendency to be high (P=0.072). A high number of diabetic patients (42%, 22 of 53) may influence the tissue characteristics of the restenotic plaques found in this study.
The present study is the first to report differences between the vascular ACE activity of the culprit lesions of patients with ACS and stable IHD. We concluded that increased ACE activity in lumen-sided coronary tissues may be involved in the pathophysiology of ACS.
This report was supported by the research fund from the Labor Welfare Corporation (to S. Hoshida, 2000), Kawasaki, Japan.
- Received October 23, 2000.
- Revision received December 13, 2000.
- Accepted December 19, 2000.
- Copyright © 2001 by American Heart Association
The MERCATOR Study Group. Does the new angiotensin converting enzyme inhibitor cilazapril prevent restenosis after percutaneous transluminal coronary angioplasty? Results of the MERCATOR study: a multicenter, randomized, double blinded, placebo-controlled trial. Circulation. 1992;86:100–110.
Diet F, Pratt RE, Berry GJ, et al. Increased accumulation of tissue ACE in human atherosclerotic coronary artery disease. Circulation. 1996;94:2756–2767.
Ohishi M, Ueda M, Rakugi H, et al. Upregulation of angiotensin-converting enzyme during the healing process after injury at the site of percutaneous transluminal coronary angioplasty in humans. Circulation. 1997;96:3328–3337.
Ribichini F, Steffenino G, Dellavalle A, et al. Plasma activity and insertion/deletion polymorphism of angiotensin I-converting enzyme: a major risk factor and a marker of risk for coronary stent restenosis. Circulation. 1998;97:147–154.
Lowry OH, Rosenbrough NJ, Farr AL, et al. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951;193:265–275.
Schieffer B, Schieffer E, Hilfiker-Kleiner D, et al. Expression of angiotensin II and interleukin 6 in human coronary atherosclerotic plaques: potential implications for inflammation and plaque instability. Circulation. 2000;101:1372–1378.
Griendling KK, Minieri CA, Ollerenshaw JD, et al. Angiotensin II stimulates NADH and NADPH oxidase activity in cultured vascular smooth muscle cells. Circ Res. 1994;74:1141–1148.
Rakugi H, Kim DK, Krieger JE, et al. Induction of angiotensin converting enzyme in the neointima after vascular injury: possible role in restenosis. J Clin Invest. 1994;93:339–346.
Ribichini F, Pugno F, Ferrero V, et al. Angiotensin-converting enzyme tissue activity in the diffuse in-stent restenotic plaque. Circulation. 2000;101:e33–e35.