Statin use is associated with a significant reduction in cholesterol content of erythrocyte membranes. A novel pleiotropic effect?

PURPOSE: High cholesterol content of erythrocyte membranes (CEM) levels is present in patients with acute coronary syndromes (ACS). Intraplaque hemorrhage and erythrocyte lysis contribute to the deposition of cholesterol on the atherosclerotic plaque and to plaque rupture. With the present study we assessed the effect of statin therapy on CEM levels, a novel marker of coronary artery disease (CAD) instability during a 1-year follow-up in CAD patients. METHODS: 212 consecutive eligible (158 men, 62 +/- 10 years) patients undergoing diagnostic coronary angiography for the assessment of angina pectoris were assessed. The study population comprised of 84 chronic stable angina (CSA) patients and 128 ACS patients. All study participants were commenced on statin treatment in equipotent doses and were followed for up to 1 year (at - 1, - 3, - 6 and - 12 months). RESULTS: Repeated measurements analysis of variance after appropriate adjustment showed a significant decrease (p < 0.001) in CEM content during follow up. CEM levels were decreasing at each time point (1 month : 100 microg/mg 95%CI 94.3-105.6, 3 months : 78.1 microg/mg 95%CI 73.2-83, 6 months : 67.2 microg/mg 95%CI 63.1-71.2, 1 year : 45.3 microg/mg 95%CI 42.2-48.3) compared to admission (112.1 microg/mg 95% CI 105.9-118.3) and to all previous measurements. CONCLUSIONS: The present study showed, that use of statins is associated with a reduction in CEM, an emerging marker of clinical instability and plaque vulnerability in CAD patients. The pleiotropic effects of statins at the cell membrane level represent a promising novel direction for research in CAD.

Cholesterol composition of erythrocyte membranes and its association with clinical presentation of coronary artery disease.

OBJECTIVES: Presence of free cholesterol in atherosclerotic plaques is a major determinant of plaque instability. It is hypothesized that extravasated erythrocytes may contribute to free cholesterol accumulation in atherosclerotic plaques through their rich in cholesterol membrane. In this study we assessed whether cholesterol in erythrocyte membranes (CEMs), that is, free (FCEM) versus esterified (ECEM), differs in patients with chronic stable angina (CSA) compared with patients presenting with acute coronary syndromes (ACSs). METHODS: Consecutive angina patients were prospectively assessed; 154 had CSA (118 men, 63 years, 56-69 years) and 164 ACS (124 men, 63 years, 55-71 years). FCEM and ECEM were measured using an enzymatic assay, and protein content was assessed by the Bradford method. RESULTS: FCEM was significantly higher (P<0.001) in the ACS patients group (94.1 microg/mg, IQ 71-116.5 microg/mg) compared with patients with CSA (61.9 microg/mg, IQ 49.3-73.1 microg/mg). ECEM levels were also significantly higher (P<0.001) in ACS patients (23.3 microg/mg, IQ 14.9-47.7 microg/mg) compared with CSA patients (10.8 microg/mg, IQ 8-22.3 microg/mg). In contrast, ratio of free-to-esterified cholesterol (P=0.110) as well as ratio of free-to-total CEM (P=0.109) were not different among CSA and ACS patients. CONCLUSION: Findings of this study show that although free cholesterol is the prevailing form of CEMs, both FCEM and ECEM levels are increased in patients with ACS compared with CSA patients. These findings suggest that it is the quantity of CEM rather than the type of cholesterol present in the erythrocyte membrane that determines plaque progression.

Interleukin-8 is increased in the membrane of circulating erythrocytes in patients with acute coronary syndrome.

AIMS: Studies have shown that erythrocyte membranes are present within necrotic cores in atherosclerotic plaques, and that circulating erythrocytes in patients with acute coronary syndrome (ACS) have increased total cholesterol content (CEM). Interleukin-8 (IL-8) binds to erythrocytes and during intraplaque haemorrhage it is released into the plaque and thus may contribute to inflammatory cascade and atherosclerotic plaque instability. The present study was undertaken to test the hypothesis that erythrocyte membrane IL-8 is elevated in patients with ACS compared with those with chronic stable angina (CSA). METHODS AND RESULTS: Consecutive patients who presented with CSA (n = 120, 92 men, 62 +/- 9 years), ACS (n = 118, 90 men, 62 +/- 10 years) or with chest pain who had normal coronary arteries (n = 36, 26 men, 60 +/- 7 years), were studied prospectively. IL-8 concentrations in erythrocyte membranes (rIL-8) and in plasma (pIL-8), C-reactive protein (CRP) and CEM were measured. rIL-8 levels [mean +/- 1 SD (standard deviation)] were higher in ACS (102.9 +/- 70.1 pg/mL) compared with CSA (44.7 +/- 22.8 pg/mL) (P < 0.001). No difference in pIL-8 levels between the two coronary artery disease groups was observed (P = 0.280). Serum CRP levels were correlated with rIL-8 levels (r = 0.294, P < 0.001); no association was found between CRP and pIL-8 levels (r = 0.025, P = 0.706). Further, rIL-8 had an independent association with ACS, when CRP and CEM were taken into consideration. CONCLUSION: This study shows for the first time that rIL-8 content was significantly higher in ACS, compared with CSA. These findings endorse results from our previous studies suggesting that erythrocytes may play an important role in the development of unstable atherosclerotic plaque.

Total cholesterol content of erythrocyte membranes is increased in patients with acute coronary syndrome: a new marker of clinical instability?

OBJECTIVES: We hypothesized that cholesterol content is increased in the circulating erythrocytes of patients with acute coronary syndrome (ACS) and may be a marker of clinical instability. We therefore sought to investigate whether cholesterol content differs in erythrocyte membranes of patients presenting with ACS compared to patients with chronic stable angina (CSA). BACKGROUND: Plaque rupture in ACS depends at least partly on the volume of the necrotic lipid core. Histopathologic studies have suggested that cholesterol transported by erythrocytes and deposited into the necrotic core of atheromatous plaques contributes to lipid core growth. METHODS: Consecutive angina patients were prospectively assessed; 120 had CSA (83 men, age 64 +/- 11 years) and 92 ACS (67 men, 66 +/- 11 years). Total cholesterol content in erythrocyte membranes (CEM) was measured using an enzymatic assay, and protein content was assessed by the Bradford method. RESULTS: The CEM (median and interquartile range) was higher (p < 0.001) in ACS patients (184 microg/mg; range 130.4 to 260.4 microg/mg) compared with CSA patients (81.1 microg/mg; range 53.9 to 109.1 microg/mg) (analysis of covariance). Total plasma cholesterol concentrations did not correlate with CEM levels (r = -0.046, p = 0.628). CONCLUSIONS: This study shows, for the first time, that CEM is significantly higher in patients with ACS compared with CSA patients. These findings suggest a potential role of CEM as a marker of atheromatous plaque growth and vulnerability. Large ad hoc studies are required to establish the clinical importance and pathogenic significance of CEM measurement.

Interleukin-18/interleukin-10 ratio is an independent predictor of recurrent coronary events during a 1-year follow-up in patients with acute coronary syndrome.

BACKGROUND: The pro-inflammatory cytokine IL-18 has been suggested to play a role in atherogenesis and atheromatous plaque rupture leading to the acute coronary syndrome (ACS). Conversely, the anti-inflammatory cytokine IL-10 seems to have an atheroprotective role. Patients with unstable coronary artery disease show an imbalance between serum levels of pro- and anti-inflammatory cytokines, and studies have shown that IL-18/IL-10 ratio is an independent predictor of adverse in-hospital events in patients with ACS. We assessed the long-term prognostic significance of admission interleukin-18 (IL-18)/interleukin-10 (IL-10) ratio for recurrent coronary events during a 1-year follow-up in patients presenting with an ACS. METHODS: We assessed independent predictors of the combined end-point using multiple logistic regression analysis, in 186 patients (138 men, 65+/-12 years) with ACS (75 STEMI, 65 NSTEMI and 46 unstable angina). The composite of cardiac death and re-hospitalization with non-fatal myocardial infarction, or unstable angina, was the pre-specified study end-point. Serum IL-10 and IL-18 levels were measured at study entry using commercially available ELISAs. RESULTS: During the 1-year follow-up, 48 (26%) patients had recurrent cardiac events and 138 (74%) were event-free. IL-18/IL-10 ratio predicted the occurrence of adverse cardiac events (OR 1.91, 95% CI 1.37-2.65, p<0.001), and was found to be an independent predictor among other established biochemical and clinical risk markers (OR 2.31, 95% CI 1.55-3.42, p<0.001). CONCLUSIONS: Serum IL-18/IL-10 ratio is an independent predictor of recurrent coronary events during long-term follow-up in patients presenting with ACS. Our study further supports the hypothesis that the balance between pro-inflammatory and anti-inflammatory cytokines may be an important determinant of patient outcome, suggesting a pathogenic role in plaque progression and instability.

Inflammatory and anti-inflammatory variable clusters and risk prediction in acute coronary syndrome patients: a factor analysis approach.

BACKGROUND: Numerous inflammatory mediators such as C-reactive protein (CRP), fibrinogen, interleukin-18 (IL-18), and inter-cellular adhesion molecule-1 (ICAM-1) have been proposed for risk stratification in acute coronary syndrome (ACS) patients. However, interactions between these markers have made it difficult to assess their true role in risk prediction. Factor analysis is a multivariable statistical technique that reduces a large number of intercorrelated variables to a smaller set of independent clusters, underlining physiological relationships. The aim of this study was to investigate, using factor analysis, a clustering of pro-inflammatory markers, anti-inflammatory cytokines such as interleukin-10 (IL-10) and HDL cholesterol, and to determine their role in prediction of risk of recurrent coronary events in ACS patients. METHODS: We assessed 320 consecutive patients (236 men; 67 years; IQ 58-74 years) admitted with ACS. The composite of cardiac death and re-hospitalization with non-fatal myocardial infarction, or unstable angina, was the pre-specified study end-point. Serum CRP, fibrinogen, HDL cholesterol, IL-10, IL-18 and ICAM-1 levels were measured at study entry. We assessed independent predictors of the combined end-point during a 1-year follow-up using multiple logistic regression analysis. RESULTS: Factor analysis identified three clusters which were arbitrarily interpreted as (1) a "systemic inflammation" cluster with positive loadings of CRP and fibrinogen, (2) a "local inflammation-endothelial dysfunction" cluster with positive loadings of IL-18 and ICAM-1 and (3) an "anti-inflammation" cluster comprising IL-10 and HDL cholesterol. Only the "anti-inflammation" cluster was a significant predictor (OR 0.66, 95% CI: 0.49-0.89) of adverse cardiac events during a 1-year follow-up and remained significant (OR 0.65, 95% CI: 0.48-0.88) in a multivariate model that included all three factors. CONCLUSIONS: Although inflammatory markers such as CRP predict future cardiovascular events in ACS patients, when all inflammatory mediators are taken into account in a prospective analysis of risk, markers reflecting anti-inflammatory mechanisms are better prognostic markers.

Apolipoprotein E genotype and circulating interleukin-10 levels in patients with stable and unstable coronary artery disease.

OBJECTIVES: This study was designed to assess the relation between apolipoprotein E (apoE) genotype and serum interleukin (IL)-10 levels in patients with acute coronary syndrome (ACS) and chronic stable angina (CSA). BACKGROUND: Genetic variations in the apoE gene affect the risk for coronary artery disease (i.e., carriers of the e4 allele have an increased risk). Increased levels of C-reactive protein (CRP), an inflammatory marker, correlate with an increased risk of acute coronary events, whereas increased IL-10 concentrations have an atheroprotective role. Studies have reported a negative association between the apoE e4 allele and CRP levels. METHODS: Apolipoprotein E genotypes were assessed in 166 consecutive ACS patients (119 men, mean age 68 years, interquartile range [IQR] 60 to 74 years) and 70 CSA patients (54 men, mean age 65 years, IQR 62 to 68 years). Serum IL-10 and CRP were assessed at study entry. RESULTS: Analysis of covariance showed that genetic variation in the apoE gene locus significantly influences serum IL-10 levels in both ACS (p = 0.009) and CSA patients (p = 0.013). Among ACS patients, IL-10 levels were lower in E3/E4 carriers compared with E3/E3 carriers (p = 0.01) and marginally lower compared with E2/E3 carriers (p = 0.065). Among CSA patients, IL-10 levels were lower in E3/E4 carriers compared with E2/E3 carriers (p = 0.004) and marginally lower compared with E3/E3 carriers (p = 0.086). CONCLUSIONS: The IL-10 concentrations differ in ACS and in CSA patients with different apoE genotypes. The e4 allele was associated with a trend toward lower IL-10 serum levels. Our results may provide an explanation of findings in previous studies that cardiovascular risk is higher in e4 carriers despite the presence of low CRP levels.

Levosimendan use reduces matrix metalloproteinase-2 in patients with decompensated heart failure.

BACKGROUND: Extracellular matrix metabolism (ECM) has an important role in left ventricular (LV) remodeling in chronic heart failure (CHF). Matrix metalloproteinases (MMPs) are involved in the regulation of extracellular matrix (ECM) metabolism. We investigated the effect of levosimendan, a novel calcium sensitizer, on serum levels of MMP-2. METHODS: Our study population consisted of 60 consecutive patients with advanced heart failure who were admitted to hospital with an acute decompensation of their CHF. Patients were randomized to levosimendan (n = 30; 18 men, aged 65 +/- 3 years) or placebo (n = 30; 15 men, aged 67 +/- 4 years). Serum MMP-2 levels were assessed before and after treatment with levosimendan or placebo, using a commercially available ELISA. RESULTS: Serum levels of MMP-2 were reduced from 427 ng/ml 95%CI 372-484 to 371 ng/ml 95%CI 329-413 in the levosimendan treated group and from 433 ng/ml 95%CI 422-444 to 425 ng/ml 95%CI 414-436 in the placebo group. Repeated measurements ANOVA showed that treatment with levosimendan significantly affected levels of MMP-2 (p = 0.019). CONCLUSIONS: The present study showed that levosimendan may beneficially affect ECM remodeling in patients with acutely decompensated CHF. Whether these effects translate into added clinical benefits, as suggested by an improved ejection fraction in the levosimendan group, deserves further investigation.