Costas Tentolouris

Role of inflammation and oxidative stress in endothelial progenitor cell function and mobilization: Therapeutic implications for cardiovascular diseases

Endothelial progenitor cells (EPCs) are mobilized from the bone marrow into the peripheral circulation, home to sites of injury, and incorporate into foci of neovascularization, thereby improving blood flow and tissue recovery. Patients with cardiovascular diseases, including coronary artery disease, heart failure, hypertension, and diabetes, have been shown to exhibit reduced number and functional capacity of EPCs. Considerable evidence indicates that EPCs constitute an important endogenous system to maintain endothelial integrity and vascular homeostasis, while reduced number of EPCs has recently been shown to predict future cardiovascular events. Thus, enhancement of EPCs could be of potential benefit for individuals with cardiovascular diseases. The interplay between inflammation and oxidative stress is involved in the initiation, progression, and complications of cardiovascular diseases. Emerging evidence from in vitro and clinical studies suggests that inflammatory and oxidative changes influence EPC mobilization. Drugs with anti-inflammatory and antioxidant properties, currently administered to patients with cardiovascular diseases, such as statins, have been demonstrated to exert beneficial effects on EPC biology. A better understanding of the inflammatory and oxidative mechanisms leading to the numerical and functional impairment of EPCs would provide additional insight into the pathogenesis of cardiovascular disease and create novel therapeutic targets.

Effects of combined administration of low dose atorvastatin and vitamin E on inflammatory markers and endothelial function in patients with heart failure.

Heart failure has been associated with impaired endothelial function, increased inflammatory process and elevated oxidative stress status. Both statins and vitamin E separately improve endothelial function in patients with hypercholesterolemia and/or advanced atherosclerosis.

Oxidative stress, antioxidant vitamins, and atherosclerosis: From basic research to clinical practice

Abstract.Oxidative stress is involved in the pathogenesis of atherosclerosis, while a variety of antioxidants has been used in clinical studies, during the past few years, for the prevention and treatment of atherosclerosis. In small clinical studies it was found that both vitamins C and E may improve endothelial function in patients with risk factors for atherosclerosis such as diabetes mellitus, smoking, hypertension, or hypercholesterolemia. However, the initial, hopeful reports regarding the beneficial role of antioxidant vitamins against atherosclerosis, derived from purely observational studies, were followed by the negative results of almost all large randomized trials. Therefore, treatment with antioxidant vitamins C and E should not be recommended for the prevention or treatment of coronary atherosclerosis. New antioxidant strategies are needed to clarify the exact role of antioxidant treatment in coronary atherosclerosis.Zusammenfassung.In der Pathogenese der Atherosklerose spielt der oxidative Stress eine Rolle. Eine Reihe von verschiedenen Antioxidanzien wurde in klinischen Untersuchungen in den letzten Jahren geprüft, und zwar für die Prävention und Behandlung der Atherosklerose. In kleinen klinischen Studien wurde belegt, dass sowohl Vitamin C als auch E die endotheliale Funktion bei Patienten mit Risikofaktoren für Atherosklerose wie Diabetes mellitus, Rauchen, Hypertonie und Hypercholesterinämie verbessern. Die initialen und hoffnungsvollen Berichte bezüglich der günstigen Rolle von oxidativen Vitaminen zur Behandlung der Atherosklerose, abgeleitet von Beobachtungsstudien, wurden von negativen Ergebnissen in großen randomisierten Studien gefolgt. Die Behandlung mit Vitamin C und E kann deshalb nicht empfohlen werden für die Prävention und Behandlung der koronaren Atherosklerose. Weitere Untersuchungen sind deshalb notwendig, um die exakte Rolle dieser Behandlungsmaßnahme bei koronarer Herzerkrankung zu klären.

Chemokines in patients with ischaemic heart disease and the effect of coronary angioplasty

Percutaneous coronary transluminal angioplasty (PTCA) may release inflammatory mediators such as chemokines. Monocyte chemoattractant protein-1 (MCP-1) and eotaxin (EOX) are monocyte- and eosinophil-specific chemokines involved in the inflammation and pathogenesis of coronary atherosclerosis. A total of 28 patients undergoing elective PTCA, 20 coronary artery disease (CAD) patients undergoing coronary angiography and 28 healthy controls were studied. In PTCA patients before the procedure, MCP-1 plasma levels (441+/-64 pg/ml) were similar to those of CAD patients (430+/-24 pg/ml), and significantly higher compared with controls (145+/-17 pg/ml, P<0.01). MCP-1 rose significantly after 3 and 6 months following PTCA (696+/-89 and 876+/-86 pg/ml, respectively, P<0.01 vs. before PTCA). EOX plasma levels (155+/-14 pg/ml) were similar to those of CAD patients (157+/-14 pg/ml), but significantly higher compared with controls (83.2+/-10 pg/ml, P<0.05). EOX rose significantly 24 h (273+/-41 pg/ml, P<0.05) but not 3 months after PTCA (160+/-20 and 158+/-19 pg/ml, respectively). These findings indicate that chemokine-induced monocyte- and eosinophil-specific chemoattraction is stimulated in patients with coronary artery disease. MCP-1 levels remain significantly elevated for at least 6 months following elective PTCA, suggesting an inflammatory stimulation.

Basal and flow-mediated nitric oxide production by atheromatous coronary arteries

OBJECTIVES This study assessed the effects of inhibition of nitric oxide synthesis on epicardial human coronary arteries and on coronary flow velocity during baseline conditions and during atrial pacing. BACKGROUND Epicardial coronary artery dilation occurs in response to an increase in heart rate. It is not known whether the dilation of both angiographically normal and diseased epicardial coronary arteries during atrial pacing is nitric oxide dependent in humans. METHODS The effects of an intracoronary infusion (4 mumol/min for 8 min) of NG-monomethyl-L-arginine (LNMMA), an inhibitor of nitric oxide synthesis, was studied in 16 patients with coronary artery disease and in 6 patients with normal coronary arteriograms. In all patients atrial pacing was performed during normal saline and during LNMMA infusion. the lumen diameter of epicardial coronary arteries was assessed by quantitative angiography, and changes in blood flow velocity were measured with a Doppler catheter. RESULTS During saline infusion a significant increase in the lumen diameter of the proximal (p < 0.05) and distal (p < 0.01) segments of both normal and diseased arteries occurred during atrial pacing. No significant lumen diameter changes occurred in either group when atrial pacing was performed during LNMMA infusion. Stenosis diameter decreased during LNMMA infusion but did not change with atrial pacing either during saline infusion or during LNMMA infusion. The mean percent change in coronary blood flow with atrial pacing was less (p < 0.05) during LNMMA infusion than during saline infusion in both groups. CONCLUSIONS These findings confirm that epicardial coronary artery dilation induced by pacing is nitric oxide dependent. Nitric oxide production contributes to the vasomotor tone of coronary resistance vessels. Nitric oxide is produced at the site of atheromatous stenosis but is unaffected by pacing.

Coronary stenosis dilatation induced by L-arginine

L-arginine, a substrate in the production of endotheliumderived nitric oxide (NO), may stimulate the release of NO. L-arginine improves the coronary blood flow response to acetylcholine in patients with normal coronary arteries and hypercholesterolaemia. However, it is unknown whether local administration of L-arginine dilates coronary stenoses in patients with stable angina. This study assessed the effects of L-arginine and D-arginine at the stenotic site in patients with coronary artery disease and stable angina. 15 patients with chronic stable angina, coronary artery disease, and a positive treadmill exercise test ( 0·1 mV ST segment depression) at between 5 and 7 METS with the modified Bruce protocol, were studied in the cardiac catheterisation laboratory of the Hippokration Hospital, Athens University. The protocol was approved by the research ethics committee of the Hippokration Hospital and each patient gave written informed consent. Antianginal medication was stopped 24 h before the study. The patients were allowed to use sublingual glyceryl trinitrate as necessary, but no study was done within 3 h of its administration. None of the study patients had taken glyceryl trinitrate on the day of the study. Following the diagnostic coronary angiogram, an optimum radiographic projection was selected and kept constant for subsequent angiograms. In ten patients (seven male, three female, mean age 58 [SD 8] years) intracoronary infusion of 0·9% saline (2 mL/min) for 2 min was followed by intracoronary infusion of 150 μmol/min of L-arginine for 8 min. In five patients (four male, one female, mean age 61 [10] years) the same protocol was used, substituting 150 μmol/min of D-arginine for L-arginine. The chosen dose and the infusion time of L-arginine administration was consistent with a study showing a response after L-arginine administration in the brachial artery. The arterial segments in each frame were analysed at the Hammersmith Hospital in random order with quantitative computerised analysis, with an automated edge contour detection analysis system. The percentage change in luminal diameter from baseline was calculated at the end of the 8 min infusions and the values for L-arginine and D-arginine and normal saline were compared with Student’s t-test. The infusions of L-arginine and D-arginine did not affect systolic blood pressure (mean [SE]) (L-arginine: mean 152 [7·1] vs 150 [7·2] mm Hg, p=0·82 D-arginine: 154 [11·2] vs 157 [11·6] mm Hg, before and after infusions, respectively, p=0·27). L-arginine administration was associated with significant dilatation of stenoses (11·7 [2·3]%: p=0·004 vs D-arginine, p=0·001 vs normal saline) (figure), but there was no significant change with D-arginine (2·0 [0·04]%) (figure). No patients were receiving longacting nitrates, and the previous antianginal treatment did not affect the response to L-arginine ( -blockers: 11·1 [6·0]%; calcium antagonists 10·5 [4·7]%; -blockers and calcium antagonists 12·5 [3·4]%). Arginine, a semiessential aminoacid serves as the substrate for the enzyme NO synthase, which converts arginine to citrulline and NO, reducing vascular tone. A recent study in experimental models suggested possible beneficial effects of L-arginine administration. Long-term oral therapy with L-arginine reduced endothelial dysfunction and inhibited the development of atherosclerosis, but the mechanisms by which it exerts its vasodilator effect are not completely understood. Possible explanations include an effect on endothelial synthesis and release of NO, and direct or indirect physiological actions on the vascular smooth muscle to augment its responsiveness to NO. In our study a significant dilation was found in coronary stenotic segments after L-arginine administration in patients with coronary artery disease and stable angina. These data suggest that the 25

L-Arginine in cardiovascular disease: dream or reality?:

l-arginine is the substrate for endothelial nitric oxide synthase (eNOS), and the precursor for the synthesis of nitric oxide (NO). This amino acid exerts a number of actions in the cardiovascular system, mainly through the production of NO. However, it also has a number of NO-independent properties, such as the ability to regulate blood and intracellular pH and the effect on the depolarization of endothelial cell membranes. It also has antihypertensive and antioxidant properties, it influences blood viscosity and the coagulation/fibrinolysis system, and it affects the metabolism of glucose, lipids and proteins. L-arginine influences a number of atherosclerosis risk factors such as hypercholesterolemia, hypertension and smoking, improving endothelial function in these patients. However, it does not affect endothelial function in patients with diabetes mellitus. The role of L-arginine in coronary artery disease is still controversial, but it seems that oral or parenteral administration of this amino acid restores endothelial function in the brachial artery and improves coronary microcirculation. The role of L-arginine in heart failure is currently under investigation, and the first results are rather hopeful. In conclusion, L-arginine seems to provide a hopeful prospect for the treatment of cardiovascular diseases. However, more data derived from large-scale prospective studies evaluating the effects of long-term treatment with L-arginine are needed.

Effects of combined administration of vitamins C and E on reactive hyperemia and inflammatory process in chronic smokers

Purpose of this study was to investigate the effect of combined administration of antioxidant vitamins C and E on endothelial function and serum levels of inflammatory markers such as tumor necrosis factor alpha (TNF-alpha), interleukines 1b (IL-1b) and 6 (IL-6), vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1) and E-selectin in chronic smokers. Forty-three smokers were randomly divided into four groups receiving vitamin C 2 g/day (group A), vitamin C 2 g/day plus vitamin E 400 IU/day (group B), vitamin C 2 g/day plus vitamin E 800 IU/day (group C) or no antioxidant treatment (group D), for 4 weeks. Forearm blood flow (FBF) was measured using venous occlusion strain gauge plethysmography. Forearm vasodilatory response to reactive hyperemia (RH%) was expressed as the percentage change from baseline to post reactive hyperemia blood flow. RH% was significantly increased in groups B (P<0.05) and C (P<0.01), but remained unaffected in groups A and D. Serum levels of IL-1b, IL-6, sVCAM-1 and sICAM-1 were significantly reduced in group C (P<0.05, respectively), but remained unaffected in groups A, B and D. Thus, short term administration of vitamins C (2 g/day) and E (800 IU/day) reduces serum levels of IL-1b, IL-6, sVCAM-1 and sICAM-1, and improves forearm vasodilatory response to reactive hyperemia in healthy young smokers, while monotherapy with vitamin C alone is ineffective.

Effects of rosuvastatin and allopurinol on circulating endothelial progenitor cells in patients with congestive heart failure: the impact of inflammatory process and oxidative stress.

OBJECTIVE Endothelial progenitor cells (EPCs) contribute to the maintenance of endothelial integrity and function. We investigated the effects of rosuvastatin and allopurinol on the number of EPCs in patients with heart failure and aimed to provide insight into the molecular inflammatory and oxidative mechanisms that could be responsible for the alterations in EPC levels after treatment. METHODS Sixty patients with systolic heart failure were randomized to receive rosuvastatin 10mg/d, allopurinol 300mg/d or placebo and followed up for 1 month. The number of CD34(+)/KDR(+) and CD34(+)/CD133(+)/KDR(+) EPCs in blood was evaluated by flow cytometry. Endothelial function was assessed by brachial artery flow-mediated dilation. Levels of markers of inflammation and oxidative stress were also determined. RESULTS Circulating EPCs were significantly increased after rosuvastatin treatment (from 230 (170-380) and 10 (8-24) to 390 (230-520) and 19 (8-33) cells/10(6) lymphomonocytes, respectively, p=0.004 and p=0.008), whereas they remained unchanged in the other groups. The increase in EPC levels was not associated with the changes in the levels of the measured inflammatory and oxidative markers. CONCLUSION Short-term treatment with rosuvastatin, but not allopurinol, significantly increases the number of circulating EPCs in patients with heart failure providing further insights into its role in these individuals. The impact of rosuvastatin on EPCs is not mediated by changes in inflammatory and oxidative status.

Vascular endothelium and inflammatory process, in patients with combined Type 2 diabetes mellitus and coronary atherosclerosis: the effects of vitamin C.

Aims  Type 2 diabetes mellitus (DM) and coronary artery disease (CAD) are both associated with endothelial dysfunction and elevated oxidative and inflammatory state. We examined the effect of vitamin C on endothelial function and levels of soluble vascular cell adhesion molecule (sVCAM‐1), interleukin‐6 (IL‐6) and tumour necrosis factor (TNF‐α), in DM patients with or without CAD and in non‐diabetic subjects.