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This study aimed to investigate whether the single nucleotide polymorphism C677T (rs1801133) of the methylene-tetrahydrofolate reductase (MTHFR) gene was associated with the risk of coronary artery disease (CAD) and circulating homocysteine (Hcy) levels in Tunisian population. 310 angiografically diagnosed CAD patients and 210 controls were enrolled in this study. The MTHFR C677T (rs1801133) polymorphism was genotyped, and the Hcy concentrations were measured. The severity of CAD was evaluated using the Gensini scoring system. Compared to the CC genotype, the TT genotype confers a higher risk for CAD severity with an OR = 9.07 and 95% CI = 3.78-21.8. The T allele was the predisposing allele for CAD and that it was probably associated with CAD severity. The area under the ROC curve for Hcy was 0.764 (95% CI 0.660 to 0.868, p = 0.001). The receiver operating characteristics curve (ROC) for Hcy showed its useful prediction of CAD. Hcy levels were not significantly associated with CAD severity expressed by Gensini Score (GS). The MTHFR C677T (rs1801133) polymorphism influences circulating Hcy levels. The MTHFR C677T polymorphism and hyperhomocysteinemia could have an important role in the prediction of the presence and not the severity expressed by GS of CAD.

Purpose: Clopidogrel non-responsiveness is multifactorial; several genetic and non-genetic factors may contribute to impaired platelet inhibition. The goal of this study is to determine the effect of the cytochrome P450 CYP2C19*2 polymorphism on the platelet response to clopidogrel in patients with and without diabetes mellitus (DM)., Methods: We conducted an observational study in patients with coronary artery disease and consequent exposure to clopidogrel therapy (75 mg/day for at least 7 consecutive days). We have analyzed two groups of patients: group I (DM patients) and group II (non-diabetes mellitus patients). Platelet reactivity was assessed by the VerifyNow P2Y12 assay and high on clopidogrel platelet reactivity (HPR) was defined as P2Y12 reaction units (PRU) ≥ 208. Genotyping for CYP2C19*2 polymorphism was performed by PCR-RFLP., Results: We have included 150 subjects (76 DM and 74 non-diabetes mellitus patients). The carriage of CYP2C19*2 allele, in DM patients, was significantly associated to HPR (odds ratio (OR) 4.437, 95% confidence interval (CI) 1.134 to 17.359; p = 0.032). Furthermore, 8.4% of the variability in percent inhibition by clopidogrel could be attributed to CYP2C19*2 carrier status. However, in non-diabetes mellitus patients, there was no significant difference in platelet response to clopidogrel according to the presence or absence of CYP2C19*2 allele carriage (OR 1.260, 95% CI 0.288 to 5.522; p = 0.759)., Conclusions: Our study suggests that the carriage of CYP2C19*2 polymorphism, in DM patients, might be a potential predictor of persisting HPR in these high-risk individuals., Trial Registration: Clinical Trials.gov NCT03373552 (Registered 13 December 2017).

In this study, we evaluate the relationships between aspirin nonresponsiveness and the cyclooxygenase-1 (Cox-1) gene C50T polymorphism in stable coronary artery disease (CAD) in Tunisian patients. One hundred twenty-five stable CAD patients were included. The Cox-1 gene C50T polymorphism was determined by the polymerase chain reaction/restriction fragment length polymorphism method. Aspirin response was evaluated by measuring the collagen epinephrine closer time and the urinary dehydro-thromboxane B2 excretion. According to the collagen epinephrine closer time values, the frequency of the -50T allele was not significantly different in bad responders when compared with good responders (36.8% vs. 15.7%; p=0.1). Similarly, the presence of the -50T mutant allele was not statistically different comparing bad and good responders according to the urinary 11-dehydro-thromboxane B2 excretion concentration (60% vs. 40%; p=0.43). Our study did not demonstrate any association between the Cox-1 gene C50T polymorphism and aspirin nonresponsiveness status in stable CAD patients.

Objectives: In this study, we investigated the association between matrix metalloproteinase-1 (MMP-1) G-1607GG, MMP-12 A-82G and MMP-12 A1082G genotypes and haplotypes and the prognosis of coronary artery disease (CAD)., Methods: A total of 129 Tunisian patients with CAD were followed prospectively for a median of 2.5 years. Genotypes were determined by a PCR-based restriction fragment length polymorphism. Two endpoints were considered: restenosis and incidence of clinical vascular events (restenosis, myocardial infarction, stroke, cardiac death)., Results: Genotypes of MMP-1 G-1607GG, MMP-12 A-82G and MMP-12 A1082G were not associated with the incidence of restenosis or clinical events. Analysis of haplotypes consisting of alleles of MMP-1 G-1607GG and MMP-12 A1082G showed that the rate of clinical events was significantly higher in patients carrying the GG-A haplotype than those with other haplotypes (0.637 vs. 0.424, respectively, odds ratio=1.45; 95% confidence interval=1.04-2.04; P<0.05; P adjusted for multiple risk factors). However, after Bonferroni correction for multiple comparisons, this difference did not reach statistical significance (P=0.093), showing that there was a tendency for the association between the GG-A haplotype and future clinical events in patients with CAD., Conclusion: These findings showed a trend of the GG-A haplotype of MMP-1 G-1607GG/MMP-12 A1082G towards the prediction of future clinical events in patients with CAD and suggested a possible importance of these loci in the prediction of the prognosis of CAD. Studies with large sample size are warranted to better investigate this association, as MMP genotyping could aid in identifying patients who are likely to have unfavourable prognosis.

Platelet glycoprotein IIb/IIIa is a membrane receptor which plays a key role in coronary artery disease and thrombotic events. However, there is a considerable controversy regarding the clinical impact of glycoprotein IIIa platelet antigen 1 (PlA1)/platelet antigen 2 (PlA2) polymorphism as a risk factor for myocardial infarction. To evaluate the association between glycoprotein IIIa PlA1/PlA2 polymorphism and 1-year cardiovascular events occurrence in aspirin-treated patients with stable coronary artery disease. We prospectively included 188 postacute coronary syndrome patients (183 men) aged 59 ± 10 years and receiving aspirin (250 mg/day). The clinical outcome at 1 year was the composite end point of nonfatal myocardial infarction, stroke, recurrent unstable angina or cardiac death. Genotyping for PlA1/PlA2 polymorphism was conducted using PCR and restriction fragment length polymorphism analysis. The genotype distribution of glycoprotein IIIa PlA1/PlA2 polymorphism was PlA1/PlA1, 55.3%; PlA1/PlA2, 39.3% and PlA2/PlA2, 4%. Incidence of composite end point in homozygous PlA1/PlA1 carriers was significantly higher than in PlA2/PlA2 and PlA1/PlA2 patients [14.4 vs. 3.6% odds ratio 4.5 (1.2-16.6, 95% confidence interval); P = 0.012]. Multivariate analysis identified three strong predictive factors of cardiac death: age more than 65 years [odds ratio = 6.8, (1.4-34, 95% confidence interval); P = 0.018], ventricular ejection fraction less than 50% [odds ratio = 8.6, (1.7-42.6, 95% confidence interval); P = 0.008] and homozygous PlA1/PlA1 genotype [odds ratio = 8.8, (1.0-78.6, 95% confidence interval); P = 0.014]. Our results demonstrated that glycoprotein IIIa PlA1/PlA1 genotype carriers have a significantly increased risks of acute vascular ischemic events associated with a poor prognosis at 1 year. These postacute coronary syndrome patients might require an optimized secondary antithrombotic prophylaxis strategy.

The aim of this pilot study was to compare the effect of two different regimens of aspirin dosage on platelet of coronary artery disease (CAD) diabetic patients. Twenty-five CAD diabetic patients were included. Initially, all patients received aspirin 100 mg/day for 10 days. At day 10, aspirin antiplatelet effect was determined by measuring the collagen/epinephrine closure time (CT) 2 h after the last aspirin dosage and the next morning at 8 a.m.. The aspirin regimen was modified to 100 mg twice daily for patients showing a non-optimal platelet-inhibitory effect (CT < 298 s at 8 a.m.). Persistent high platelet reactivity (HPR) was defined by a CT < 160 s. During the 100 mg/day aspirin regimen, the prevalence of HPR at 8 a.m. was 48%, and only 7 patients (28%) had showed an optimal platelet-inhibitory effect. Bridging to the twice-daily regimen, the HPR was significantly reduced (p=0.025), and the optimal platelet-inhibitory effect was reached for 3 other patients. Our results showed that 100 mg aspirin twice-daily dosing rather than a once-daily dose significantly improves the aspirin effect on platelet of CAD diabetic patients. However, large prospective studies were needed to confirm whether this strategy will be clinically relevant and safe.

Peroxisome proliferator-activated receptor delta (PPAR-delta) is a transcription factor implicated in metabolism and inflammation. The +294T/C polymorphism in the PPAR-delta gene is associated with risk of coronary artery disease (CAD) in dyslipidemic women and hypercholesterolemic men. Whether this polymorphism influences the risk of CAD in the absence of dyslipidemia was not known, so we investigated a possible association of this polymorphism with plasma lipid and lipoprotein levels and with risk and outcome of CAD in a normolipidemic Tunisian population. Genotyping was performed by PCR-RFLP in 112 CAD patients and 113 healthy volunteers. The C-allele was significantly more frequent in patients than in controls (0.320 vs 0.189, P = 0.001). This association remained significant after adjustment for age, gender, body mass index, smoking, hypertension, and high-density lipoprotein cholesterol. Subjects carrying either one or two copies of the C-allele had a 2.7-fold higher risk of CAD than subjects homozygous for the T-allele. PPAR-delta genotypes were not associated with lipoprotein concentrations or outcome of CAD. We conclude that PPAR-delta +294T/C polymorphism is an independent risk factor of CAD in normolipidemic Tunisian subjects. The lack of association with lipoprotein concentrations suggests that the effect of the polymorphism on CAD is not mediated through lipoprotein levels in this population and that it may influence the atherosclerotic process through mechanisms involving inflammation.

Background: Various genetic polymorphisms have been proposed to explain the persistent platelet hyperactivity (HPR) under aspirin treatment. PlA polymorphism of platelet GPIIIa receptor has been largely studied. However, its influence on platelet sensitivity to aspirin remains controversial., Objectives: The aim of this prospective study is to investigate whether this PlA polymorphism is associated with a greater prevalence of HPR in stable coronary artery disease patients Material and Methods: 188 stable coronary artery disease patients were included. Platelet aspirin inhibitory effect was determined with PFA-100 using Collagen/Epinephrine closure time (CEPI-CT). A CEPI-CT<160sec was defining the HPR status. GPIIIa PlA polymorphism was established using polymerase chain reaction and classical restriction fragments length polymorphism., Results: The observed frequencies of different genotypes were not different from those predicted by the Hardy-Weinberg equilibrium: PlA1/lA1 (55.3%), PlA1/PlA2 (39.4%) and PlA2/PlA2 (5.3%). HPR patients with inadequate aspirin inhibition were significantly more often homozygous PlA1/A1 (65.4% vs. 47.7%, p=0.015). After multivariate analysis, PlA1/A1 genotype was the only independent risk factor for persistent HPR (OR: 2.07; 95% CI [1.14 to 3.76; p=0.016)., Conclusion: In CAD patients receiving daily low dose of aspirin, there is a significant and independent association between the expression of GPIIIa PlA1 allele and the occurrence of persistent HPR detected with PFA-100., (Copyright 2009 Elsevier Ltd. All rights reserved.)

The aim of this study was to assess the association between "aspirin non responsiveness" in patients with coronary artery diseases (CAD) and the risk of major adverse cardiovascular events (MACE). 204 patients with CAD receiving aspirin (250 mg/d) were included. Both Collagen/Epinephrine Closure Time (CEPI-CT) and urinary Thromboxane B2 (uTxB2) concentration was used to determine the patients aspirin responsiveness. The clinical primary endpoint was the occurrence of MACE including: cardiovascular death, MI, stroke or transient ischemic attack. The secondary endpoint was the occurrence of Recurrent Acute Vascular Event (RAVE: MI, stroke or transient ischemic attack). After 1-year follow-up, no responders diagnosed by CEPI-CT had a trend for higher risk of MACE (13% vs 7.4%; P = 0.22) and significant higher risk of RAVE (OR = 2.1; 95%CI: 1.7-2.4; P = 0.01) when compared to good responders. Multivariate analysis showed that CEPI-CT < 143 s was the only independent predictor of RAVE (OR = 6.3; 95% CI: 1.2-32.2; P = 0.026). Aspirin non-responsiveness, diagnosed by the uTxB2, was not associated with an increased risk of either MACE or RAVE. Our results, reinforce the importance of being able to diagnose laboratory "aspirin non responsiveness", and extend the evidence that aspirin non responsiveness may explain in part the occurrence of RAVE.

Aim: To analyze apolipoprotein gene polymorphisms in the Tunisian population and to check the relation of these polymorphisms and homocysteine, lipid and apolipoprotein levels to the coronary artery disease (CAD)., Methods: In healthy blood donors and in patients with CAD complicated by myocardial infarction (MI) four apolipoprotein gene polymorphisms [APO (a) PNR, APO E, APO CI and APO CII] were determined and plasma levels of total homocysteine, total cholesterol (TC), triglycerides (TG), HDL-cholesterol (HLD-C) and apolipoproteins (apo A-I, Apo B, Apo E) were measured., Results: Analysis of the four apolipoprotein gene polymorphisms shows a relative genetic homogeneity between Tunisian population and those on the other side of Mediterranean basin. Compared to controls, CAD patients have significantly higher main concentrations of TC, TG, LDL-C, apo B and homocysteine, and significantly lower ones of HDL-C, apo A-I and apo E. The four apolipoprotein gene polymorphisms have not showed any significant differences between patients and controls. However, the APO E4 allele appears to be associated to the severity of CAD and to high levels of atherogenic parameters and low level of apo E, which has very likely an anti-atherogenic role., Conclusion: Although APO (a) PNR, APO CI and APO CII genes are analyzed in only few populations, they show a frequency distribution, which is not at variance with that of APO E gene and other widely studied genetic markers. In the Tunisian population the APO E 4 appears to be only indirectly involved in the severity of CAD. In the routine practice, in addition of classic parameters, it will be useful to measure the concentration of apo E and that of Homocysteine and if possible to determine the APO E gene polymorphism.

Several matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) have been implicated in the development and outcome of coronary artery disease (CAD). We investigated whether MMP-12 and TIMP-1 levels were associated with risk, severity, and outcome of CAD. Plasma MMP-12 and TIMP-1 levels are measured in 50 and 44 patients with CAD, respectively, by enzyme-linked immunosorbent assay. Of all patients, 16 were taking statins. Patients who were not on statins were classified into 3 groups according to number of >50% stenotic vessels. Compared with 29 volunteers without CAD, patients without statins (n = 34) had higher MMP-12 concentrations (1.71 vs 1.08 ng/ml, p = 0.021). MMP-12 levels were significantly lower in patients with than in those without statin treatment (0.99 vs 1.71 ng/ml, p = 0.008). There was no association between MMP-12 levels and number of >50% stenotic vessels. MMP-12 concentrations were not associated with outcome of CAD. However, plasma TIMP-1 levels were associated with restenosis independently of number of stenotic vessels and age (p = 0.035) but not with risk or severity of CAD. In conclusion, plasma MMP-12 concentration was associated with the presence of CAD. Statin therapy decreases plasma MMP-12 levels in patients with CAD. Increased TIMP-1 levels may prevent restenosis after angioplasty.

Background: Aspirin (ASA) failure to inhibit in vitro platelet function had been termed ASA resistance. The prevalence of this phenomenon as measured with different platelet function tests varies widely among studies., Objectives: In this study, we propose to determine the prevalence of ASA non-responsiveness in stable coronary artery patients using three different tests., Patients and Methods: One hundred ninety-one patients with a stable coronary artery disease and receiving secondary ASA prophylaxis (250 mg/day) were tested. For each patient the ASA-induced platelet inhibition was determined using three different tests: Ivy Bleeding time (BT), collagen/epinephrine closure time (CEPI-CT; PFA-100, Dade-Behring) and urinary 11-dehydrothromboxane B2 (uTxB2) excretion level. The agreement between these tests was evaluated by kappa statistics test., Results: The prevalence of biological ASA resistance was 15.7% (n=30), 20.4% (n=39) and 24.6% (n=47) by BT, PFA-100 and UTxB2, respectively. Only fourteen patients (7.3%) were non-responders for two tests: 6 (3.1%) BT/ PFA-100; 1 (0.5%) BT/UTxB2; 7 (3.7%) PFA-100/UTxB2). A poor agreement was found between these three methods and only 3 patients were resistant with all the tests (1.6%)., Conclusion: The lack of agreement supposed that different types of aspirin resistance exist. Thus, combination of two tests or more could be a primary solution for a better identification of ASA resistant patients. This hypothesis must be confirmed by a large-scale randomized study with clinically well-defined endpoints.

Abstract: Objective: Coronary artery disease (CAD) is a complex multifactorial disease due to the interaction of multiple genes variations and environmental factors. Genetic variants of lipoprotein lipase (LPL), a key enzyme in the hydrolysis of triglyceride rich particles, may contribute to CAD. We analysed here the frequency of LPL variants (p.Asp9Asn, p.Asn291Ser and p.Ser447X) in a Tunisian population as well as their association with circulating lipid level and risk of CAD. Patients and methods: LPL variations were investigated by PCR-RFLP and lipid parameters were measured in 135 patients and 109 controls. Results: The frequency of the p.Asp9Asn variation was 10.37% in CAD patients versus 3.66% in controls. The frequency for the p.Ser447X variation was 8.8% in CAD patients versus 13.7% in controls. There was no significant association between these two variants and CAD. The p.Asn291Ser mutation variation was absent in this population. In healthy subjects, heterozygote carriers of the p.Asp9Asn substitution had a significant increase level of total cholesterol (4.2±0.9mmol/L vs 5.6±1.2mmol/L; P =0.01) and a decreased level of HDL-cholesterol (1.36±0.3mmol/L vs 0.93±0.1mmol/L; P =0.045). Conclusion: There was no significant association between genetic variants of the LPL gene and CAD in this Tunisian population. The very low frequency of the p.Asn291Ser variation may be an ethnic specificity of Tunisians. [Copyright &y& Elsevier]