The objective of this study is to investigate the potential association of the
Coronary artery disease (CAD), characterized pathologically by atherosclerosis, is the most important cause of death in most countries and acts as a major economic burden on their public health systems [
As a multiprotein complex, NLRP3 inflammasome includes the NLRP3 polypeptide, caspase recruitment domain-containing protein 8 (CARD8), the PYD and CARD domain-containing protein (PYCARD), and caspase-1. NLRP3 inflammasome forms and is activated when pattern-recognition receptors detect endogenous and exogenous “danger signals” such as cholesterol crystals [
NLRP3 polypeptide and CARD8 control the activity of inflammatory caspase-1 through the formation of NLRP3 inflammasome complexes [
This study was approved by the Ethics Committee of the First Affiliated Hospital of Xi’an Jiaotong University (Xi’an, Shaanxi, China) in accordance with the Declaration of Helsinki guidelines. All participants were fully aware of the research and signed informed consents.
In the case-control study, a total number of 916 consecutive Chinese Han subjects (aged from 24 to 80 years) were admitted to the Department of Cardiology at the First Affiliated Hospital of Xi’an Jiaotong University from January 2012 to April 2014 because of chest pain. Based on medical history, typical electrocardiogram changes, myocardial enzymes, and coronary angiography (CAG), the subjects were classified into the CAD group (
The prospective cohort study consisted of the 515 CAD patients. All the patients underwent a PCI procedure because of serious coronary artery stenosis and were followed up for a median period of 32 months (range, 14–40 months). Major adverse cardiovascular events (MACEs) are defined as nonfatal MI, cardiovascular death, unstable angina, nonfatal ischemic stroke, and revascularization procedure. According to whether MACEs occurred during the follow-up, the patients were divided into the MACE group and the non-MACE group.
Patients with any of the following were excluded from the study: acute infections, postrevascularization, serious liver or kidney disease, cancer, or acute stroke.
Clinical data of the subjects was collected by well-trained investigators. A history of smoking was defined as either past or current use of cigarettes. Hypertension was defined as systolic blood pressure ≥140 mmHg and/or diastolic blood pressure ≥90 mmHg. Diabetes mellitus (DM) was defined as an elevated fasting plasma glucose concentration >126 mg/dL, or serum glycosylated hemoglobin A1C (HbA1C) level ≥6.5%. The weight (kg) and height (m) of every subject were acquired during the initial visit. Body mass index (BMI) was calculated by the equation: weight (kg)/height (m2).
Coronary angiographies and PCI procedure were performed according to the Judkins technique. CAD was determined by coronary angiographies with an epicardial coronary artery narrowing of ≥50%. The severity of stenosis in the coronary artery was reflected by Gensini scores [
All blood samples (5 mL) were collected into tubes containing anticoagulant and then were centrifuged for 10 minutes at 3000 rpm. Levels of serum high sensitivity C-reactive protein (hs-CRP), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), and triglyceride (TG) were measured at the laboratory of our hospital. Low-density lipoprotein cholesterol (LDL-C) was calculated using the Friedewald formula. Interleukin-1
In the process of SNP selection of NLRP3 inflammasome, we included in our study those SNPs with a MAF (Minor Allele Frequency) >0.05 in the Chinese Han population. MAFs were analyzed using online tools (
The 515 CAD patients were followed up at months 1, 6, 12, 24, 36, and 40 (median follow-up time 32 months) after discharge and 12 subjects (2.33%) were lost to follow-up. Information of the patients was obtained through telephone communications or face-to-face interviews with the patients or their family members by a trained research cardiologist using a structured questionnaire. MACE incidence of subjects of different genotypes was calculated.
Categorical variable was presented as
The clinical characteristics of the 401 control subjects and 515 CAD patients are summarized in Table
Baseline characteristics of the control and CAD subjects.
Control | CAD |
|
|
---|---|---|---|
Total, |
401 | 515 | |
Male, |
241 (60.10%) | 364 (70.68%) | 0.001 |
Age, y (mean |
57.49 |
59.50 |
0.031 |
Hypertension (%) | 194 (48.38%) | 286 (55.53%) | 0.033 |
DM (%) | 69 (17.21%) | 132 (25.63%) | 0.002 |
Smoking (%) | 151 (37.66%) | 248 (48.16%) | 0.002 |
SBp (mmHg) | 127.16 |
125.53 |
0.242 |
DBp (mmHg) | 79.26 |
78.48 |
0.389 |
HR (beats/min) | 72.75 |
72.86 |
0.912 |
BMI (kg/m2) | 24.18 |
24.75 |
0.003 |
LVEF (%) | 63.06 |
57.65 |
0.156 |
Hs-CRP (mg/L) | 2.71 |
6.38 |
|
TC (mmol/L) | 3.83 |
3.95 |
0.051 |
TG (mmol/L) | 1.65 |
1.70 |
0.334 |
HDL-C (mmol/L) | 1.00 |
0.96 |
0.016 |
LDL-C (mmol/L) | 2.16 |
2.31 |
0.003 |
CAD: coronary artery disease; SBp: systolic blood pressure; DBp: diastolic blood pressure; HR: heart rate; BMI: body mass index; LVEF: left ventricular ejection fraction; Hs-CRP: high-sensitivity C-reactive protein; TC: total cholesterol; TG: triglyceride; HDL-C: high-density lipoprotein cholesterol; LDL-C: low-density lipoprotein cholesterol; DM: diabetes mellitus.
The distributions of the genotypes are shown in Table
Genotype frequencies of
Control | CAD | AOR (95% CI) |
|
|
---|---|---|---|---|
( |
( |
|||
|
||||
CC | 142 (35.41%) | 149 (28.93%) | 1.000 (reference) | |
CG | 200 (49.88%) | 263 (51.07%) | 1.294 (0.952–1.758) | 0.099 |
GG | 59 (14.71%) | 103 (20.00) | 1.630 (1.080–2.459) | 0.020 |
Dominant model (CC versus CG + GG) | 1.371 (1.024–1.835) | 0.034 | ||
Recessive model (CC + CG versus GG) | 1.392 (0.965–2.007) | 0.077 | ||
C allele | 484 (60.35%) | 561 (54.47%) | 1.000 (reference) | |
G allele | 318 (39.65%) | 469 (45.53%) | 1.263 (1.041–1.534) | 0.018 |
HWE |
0.713 | 0.450 | ||
HWE |
0.399 | 0.502 | ||
|
||||
|
||||
AA | 175 (43.64%) | 210 (40.78%) | 1.000 (reference) | |
AT | 186 (46.38%) | 244 (47.38%) | 1.160 (0.870–1.545) | 0.311 |
TT | 40 (9.98%) | 61 (11.84%) | 1.252 (0.787–1.990) | 0.343 |
Dominant model (AA versus AT + TT) | 1.176 (0.894–1.547) | 0.245 | ||
Recessive model (AA + AT versus TT) | 1.158 (0.747–1.795) | 0.513 | ||
A allele | 536 (66.83%) | 664 (64.47%) | 1.000 (reference) | |
T allele | 266 (33.17%) | 366 (35.53%) | 1.126 (0.921–1.377) | 0.248 |
HWE |
0.858 | 0.600 | ||
HWE |
0.354 | 0.439 |
CAD: coronary artery disease; AOR: adjusted odds ratio; HWE: Hardy-Weinberg equilibrium; 95% CI: 95% confidence interval.
Since the MAF of
Gensini scoring system is a well-used method for evaluating the severity of coronary atherosclerosis based on angiographic findings. Gensini scores for
The Gensini scores for
Among 503 patients who accepted follow-up during a median follow-up period of 32 months, 145 (28.83%) had MACEs including 33 deaths, 19 acute myocardial infarctions, 23 coronary revascularizations, 51 unstable anginas, and 19 strokes. The clinical characteristics of patients with or without MACE are summarized in Table
Baseline characteristics of CAD patients with and without MACE during follow-up.
Patients without MACE | Patients with MACE |
| |
---|---|---|---|
Total, |
358 | 145 | |
Male, |
240 (67.04%) | 118 (81.38%) | 0.001 |
Age, y (mean |
58.52 |
61.75 |
0.004 |
Hypertension (%) | 188 (52.51%) | 93 (64.14%) | 0.018 |
DM (%) | 77 (21.51%) | 52 (35.86%) | 0.001 |
Smoking (%) | 159 (44.41%) | 84 (57.93%) | 0.008 |
SBp (mmHg) | 125.79 |
125.34 |
0.843 |
DBp (mmHg) | 78.73 |
78.39 |
0.802 |
HR (beats/min) | 73.44 |
71.37 |
0.212 |
BMI (kg/m2) | 24.63 |
25.20 |
0.045 |
LVEF (%) | 57.58 |
55.54 |
0.085 |
Hs-CRP (mg/L) | 5.76 |
6.34 |
0.029 |
TC (mmol/L) | 3.91 |
4.09 |
0.060 |
TG (mmol/L) | 1.69 |
1.73 |
0.590 |
HDL-C (mmol/L) | 0.99 |
0.88 |
|
LDL-C (mmol/L) | 2.26 |
2.51 |
0.001 |
ACE-I/ARB, |
247 (68.99%) | 96 (66.21%) | 0.527 |
Statins, |
343 (95.81%) | 134 (92.41%) | 0.124 |
|
246 (68.72%) | 105 (72.41%) | 0.454 |
Aspirin, |
344 (96.09%) | 137 (94.48%) | 0.471 |
CAD: coronary artery disease; MACE: major adverse cardiacevent; SBp: systolic blood pressure; DBp: diastolic blood pressure; HR: heart rate; BMI: body mass index; LVEF: left ventricular ejection fraction; Hs-CRP: high-sensitivity C-reactive protein; TC: total cholesterol; TG: triglyceride; HDL-C: high-density lipoprotein cholesterol; LDL-C: low-density lipoprotein cholesterol; DM: diabetes mellitus; ACEI: angiotensin converting enzyme inhibitor; ARB: angiotensin receptor antagonist.
Genotype frequencies of
Patients without MACE | Patients with MACE | Adjusted HR (95% CI) |
Log-rank |
|
---|---|---|---|---|
( |
( |
|||
|
||||
CC | 113 (31.56%) | 32 (22.07%) | 1.000 (reference) | |
CG | 180 (50.28%) | 75 (51.72%) | 1.503 (0.990–2.282) | 0.056 |
GG | 65 (18.16%) | 38 (26.21%) | 1.790 (1.107–2.893) | 0.017 |
Dominant model (CC versus CG + GG) | 1.585 (1.065–2.359) | 0.023 | ||
Recessive model (CC + CG versus GG) | 1.363 (0.934–1.989) | 0.108 | ||
C allele | 406 (56.70%) | 139 (47.93%) | 1.000 (reference) | |
G allele | 310 (43.30%) | 151 (52.07%) | 1.502 (1.182–1.909) | 0.001 |
HWE |
0.206 | 0.191 | ||
HWE |
0.650 | 0.662 | ||
|
||||
|
||||
AA | 146 (40.78%) | 60 (41.38%) | 1.000 (reference) | |
AT | 168 (46.93%) | 68 (46.90%) | 1.193 (0.831–1.711) | 0.338 |
TT | 44 (12.29%) | 17 (11.72%) | 0.850 (0.483–1.497) | 0.574 |
Dominant model (AA versus AT + TT) | 1.111 (0.786–1.571) | 0.551 | ||
Recessive model (AA + AT versus TT) | 1.044 (0.981–1.111) | 0.174 | ||
A allele | 460 (64.25%) | 188 (64.83%) | 1.000 (reference) | |
T allele | 256 (35.75%) | 102 (35.17%) | 0.992 (0.774–1.272) | 0.949 |
HWE |
0.165 | 0.117 | ||
HWE |
0.685 | 0.733 |
CAD: coronary artery disease; HR: hazard risk; MACE: major adverse cardiovascular event. HWE: Hardy-Weinberg equilibrium; 95% CI: 95% confidence interval.
Kaplan-Meier survival curves of the freedom of MACEs and death in CAD patients with different
In order to identify the pathogenic mechanisms of the
The comparison of the serum levels of IL-1
In this study, our current results demonstrate a clear correlation between the
Atherosclerosis is the most important pathophysiological characteristic of CAD. There is a consensus that inflammatory response plays an important role in the progression of atherosclerosis. Many kinds of inflammatory cytokines, such as IL-1
Research has shown that the gene polymorphism of NLRP3 rs10754558 is associated with common inflammatory diseases or a known functional effect [
CARD8 has been proved to be associated with several inflammatory diseases [
Even though we carried out a comprehensive analysis, some limitations of this study need to be noted in evaluating our results. The sample size was relatively small. Further studies with larger sample sizes, longer follow-up periods, and more detailed examination of the mechanism of G allele in atherosclerosis should be conducted to assess the effect of
In conclusion, the present study shows that the G allele of
The authors declare that they have no competing interests.
Dong Zhou and Xinhong Wang contributed equally to this study. Dong Zhou and Xinhong Wang contributed with clinical data collection, Tao Chen and Wen Wen performed the laboratory experiments, Yue Wu and Zuyi Yuan contributed with study design, Dong Zhou and Yang Liu did statistical analysis, and Dong Zhou and Xinhong Wang wrote the paper.
This study was supported by the National Basic Research Program of China (“973 Project” no. 2012CB517804) and the National Natural Science Fund for Distinguished Young Scholars (81025002).