Alexandros D. Tselepis

Inflammation, bioactive lipids and atherosclerosis: potential roles of a lipoprotein-associated phospholipase A2, platelet activating factor-acetylhydrolase.

It is well established that inflammation is an integral feature of atherosclerosis and of the cardiovascular diseases which it underlies. Oxidative stress is also recognized as a key actor in atherogenesis, in which it is closely associated with the inflammatory response and bioactive lipid formation. Several bioactive lipids have been identified in the atherosclerotic plaque, including the potent inflammatory mediator platelet activating factor (PAF), PAF-like lipids, oxidised phospholipids (oxPL) and lysophosphatidylcholine (lyso-PC). Recent evidence has established a central role of two phospholipases (PL) in atherogenesis, the non-pancreatic Type II secretory phospholipase A(2) (sPLA(2)) and the lipoprotein-associated PLA(2)-alternatively termed as PAF-acetylhydrolase (PAF-AH). sPLA(2) is calcium-dependent and hydrolyses the sn-2 acyl group of glycerophospholipids of lipoproteins and cell membranes to produce lyso-PC and free fatty acids. It is also implicated in isoprostane production from oxPL. sPLA(2) is an acute phase reactant, which is upregulated by inflammatory cytokines and may represent a new independent risk factor for coronary heart disease. In contrast to sPLA(2), PAF-AH is calcium-independent and is specific for short acyl groups at the sn-2 position of the phospholipid substrate and with the exception of PAF, can equally hydrolyze oxPL to generate lyso-PC and oxidized fatty acids. Thus PAF-AH plays a key role in the degradation of proinflammatory oxPL and in the generation of lyso-PC and oxidized fatty acids. PAF-AH equally can also hydrolyze short-chain diacylglycerols, triacylglycerols, and acetylated alkanols, and displays a PLA(1) activity. Whereas sPLA(2) may represent a new independent risk factor for coronary artery disease, the potential relevance of PAF-AH to atherosclerosis remains the subject of debate, and recent results suggest that the potential role of the LDL-associated PAF-AH in atherogenesis may be distinct to that of the HDL-associated enzyme. This review is focused on the main structural and catalytic features of plasma PAF-AH, on the association of the enzyme with distinct lipoprotein particle subspecies, on its cellular sources, and finally on the potential significance of this lipoprotein-associated PLA(2) in cardiovascular disease.

PAF-Degrading Acetylhydrolase Is Preferentially Associated With Dense LDL and VHDL-1 in Human Plasma Catalytic Characteristics and Relation to the Monocyte-Derived Enzyme

In human plasma, platelet activating factor (PAF)-degrading acetylhydrolase (acetylhydrolase) is principally transported in association with LDLs and HDLs; this enzyme hydrolyzes PAF and short-chain forms of oxidized phosphatidylcholine, transforming them into lyso-PAF and lysophosphatidylcholine, respectively. We have examined the distribution, catalytic characteristics, and transfer of acetylhydrolase activity among plasma lipoprotein subspecies separated by isopycnic density gradient ultracentrifugation; the possibility that the plasma enzyme may be partially derived from adherent monocytes has also been evaluated. In normolipidemic subjects with Lp(a) levels < 0.1 mg/mL, acetylhydrolase was associated preferentially with small, dense LDL particles (LDL-5; d = 1.050 to 1.063 g/mL) and with the very-high-density lipoprotein-1 subfraction (VHDL-1; d = 1.156 to 1.179 g/mL), representing 23.9 +/- 1.7% and 20.6 +/- 3.2%, respectively, of total plasma activity. The apparent Km values for PAF of the enzyme associated with such lipoproteins were 89.7 +/- 23.4 and 34.8 +/- 4.5 mumol/L for LDL-5 and VHDL-1, respectively: indeed, the Km value for LDL-5 was some 10-fold higher than that of the light LDL-1, LDL-2, and LDL-3 subspecies, whereas the Km of VHDL-1 was some twofold greater than those of the HDL-2 and HDL-3 subspecies. Furthermore, when expressed on the basis of unit plasma volume, the Vmax of the acetylhydrolase associated with LDL-5 was some 150-fold greater than that in LDL-1 (d = 1.019 to 1.023 g/mL). No significant differences in the pH dependence of enzyme activity or in sensitivity to protease inactivation, sulfydryl reagents, the serine protease inhibitor Pefabloc, or the PAF antagonist CV 3988 could be detected between apo B-containing and apo A-I-containing lipoprotein particle subspecies. Incubation of LDL-1 (Km = 8.4 +/- 2.6 mumol/L) and LDL-2 (d = 1.023 to 1.029 g/mL; Km = 8.4 +/- 3.3 mumol/L) subspecies with LDL-5, in which acetylhydrolase had been inactivated by pretreatment with Pefabloc, demonstrated preferential transfer of acetylhydrolase to LDL-5. Acetylhydrolase transferred to LDL-5 from the light LDL subspecies exhibited a Km of 9.4 +/- 2.2 mumol/L, a value characteristic of the particle donors. Finally, acetylhydrolase (Km = 23.4 +/- 7.6 mumol/L) released by adherent human monocytes in culture was found to bind preferentially to small, dense LDL subspecies upon incubation of Pefabloc-inactivated plasma with monocyte supernatant.(ABSTRACT TRUNCATED AT 400 WORDS)

Atherogenic lipid profile is a feature characteristic of patients with early rheumatoid arthritis: effect of early treatment – a prospective, controlled study

We investigated lipid profiles and lipoprotein modification after immuno-intervention in patients with early rheumatoid arthritis (ERA). Fifty-eight patients with ERA who met the American College of Rheumatology (ACR) criteria were included in the study. These patients had disease durations of less than one year and had not had prior treatment for it. Smokers or patients suffering from diabetes mellitus, hypothyroidism, liver or kidney disease, Cushings syndrome, obesity, familiar dyslipidemia and those receiving medications affecting lipid metabolism were excluded from the study. Sixty-three healthy volunteers (controls) were also included. Patients were treated with methotrexate and prednisone. Lipid profiles, disease activity for the 28 joint indices score (DAS-28) as well as ACR 50% response criteria were determined for all patients. The mean DAS-28 at disease onset was 5.8 ± 0.9. After a year of therapy, 53 (91.3%) patients achieved the ACR 20% response criteria, while 45 (77.6%) attained the ACR 50% criteria. In addition, a significant decrease in the DAS-28, C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) were observed. ERA patients exhibited higher serum levels of total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and triglycerides, whereas their serum high-density lipoprotein cholesterol (HDL-C) levels were significantly lower compared to controls. As a consequence, the atherogenic ratio of TC/HDL-C as well as that of LDL-C/HDL-C was significantly higher in ERA patients compared to controls. After treatment, a significant reduction of the atherogenic ratio of TC/HDL-C as well as that of LDL-C/HDL-C was observed, a phenomenon primarily due to the increase of serum HDL-C levels. These changes were inversely correlated with laboratory changes, especially CRP and ESR. In conclusion, ERA patients are characterized by an atherogenic lipid profile, which improves after therapy. Thus, early immuno-intervention to control disease activity may reduce the risk of the atherosclerotic process and cardiovascular events in ERA patients.

“European Panel on Low Density Lipoprotein (LDL) Subclasses”: A Statement on the Pathophysiology, Atherogenicity and Clinical Significance of LDL Subclasses

Aim of the present Consensus Statement is to provide a comprehensive and up to-date document on the pathophysiology, atherogenicity and clinical significance of low density liproproteins (LDL) subclasses. We sub-divided our statement in 2 sections. section I discusses the pathophysiology, atherogenicity and measurement issues, while section II is focused on the effects of drug and lifestyle modifications. Suggestions for future research in the field are highlighted at the end of section II. Each section includes Conclusions.

Atorvastatin Does Not Affect the Antiplatelet Potency of Clopidogrel When It Is Administered Concomitantly for 5 Weeks in Patients With Acute Coronary Syndromes

Background—The antiplatelet effect of clopidogrel may be attenuated by short-term coadministration of lipophilic statins. We investigated whether the coadministration of atorvastatin for 5 weeks in patients with acute coronary syndromes (ACS) could affect the antiplatelet potency of clopidogrel. Methods and Results—Forty-five hypercholesterolemic patients with the first episode of an ACS were included in the study. Patients were randomized to receive daily either 10 mg of atorvastatin (n=21) or 40 mg of pravastatin (n=24). Thirty patients who underwent percutaneous coronary intervention (PCI) received a loading dose of 375 mg of clopidogrel, followed by 75 mg/d for at least 3 months. In the remaining 15 patients who refused to undergo PCI, clopidogrel therapy was not administered. Eight normolipidemic patients with the first episode of an ACS were also included and received only clopidogrel. The serum levels of soluble CD40L and the adenosine 5′-diphosphate– or thrombin receptor activating peptide-14–induced platelet aggregation, as well as P-selectin and CD40L surface expression, were studied at baseline (within 30 minutes after admission) and 5 weeks later. Neither atorvastatin nor pravastatin significantly influenced the clopidogrel-induced inhibition of platelet activation, nor did clopidogrel influence the therapeutic efficacy of atorvastatin. Conclusions—Atorvastatin does not affect the antiplatelet potency of clopidogrel when coadministered for 5 weeks in ACS patients.

Laparoscopic surgery-induced changes in oxidative stress markers in human plasma.

Background: The induction of the pneumoperitoneum increases intraabdominal pressure (IAP), causing splanchnic ischemia, whereas its deflation normalizes IAP and splanchnic blood flow. This procedure appears to represent an ischemia-reperfusion model in humans. Methods: Thirty laparoscopic cholecystectomies (LC) were performed in 30 patients with a mean age of 54.6 ± 15.6 years. A group of 20 patients mean age, 57.3 ± 9.65 who underwent open cholecystectomy (OC) was also studied. Vein plasma levels of thiobarbituric acid-reactive substances (TBARS), a marker of free radical production; plasma total antioxidant status (TAS); and uric acid (UA) levels were measured preoperatively, 5 min after deflation of the pneumoperitoneum or at the end of operation, and 24 h postoperatively. Aspartate aminotransferase (AST), alanine aminotransferase (ALT), and total bilirubin (TBL) levels were measured preoperatively and 24 h after the operation. Results: In the LC group, significant elevations in the concentration of TBARS were observed in the early postoperative measurements in comparison with the preoperative measurements. TAS and UA levels were decreased significantly 24 h postoperatively compared to preoperative levels. The postoperative levels of AST, ALT, and TBL increased significantly in comparison with the preoperative levels. In the OC group, no alterations in the concentration of TBARS were observed in the postoperative period. The other parameters had changes similar to those recorded for the LC group. Conclusions: Free radical-induced lipid peroxidation associated with a decrease in plasma antioxidant capacity and UA levels as well as altered hepatic function is observed after deflation of the pneumoperitoneum. These results suggest that free radicals are generated at the end of a laparoscopic procedure, possibly as a result of an ischemia-reperfusion phenomenon induced by the inflation and deflation of the pneumoperitoneum.

The role of lipoprotein-associated phospholipase A2 in atherosclerosis may depend on its lipoprotein carrier in plasma.

Platelet-activating factor (PAF) acetylhydrolase exhibits a Ca(2+)-independent phospholipase A2 activity and degrades PAFas well as oxidized phospholipids (oxPL). Such phospholipids are accumulated in the artery wall and may play key roles in vascular inflammation and atherosclerosis. PAF-acetylhydrolase in plasma is complexed to lipoproteins; thus it is also referred to as lipoprotein-associated phospholipase A2 (Lp-PLA2). Lp-PLA2 is primarily associated with low-density lipoprotein (LDL), whereas a small proportion of circulating enzyme activity is also associated with high-density lipoprotein (HDL). The majority of the LDL-associated Lp-PLA2 (LDL-Lp-PLA2) activity is bound to atherogenic small-dense LDL particles and it is a potential marker of these particles in plasma. The distribution of Lp-PLA2 between LDL and HDL is altered in various types of dyslipidemias. It can be also influenced by the presence of lipoprotein (a) [Lp(a)] when plasma levels of this lipoprotein exceed 30 mg/dl. Several lines of evidence suggest that the role of plasma Lp-PLA2 in atherosclerosis may depend on the type of lipoprotein particle with which this enzyme is associated. In this regard, data from large Caucasian population studies have shown an independent association between the plasma Lp-PLA2 levels (which are mainly influenced by the levels of LDL-Lp-PLA2) and the risk of future cardiovascular events. On the contrary, several lines of evidence suggest that HDL-associated Lp-PLA2 may substantially contribute to the HDL antiatherogenic activities. Recent studies have provided evidence that oxPL are preferentially sequestered on Lp(a) thus subjected to degradation by the Lp(a)-associated Lp-PLA2. These data suggest that Lp(a) may be a potential scavenger of oxPL and provide new insights into the functional role of Lp(a) and the Lp(a)-associated Lp-PLA2 in normal physiology as well as in inflammation and atherosclerosis. The present review is focused on recent advances concerning the Lp-PLA2 structural characteristics, the molecular basis of the enzyme association with distinct lipoprotein subspecies, as well as the potential role of Lp-PLA2 associated with different lipoprotein classes in atherosclerosis and cardiovascular disease.

Differential Effect of Hypolipidemic Drugs on Lipoprotein-Associated Phospholipase A2

Objective— Lipoprotein-associated phospholipase A2 (Lp-PLA2) is a predictor for incident atherosclerotic disease. We investigated the effect of 3 hypolipidemic drugs that exert their action through different mechanisms on plasma and lipoprotein-associated Lp-PLA2 activity and mass. Methods and Results— In 50 patients with Type IIA dyslipidemia were administered rosuvastatin (10 mg daily), whereas in 50 Type IIA dyslipidemic patients exhibiting intolerance to previous statin therapy were administered ezetimibe as monotherapy (10 mg daily). Fifty patients with Type IV dyslipidemia were given micronised fenofibrate (200 mg daily). Low- and high-density lipoprotein (LDL and HDL, respectively) subclass analysis was performed electrophoretically, whereas lipoprotein subfractions were isolated by ultracentrifugation. Ezetimibe reduced plasma Lp-PLA2 activity and mass attributable to the reduction in plasma levels of all LDL subfractions. Rosuvastatin reduced enzyme activity and mass because of the decrease in plasma levels of all LDL subfractions and especially the Lp-PLA2 on dense LDL subfraction (LDL-5). Fenofibrate preferentially reduced the Lp-PLA2 activity and mass associated with the VLDL+IDL and LDL-5 subfractions. Among studied drugs only fenofibrate increased HDL-associated Lp-PLA2 (HDL-Lp-PLA2) activity and mass attributable to a preferential increase in Lp-PLA2 associated with the HDL-3c subfraction. Conclusion— Ezetimibe, rosuvastatin, and fenofibrate reduce Lp-PLA2 activity and mass associated with the atherogenic apoB-lipoproteins. Furthermore, fenofibrate improves the enzyme specific activity on apoB-lipoproteins and induces the HDL-Lp-PLA2. The clinical implications of these effects remain to be established.

Clinical importance and therapeutic modulation of small dense low-density lipoprotein particles

The National Cholesterol Education Programme Adult Treatment Panel III accepted the predominance of small dense low-density lipoprotein (sdLDL) as an emerging cardiovascular disease (CVD) risk factor. Most studies suggest that measuring low-density lipoprotein (LDL) particle size, sdLDL cholesterol content and LDL particle number provides additional assessment of CVD risk. Therapeutic modulation of small LDL size, number and distribution may decrease CVD risk; however, no definitive causal relationship is established, probably due to the close association between sdLDL and triglycerides and other risk factors (e.g., high-density lipoprotein, insulin resistance and diabetes). This review addresses the formation and measurement of sdLDL, as well as the relationship between sdLDL particles and CVD. The effect of hypolipidaemic (statins, fibrates and ezetimibe) and hypoglycaemic (glitazones) agents on LDL size and distribution is also discussed.

Increased plasma levels of visfatin/pre-B cell colony-enhancing factor in obese and overweight patients with metabolic syndrome.

Background: Visfatin [pre-B cell colony-enhancing factor (PBEF)] is a cytokine highly expressed in visceral fat that exhibits insulin-mimetic properties. However, its role in insulin-resistant states, such as in metabolic syndrome (MetS), remains largely unknown. Objective: To investigate the possible differences of plasma visfatin levels between obese and overweight subjects with and without MetS. Design and patients: Plasma visfatin concentrations were measured with enzyme-linked immunosorbent assay (ELISA) in 28 overweight and obese [body mass index (BMI)>28 kg/m2] subjects with Mets and 28 age- and sex-matched overweight and obese (BMI>28 kg/m2) individuals without MetS (control group). Results: Patients with MetS exhibited significantly elevated waist circumference (WCR) values, higher blood pressure readings, higher fasting glucose and triglyceride concentrations as well as lower levels of HDL cholesterol (HDL-C) compared with controls. Total and LDL cholesterol (LDL-C) concentrations did not differ significantly between the two groups. Plasma visfatin concentrations were significantly higher in subjects with MetS compared with controls [27 (16–65) ng/ml vs 19 (10–47) ng/ml, p<0.05]. The same results were observed even after adjustment for age, sex and BMI. Plasma visfatin levels were positively correlated with age (r=0.32, p<0.05), WCR (r=0.31, p<0.05), triglyceride (r=0.59, p<0.01) and glucose (r=0.33, p<0.05) levels and were negatively correlated with HDL-C levels (r=−0.38, p<0.05). Multiple linear regression analysis revealed similar results. Conclusion: Plasma visfatin levels are increased in overweight and obese subjects with MetS compared with those individuals who do not fulfil the criteria for the diagnosis of MetS.