Luca Puccetti

Time‐dependent effect of statins on platelet function in hypercholesterolaemia

Background Reduction of platelet activity induced by statins has been described as a positive effect exerted by such molecules on vascular thrombotic events. However, the relations among cholesterol (LDL‐C) reduction, the timing of the antiplatelet effect, the involved mechanisms and the doses of each statin able to reduce platelet function are not actually well known. The aim of our study was to evaluate the impact of simvastatin (20 mg day−1), atorvastatin (10 mg day−1), fluvastatin (40 mg day−1) and pravastatin (40 mg day−1) on platelet function in hypercholesterolaemic subjects with relation to (LDL‐C), oxidized‐LDL (ox‐LDL) and antiport mechanism modifications.

Investigations of the esterase, phosphatase, and sulfatase activities of the cytosolic mammalian carbonic anhydrase isoforms I, II, and XIII with 4-nitrophenyl esters as substrates.

The esterase, phosphatase, and sulfatase activities of carbonic anhydrase (CA, EC 4.2.1.1) isozymes, CA I, II, and XIII with 4-nitrophenyl esters as substrates was investigated. These enzymes show esterase activity with 4-nitrophenyl acetate as substrate, with second order rate constants in the range of 753-7706M(-1)s(-1), being less effective as phosphatases (k(cat)/K(M) in the range of 14.89-1374.40M(-1)s(-1)) and totally ineffective sulfatases. The esterase/phosphatase activities were inhibited by sulfonamide CA inhibitors, proving that the zinc-hydroxide mechanism responsible for the CO(2) hydrase activities of CAs is also responsible for their esterase/phosphatase activity. CA XIII was the most effective esterase and phosphatase. CA XIII might catalyze other physiological reactions than CO(2) hydration, based on its relevant phosphatase activity.

Selective Inhibition of Carbonic Anhydrase IX Decreases Cell Proliferation and Induces Ceramide-Mediated Apoptosis in Human Cancer Cells

Recently, carbonic anhydrase (CA) inhibitors have been proposed as a potential new class of antitumor agents. The aim of this study was to evaluate the antitumor activity of three CA inhibitors, namely acetazolamide (AZ) and two newly synthesized aromatic sulfonamides with high affinity for CA IX, 2-(4-sulfamoylphenyl-amino)-4,6-dichloro-1,3,5-triazine (TR1) and 4-[3-(N,N-dimethylaminopropyl)thioreidophenylsulfonylaminoethyl]benzenesulfonamide (GA15), against human tumor cells. The effects of AZ, TR1, and GA15 on cell proliferation and apoptosis were evaluated in CA IX-positive HeLa and 786-O cells and CA IX-negative 786-O/von Hippel-Lindau (VHL) cells. We also investigated whether the potential antitumor activity of these molecules might be mediated by an increase in ceramide production. AZ, TR1, and GA15 could significantly reduce cell proliferation and induce apoptosis in HeLa and 786-O cells. Moreover, all three inhibitors could decrease intracellular pH (pHi) and increase ceramide production in the same cells. Treatment with the ceramide synthase inhibitor fumonisin B1 prevented the apoptotic effects of the three CA inhibitors. In all experiments, the effects of aromatic sulfonamides were more pronounced than those of AZ. The three inhibitors did not show any antitumor activity in CA IX-negative 786-O/VHL cells and failed to lower pHi or increase intracellular ceramide levels in the same cells. In conclusion, CA inhibition can decrease cell proliferation and induce apoptosis in human tumor cells. The ability of CA inhibitors to decrease pHi might trigger cell apoptosis through mediation of ceramide synthesis. Activation of this apoptotic cascade probably is mediated by inhibition of the CA IX isoform.

Platelet hyperactivity after statin treatment discontinuation

Hydroxymethyl-glutaryl-CoA-reductase inhibitors (statins) reduce cardiovascular events by cholesterol lowering as well as by non-lipid related actions. Among them, the modulation of platelet activity could play a relevant role in vascular protection. Furthermore withdrawal of statins has been associated with increased cardiovascular event rate. The aim of our study was to evaluate platelet activity after cerivastatin discontinuation in eighteen subjects that did not accept other drugs and in sixteen subjects continuing treatment with simvastatin. Fourteen subjects at the end of the discontinuation period decided to receive other drugs (simvastatin) and they were evaluted six weeks later. We measured complete lipid profile by the chromogenic method (LDL-C was calculated); oxidized-LDL (ox-LDL; ELISA), platelet P-selectin (P-sel) expression (flow cytometry detection), platelet aggregation (% change of transmitted light), intracellular citrullin production (iCit; HPLC) as an indicator of intracellular NO synthase activity at baseline and 7, 14, 28, 60 days after statin discontinuation. P-sel expression and platelet aggregation were increased at 14 days (p < 0.001 and p < 0.05) in association with raised ox-LDL (r = 0.30, p < 0.05) and decreased iCit (r = 0.53, p < 0.01). Increased LDL-C was related to P-sel and platelet aggregation at 28 days (r = 0.30, p < 0.05). Subjects continuing statin treatment had no significant changes of P-sel at 28 (p = 0.221) and 60 days (p = 0.238). Subjects treated with simvastatin after 60 days of diet showed a significant reduction of P-sel and platelet aggregation after six weeks of treatment (p < 0.01). Our data suggest a platelet hyperactivation state in the second week after statin discontinuation which is partially related to raised LDL-C. Such a finding could participate in the increased cardiovascular event rate after statin discontinuation.

Atorvastatin reduces platelet-oxidized-LDL receptor expression in hypercholesterolaemic patients

Background  Oxidized‐LDL (ox‐LDL) are proatherogenic and platelet‐activating molecules. Atorvastatin reduces platelet activity before cholesterol‐lowering action. CD36 and lectin‐like oxidized‐LDL receptor‐1 (LOX‐1) are specific ox‐LDL receptors expressed also in platelets. This study was planned to address whether the possible rapid effect of atorvastatin on platelets could be related to modulation of ox‐LDL receptors.

Erratum to “Pro/Anti-Inflammatory Cytokine Imbalance in Postischemic Left Ventricular Remodeling”

The authors would like to inform the correct names for the present paper as stated above.

Effects of carvedilol on left ventricular remodeling and systolic function in elderly patients with heart failure

Recent studies have shown that carvedilol therapy in patients with heart failure improves clinical outcome and survival, however, the effects of such treatment on left cardiac morphology and function in elderly patients with severe heart failure has not been widely studied.

Pro-inflammatory/anti-inflammatory cytokine imbalance in acute coronary syndromes

The aim of this study was to evaluate the presence of an imbalance between proinflammatory and anti-inflammatory mediators in patients affected by acute coronary syndromes (ACS). We considered two groups of 26 and 28 patients with acute myocardial infarction (AMI) and unstable angina (UA) respectively, compared with a group of 30 patients with stable angina and 30 healthy volunteers. We evaluated the production in cultured and stimulated lymphomonocytes of interferon (IFN)γ and tumour necrosis factor (TNF)α, which are well known to possess proinflammatory effects, and of interleukin (IL)10, which has been shown to have a protective anti-inflammatory activity. We also assessed the clinical characteristics of groups and, particularly, we evaluated the circulating levels of C-reactive protein (hs-CRP). We found a significant increase of IFNγ and TNFα production (P<0.01) and a significant decrease of IL10 production (P<0.05) in cultures of lymphomonocytes taken from patients with AMI and UA compared with SA patients and controls. No significant changes where found between AMI and UA patients and SA patients and controls. Circulating levels of hs-CRP were significantly increased (P<0.01) in patients with ACS compared with the other control groups. Our data showed an increased production of proinflammatory mediators in ACS that may be detectable both in circulating blood and in cell cultures where it is possible to evaluate in a better way the functional state of cells; this finding was associated with a reduced production of the antiinflammatory cytokine IL10. In conclusion, a relevant imbalance is present in ACS and this fact could contribute to plaque instability and clinical manifestations.

Different effect of statins on platelet oxidized-LDL receptor (CD36 and LOX-1) expression in hypercholesterolemic subjects.

Hydroxymethyl-glutaryl-CoA-reductase inhibitors (statins) reduce cardiovascular mortality by decreasing cholesterol as well as by non-lipid-related actions. Oxidized low-density lipoproteins (ox-LDL) are pro-atherogenic molecules and potent platelet agonists. CD36 and lectin-like ox-LDL receptor-1 (LOX-1) are specific ox-LDL receptors also expressed in platelets. This study was planned to address whether treatment with atorvastatin 10 mg/day, pravastatin 40 mg/day or simvastatin 20 mg/day could affect platelet CD36 and LOX-1 expression. Twenty-four patients for each treatment were evaluated after 3, 6, and 9 days and at 6 weeks for complete lipid profile (chromogenic), ox-LDL (ELISA), platelet P-selectin (P-sel), CD36, LOX-1 (FACS), and intracellular citrullin recovery (iCit) (HPLC). Data show hyperactivated platelets (P-sel absolute values, percent variation in activated cells, all p < 0.001), and CD36 and LOX-1 overexpression (all p < 0.001) in patients at baseline. P-sel, CD36, and LOX-1 were significantly decreased by atorvastatin and simvastatin (all p < 0.01) and related with iCit increase (r = 0.58,p < 0.001) and platelet-associated ox-LDL (r = 0.51, p < 0.01) at 9 days. Pravastatin reduced LOX-1 and P-sel (p < 0.05) at 6 weeks in relation with decreased LDL and ox-LDL (r = 0.39, p < 0.01 and r = 0.37, p < 0.01, respectively). These data suggest that atorvastatin and simvastatin reduce platelet activity by exposure of CD36 and LOX-1 before significant LDL reduction, whereas pravastatin action is detected later and in relation with LDL and ox-LDL lowering. Rapid and consistent reduction of CD36 and LOX-1 could be considered a direct anti-atherothrombotic mechanism related to the role of ox-LDL in platelet activation, platelet-endothelium interactions, and NO synthase activity.

Effects of carvedilol therapy on restrictive diastolic filling pattern in chronic heart failure

BACKGROUND Carvedilol therapy during congestive heart failure demonstrated a good efficacy in mortality rate reduction and in improvement of left ventricular (LV) systolic performance. However, currently there is not any finding about the drugs effect on diastolic filling. The aim of this study was to evaluate the effects of beta-blocker treatment on LV diastolic function with an eco-pulsed Doppler ultrasound scanning examination at transmitral level in a group of patients who were affected by heart failure with a restrictive filling pattern. METHODS We studied 27 patients with idiopathic or ischemic dilated cardiomyopathy with LV severe systolic disfunction (ejection fraction <35%). Fourteen patients were randomized to receive carvedilol treatment (carvedilol group), and 13 patients continued to receive standard therapy with angiotensin-converting enzyme inhibitors, diuretics, and vasodilators (placebo group). All patients underwent an echo-Doppler ultrasound scanning examination at the beginning of the study and after 4 and 12 months of treatment. RESULTS In the carvedilol group, we found a progressive improvement of Doppler ultrasound scanning parameters after 4 months, with a significant increase of A wave (P <.005), deceleration time (DT; P <.02) and isovolumetric relaxation time (IVRT; P <.02). These improvements were confirmed after 1 year of follow-up, whereas patients in the placebo group did not shown any significant modifications. After 1 year, the differences in these groups were more significant for A wave (39 +/- 4 cm/sec carvedilol group vs 30 +/- 4 cm/sec placebo group; P <.0001), for E/A ratio (1.8 +/- 0.2 carvedilol group vs 2.6 +/- 0.5 placebo group; P <.0002), for DT 1(40 +/- 16 msec carvedilol group vs 112 +/- 13 msec placebo group; P <.001), and for IVRT (74 +/- 8 msec carvedilol group vs 57 +/- 7 mesc placebo group; P <.0002). These changes seem to happen before systolic and morphological modifications. CONCLUSION Our results show that carvedilol therapy is a means of modifying parameters of diastolic filling favorably in patients with heart failure. These effects seem to be independent of those of systolic function. The improvement of systolic performance occurs after 1 year of treatment. The restrictive filling pattern, related to an unfavorable prognosis, changes toward pseudonormal or altered relaxation pattern during carvedilol therapy. Further investigations with a greater sample size will be necessary to confirm our findings.