2.1. Compliance with Ethical Standards
From October 2016 to October 2019, we conducted a prospective, single-center, randomized, double-blind study in the Chinese PLA General Hospital in Beijing.
The study complied with the Declaration of Helsinki, which was approved by the Beijing Ethics Association and the Ethics Committee of the Chinese PLA General Hospital.
Informed consent was obtained from all individual participants included in the study.
The trial was registered on ClinicalTrials.gov (registration number: NCT02826616).There is no potential conflicts of interest.
2.2. Study population
The study population included patients diagnosed with STEMI for the first time and underwent primary PCI within 12 hours of the onset of chest pain. Diagnosis of STEMI was based on the concurrence of symptoms consistent with STEMI for > 30 minutes; and ST-segment elevation ≥ 1 mm in at least 2 or more inferior leads or ≥ 2 mm in at least 2 or more contiguous precordial leads. Patients were excluded if they met any one of the bellow conditions: already treated with TMZ, history of myocardial infarction, mechanical complications, previous coronary artery bypass grafting (CABG) or PCI, contraindications of CMR, liver or kidney failure, malignant tumor.
2.3. PCI Procedures
Eligible patients were randomly assigned to receive TMZ or placebo before primary PCI. Primary PCI was performed by 2 operators using standard techniques. Patients took 300 mg aspirin in the emergency department, followed by 100 mg aspirin per day orally thereafter. A loading dose of 180 mg ticagrelor was administered before catheterization, followed by 90 mg ticagrelor twice daily for the next 12 months. Drug-eluting stents were the stents of choice. All patients were given enoxaparin treatment every 12 hours after primary PCI for 7 days. Blood samples were taken for troponin T testing before and at 6, 12, 24, 48 hours after PCI. The application of thrombectomy and intro-aortic balloon pump (IABP) was at the discretion of the operators. The PCI data analysts were kept blind from grouping.
2.4. Experimental Treatment Protocol
All patients were informed of the potential benefits and risks of the trial before they signed written informed consents and enrolled in the study.
Patients were randomly divided into placebo and TMZ groups by using a computer-generated sequence at a ratio of 1:1. Patients were treated with a dose of 60 mg TMZ (Servier companies) or placebo orally prior to reperfusion by primary PCI and followed by 20 mg TMZ or placebo three times a day for 12 months. Other medications including aspirin, ticagrelor, statins, β-blockers and ACEI/ARB were given in accordance with the European Cardiology Society STEMI guideline [
11].
2.5. Primary endpoint and clinical Follow-Up
The primary endpoint was the infarction size measured by CMR at 7 days after primary PCI. The secondary endpoint was MSI measured by CMR at 7 days after primary PCI and main adverse cardiac events (MACEs) in following 12 months. We followed up these patients by routine clinical visits and recorded any MACEs until 12 months after the PCI.
Follow-up information was obtained at the outpatient clinic.
Good clinical practice training was required for all personnel involved in the trial. MACEs were defined as stroke, repeat revascularization, and readmission due to acute heart failure, nonfatal myocardial infarction and all-cause death. All end-point events were adjudicated by an independent clinical events committee based on medical record. All were blinded to treatment group.
2.6. Cardiovascular magnetic resonance acquisition and analysis
The scan was performed at 7 days after primary PCI so as to assess the final infarction size, microvascular obstruction (MVO), area at risk (AAR) and MSI.
All participants took a CMR examination on a 1.5T MR Scanner (Achieva; Philips Medical Systems, Best, the Netherlands), using the 32-channel phased-array body coil.
All CMR images were acquired during breath-hold and with ECG triggering. Retrospective ECG gated cine CMR imaging was performed with steady-state free precession (SSFP) sequences using the standard protocol covering short axis and long axis in the 2- and 4- chamber views. The SSFP cine images were acquired continuously from the mitral annulus to the apical level without gaps on the short-axis. Based on the retrospective triggering, 25–30 cardiac phases covering systole and diastole within a cardiac cycle were reconstructed. Infarction size was acquired by the CMR LGE method which acquired the images after 15 min injection of gadolinium (0.1 mmol/kg at 3ml/s). Inversion-recovery CMR LGE images were obtained at end-diastolic on short axis and the inversion time was manually adjusted to null the signal from remote myocardium. Myocardial edema-sensitive black-blood T2-weighted short tau inversion-recovery (STIR) sequence was performed using a fat-saturation triple inversion-recovery sequence. A full stack of LGE and T2 were acquired with about 10 slices of left ventricle (LV) chamber from base to apex, 8 mm apart. Each slice of LGE and T2 owed the same parametric location which was available for the precise analysis afterward.
The CMR data were analyzed by 2 CMR readers using the freely available validated cardiovascular image analysis software CVI42 5.11.2 (Circle Cardiovascular Imaging Inc, Calgary, Canada, version 5.10.1). Cine images on short-axis were used to analyze LV systolic function. The LV function analysis included all slices from end-diastole to end-systole. Left ventricular ejection fraction (LVEF) was calculated by manually tracing the endocardial borders in all phases on an ECG-triggered balanced steady-state-free procession cine sequence by applying multiple slices in the short-axis image covering the entire LV. Specifically, after manual tracing of the epicardial and endocardial borders, the areas of STEMI were quantified in LGE images detected by semi-automated software, by the method of mean + 5SD Ref ROI with manual correction (Fig. 1A-B). We removed the artifact in the remote myocardium which would affect the infarction size calculation in the LGE image. Infarction size was expressed both in grams and in the percentage of the total LV mass. The myocardial AAR was assessed as edema on the CMR scan using T2-weighted STIR sequence (Fig. 1C-D). AAR was defined as the long signal area on T2-weighted images, and was semi-automated recognized by mean + 2SD Ref ROI. We removed the artifact in the remote non-infarcted myocardium in the T2 STIR image with the manual correction to analyze AAR, and the area of MVO should be manually included in AAR. The MSI was calculated as follows: (AAR, g - infarction size, g)/ AAR, g. MVO was identified in LGE images as a subendocardial region with lower enhancement than the surrounding area which was semi-automated discerned via the dark areas inside the infarcted myocardium tissue with manual correction. All the data were validated by inter-and intra-observer analysis of reproducibility method.
2.7. Statistical analysis
Continuous variables were presented as mean ± SD and compared by the t-test for independent samples. Non-normally distribution variables were presented as a median and interquartile range, and compared by the Wilcoxon rank-sum test. Categorical data were presented as percentages and compared by the χ 2 test or the Fisher exact test when there were less than 5 values in a given cell. All statistical analyses were performed using SPSS statistical software (version 18.0, SPSS, Chicago, Illinois).