Gastric distension causes changes in heart rate and arterial blood pressure by affecting the crosstalk between vagal and splanchnic systems in anesthetised rats.

Various hindbrain nuclei have been demonstrated to be involved in the control of the cardiovascular reflexes elicited by both non-noxious and noxious gastric distension, through parasympathetic and sympathetic activation. The different role played by the branches of autonomic nervous system in exerting these effects and their crosstalk in relation to low-/high-pressure distension rate has not been examined yet. Therefore, in the present work, monolateral and bilateral vagotomy and splanchnicotomy were performed in anesthetised rats to analyse the involvement of hindbrain nuclei in haemodynamic changes caused by gastric distension at high (80 mmHg) and low (15 mmHg) pressure. The analysis of c-Fos expression in neuronal areas involved in cardiovascular control allowed us to examine their recruitment in response to various patterns of gastric distension and the crosstalk between vagal and splanchnic systems. The results obtained show that the low-pressure (non-noxious) gastric distension increases both heart rate and arterial blood pressure. In addition, the vagus nerve and hindbrain nuclei, such as nucleus ambiguous, ventrolateral medulla and lateral reticular nucleus, appear to be primarily involved in observed responses. In particular, we have found that although vagus nerve plays a central role in exerting those cardiovascular reflex changes at low gastric distension, for its functional expression an intact splanchnic system is mandatory. Hence, the absence of splanchnic input attenuates pressor responses or turns them into depressor responses. Instead at high-pressure (noxious) gastric distension, the splanchnic nerve represents the primary component in regulating the reflex cardiovascular effects.

Human Chorionic Gonadotropin Protects Vascular Endothelial Cells from Oxidative Stress by Apoptosis Inhibition, Cell Survival Signalling Activation and Mitochondrial Function Protection.

BACKGROUND/AIM: Previous reports have made it hypothetically possible that human chorionic gonadotropin (hCG) could protect against the onset of pregnancy-related pathological conditions by acting as an antioxidant. In the present study we planned to examine the effects of hCG against oxidative stress in human umbilical vein endothelial cells (HUVEC). METHODS: HUVEC were subjected to peroxidation by hydrogen peroxide. The modulation of nitric oxide (NO) release by hCG and its effects on cell viability, glutathione (GSH) levels, mitochondrial membrane potential and mitochondrial transition pore opening (MPTP) were examined by specific dyes. Endothelial and inducible NO synthase (eNOS and iNOS), Akt and extracellular -signal-regulated kinases 1/2 (ERK1/2) activation and markers of apoptosis were analyzed by Western Blot. RESULTS: In HUVEC, hCG reduced NO release by modulating eNOS and iNOS. Moreover, hCG protected HUVEC against oxidative stress by preventing GSH reduction and apoptosis, by maintaining Akt and ERK1/2 activation and by keeping mitochondrial function. CONCLUSION: The present results have for the first time shown protective effects exerted by hCG on vascular endothelial function, which would be achieved by modulation of NO release, antioxidant and antiapoptotic actions and activation of cell survival signalling. These findings could have clinical implications in the management of pregnancy-related disorders.

Uric acid: a potential biomarker of multiple sclerosis and of its disability.

BACKGROUND: Uric acid (UA) is a strong natural scavenger of reactive oxygen and nitrogen species, with evidence of possible use in the treatment of animal models of multiple sclerosis (MS). Consequently, serum UA has gained much attention as a possible biomarker of MS. We aim to investigate differences in serum UA levels between MS subjects and controls and evaluate possible relationships of UA with MS clinical features. METHODS: We recruited relapsing-remitting and secondary progressive MS subjects and healthy controls and measured their serum UA levels. We excluded subjects presenting concomitant conditions affecting UA levels. RESULTS: MS subjects (n=362) and controls (n=181) were recruited by propensity score matching (PSM). Statistical analyses were corrected for age, gender, and renal function. MS subjects presented significantly lower serum UA levels than controls (analysis of variance, p=0.014, adjusted r2=0.3036). Linear regression analysis showed a relationship between UA levels and disease duration (p<0.001, adjusted r2=0.3158, coefficient -0.00039), time from diagnosis (p<0.001, adjusted r2=0.3100, coefficient -0.0012), and Expanded Disability Status Scale (EDSS) (p<0.001, adjusted r2=0.3230, coefficient -0.1). CONCLUSIONS: Our findings support the importance of serum UA as a biomarker of MS disability and progression. Further studies with longitudinal design should be specifically designed to evaluate the importance of UA in the different stages of MS and in relation to distinct therapeutic strategies.

Effects of Artemetin on Nitric Oxide Release and Protection against Peroxidative Injuries in Porcine Coronary Artery Endothelial Cells.

Artemetin is one of the main components of Achillea millefolium L. and Artemisia absinthium, which have long been used for the treatment of various diseases. To date, however, available information about protective effects of their extracts on the cardiovascular system is scarce. Therefore, we planned to analyze the effects of artemetin on nitric oxide (NO) release and the protection exerted against oxidation in porcine aortic endothelial (PAE) cells. In PAE, we examined the modulation of NO release caused by artemetin and the involvement of muscarinic receptors, ß2-adrenoreceptors, estrogenic receptors (ER), protein-kinase A, phospholipase-C, endothelial-NO-synthase (eNOS), Akt, extracellular-signal-regulated kinases 1/2 (ERK1/2) and p38 mitogen activated protein kinase (p38 MAPK). Moreover, in cells treated with hydrogen peroxide, the effects of artemetin were examined on cell survival, glutathione (GSH) levels, apoptosis, mitochondrial membrane potential and transition pore opening. Artemetin increased eNOS-dependent NO production by the involvement of muscarinic receptors, ß2-adrenoreceptors, ER and all the aforementioned kinases. Furthermore, artemetin improved cell viability in PAE that were subjected to peroxidation by counteracting GSH depletion and apoptosis and through the modulation of mitochondrial function. In conclusion, artemetin protected endothelial function by acting as antioxidant and antiapoptotic agent and through the activation of ERK1/2 and Akt. Copyright © 2015 John Wiley & Sons, Ltd.

Protective effects elicited by levosimendan against liver ischemia/reperfusion injury in anesthetized rats.

As in other organs, oxidative stress-induced injury and cell death may result from free oxygen radical-dependent mechanisms and alterations in signal transduction pathways leading to apoptosis. Among the new suggested therapies for injuries caused by oxidative stress, the use of levosimendan has been reported to be quite promising. In the present study, we aimed to examine the protective effects of levosimendan against liver oxidative stress in anesthetized rats and to analyze the involvement of mitochondrial adenosine triphosphate-dependent potassium (mitoK(ATP)) channels and nitric oxide (NO). In 50 anesthetized rats, liver ischemia/reperfusion (I/R) was performed via nontraumatic portal occlusion. In some animals, levosimendan was infused into the portal vein at the onset of reperfusion, whereas other rats received the vehicle only. Moreover, in some rats, levosimendan was given after the intraportal administration of L-Nω-nitro-arginine methyl ester (L-NAME) or 5-hydroxydecanoate (5HD). The portal vein blood flow was measured, and blood samples were taken for the determination of transaminases, thiobarbituric acid reactive substances (TBARS), and reduced glutathione (GSH); liver biopsy samples were used for B cell lymphoma 2-associated X protein, caspase-9, Akt, and endothelial nitric oxide synthase (eNOS) activation through western blotting. Also, caspase-3 activity was measured. In rats, I/R caused an increase in apoptotic markers, transaminases, and TBARS and a decrease in GSH and Akt activation. Levosimendan administration was able to counteract oxidative damage and apoptosis in a dose-dependent way and to increase GSH, Akt, and eNOS activation. All effects of levosimendan were abolished by pretreatment with L-NAME and 5HD. In conclusion, the results of the present study show that levosimendan can exert protection against ischemic liver damage through mechanisms related to NO production and mitoKATP channel function. These data provide interesting perspectives into the use of levosimendan in hepatic surgery and transplantation.

1α,25-Dihydroxycholecalciferol (Vitamin D3) induces NO-dependent endothelial cell proliferation and migration in a three-dimensional matrix.

BACKGROUND/AIMS: The 1α,25-dihydroxycholecalciferol (Vit. D) induces eNOS dependent nitric oxide (NO) production in human umbilical vein endothelial cells (HUVEC). To our knowledge, there are no reports directly relating Vit. D induced NO production to proliferation and/or migration in endothelial cells (EC). The aim of this study was to evaluate whether Vit. D addition to porcine EC could affect their proliferation and/or migration in a three-dimensional matrix via NO production. MATERIALS AND METHODS: Porcine aortic endothelial cells (PAE) were used to evaluate Vit. D effects on cell proliferation and migration in a three-dimensional matrix. RESULTS: Vit. D induced NO production in PAE cells. Moreover, it induced a significant increase in cellular proliferation and migration in a three-dimensional matrix. These effects were NO dependent, as inhibiting eNOS activity by L-NAME PAE migration was abrogated. This effect was strictly related to MMP-2 expression and apparently dependent on Vit. D and NO production. CONCLUSIONS: Vit. D can promote both endothelial cells proliferation and migration in a three-dimensional matrix via NO-dependent mechanisms. These findings cast new light on the role of Vit. D in the angiogenic process, suggesting new applications for Vit. D in such fields as tissue repair and wound healing.

Intracoronary secretin increases cardiac perfusion and function in anaesthetized pigs through pathways involving ß-adrenoceptors and nitric oxide.

Secretin has been implicated in cardiovascular regulation through its specific receptors, as well as through ß-adrenoceptors and nitric oxide, although data on its direct effect on coronary blood flow and cardiac function have remained scarce. The present study aimed to determine the primary in vivo effect of secretin on cardiac function and perfusion and the mechanisms related to the autonomic nervous system, secretin receptors and NO. In addition, in coronary endothelial cells the intracellular pathways involved in the effects of secretin on NO release were also examined. In 30 pigs, intracoronary secretin infusion at 2.97 pg for each millilitre per minute of coronary blood flow at constant heart rate and aortic blood pressure increased coronary blood flow, maximal rate of change of left ventricular pressure, segmental shortening, cardiac output and coronary NO release (P<0.05). These responses were graded in a further five pigs. Moreover, while blockade of muscarinic cholinoreceptors (n=5) and of α-adrenoceptors (n=5) did not abolish the observed responses to secretin, blockade of ß1-adrenoceptors (n=5) prevented the effects of secretin on cardiac function. In addition, blockade of ß2-adrenoceptors (n=5) and NO synthase inhibition (n=5) prevented the coronary response and the effect of secretin on NO release. All these effects were abolished by a secretin receptor inhibitor (n=5). In coronary endothelial cells, the increased NO production caused by secretin was found to be related to cAMP/protein kinase A signalling activated as downstream effectors of stimulation of secretin receptors and ß2-adrenoceptors. In conclusion, in anaesthetized pigs secretin primarily increased cardiac function and perfusion through the involvement of specific receptors, ß-adrenoceptors and NO release.

Chronic cerebrospinal venous insufficiency in multiple sclerosis: a highly prevalent age-dependent phenomenon.

BACKGROUND: This study aimed to investigate the prevalence and clinical relevance of chronic cerebrospinal venous insufficiency (CCSVI) in multiple sclerosis (MS) patients and healthy controls using extra- and intracranial colour Doppler sonography. METHODS: We examined 146 MS patients, presenting with a clinically isolated syndrome, relapsing-remitting, secondary progressive, or primary progressive MS, and 38 healthy controls. Sonographic examination was performed according to Zamboni's protocol and was performed by three independent sonographers. The results of sonographic examination were compared with clinical and demographic characteristics of the patients. RESULTS: CCSVI, defined as the presence of at least two positive Zamboni's criteria, was found in 76% of MS patients and 16% of control subjects. B-mode anomalies of internal jugular veins, such as stenosis, malformed valves, annuli, and septa were the most common lesions detected in MS patients (80.8%) and controls (47.4%). We observed a positive correlation between sonographic diagnosis of CCSVI and the patients' age (p = 0.003). However, such a correlation was not found in controls (p = 0.635). Notably, no significant correlations were found between sonographic signs of CCSVI and clinical characteristics of MS, except for absent flow in the jugular veins, which was found more often in primary (p<0.005) and secondary (p<0.05) progressive patients compared with non-progressive patients. Absent flow in jugular veins was significantly correlated with patients' age (p < 0.0001). CONCLUSIONS: Sonographically defined CCSVI is common in MS patients. However, CCSVI appears to be primarily associated with the patient's age, and poorly correlated with the clinical course of the disease.

Levosimendan protection against kidney ischemia/reperfusion injuries in anesthetized pigs.

Ischemia/reperfusion (I/R) injury is an important cause of acute renal failure because of oxidative, inflammatory, and apoptotic mechanisms. The aim of the present study was to examine any possible protective effects of levosimendan in an in vivo pig model of renal I/R injury. In 40 anesthetized pigs (eight groups of five pigs each), I/R was induced by clamping-reopening the left renal artery. During ischemia, in three groups of pigs, levosimendan and the multiorgan preservation solution Custodiol, alone or in combination with levosimendan, were infused in the renal artery. In two other groups of animals, levosimendan in combination with Custodiol was administered after the intrarenal nitric-oxide (NO) synthase blocker N(ω)-nitro-L-arginine methyl ester (L-NAME) or the mitochondrial ATP-sensitive K(+) channel (K(ATP) channel) inhibitor 5-hydroxydecanoate (5-HD). In the other animals, saline, L-NAME, or 5-HD were administered alone. Throughout the experiments, urinary N-acetyl-ß-glucosaminidase (NAG) release was measured, and renal function was assessed. Moreover, renal biopsy samples were taken for the detection of apoptosis and tissue peroxidation. In pigs treated with levosimendan or the combination of levosimendan and Custodiol, NAG, peroxidation, and apoptotic markers were lower than in animals treated with Custodiol alone. In addition, renal function was better preserved, and cell survival and antioxidant systems were more activated. All beneficial effects were prevented by L-NAME and 5-HD. In conclusion, levosimendan alone or in combination with Custodiol exerted better protection against renal I/R injuries than Custodiol alone through antioxidant, antiapoptotic, and prosurvival actions depending on mitochondrial K(ATP) channels and NO-related mechanisms.

1α,25-dihydroxycholecalciferol induces nitric oxide production in cultured endothelial cells.

BACKGROUND: Recently, 1α,25-dihydroxycholecalciferol (vitD) has received increasing interest for its effects on many tissues and organs other than bone. A number of experimental studies have shown that vitD may have an important role in modifying risk for cardiovascular disease. AIMS: This study was planned to test the effects of vitD on endothelial nitric oxide (NO) production and to study the intracellular pathways leading to NO release. METHODS: In human umbilical vein endothelial cells (HUVEC) cultures the effects of vitD on NO production and p38, Akt, ERK and eNOS phosphorylations were examined in absence or in presence of the NO synthase inhibitor L-NAME and protein kinases specific inhibitors SB203580, wortmannin and UO126. RESULTS: VitD caused a concentration-dependent increase in NO production. The maximum effect was observed at a concentration of 1 nM and the optimal time of stimulation was 1 min. Effects induced by vitD were abolished by L-NAME and by pre-treatment with protein kinases inhibitors. To verify the effective involvement of vitD receptor (VDR) in the action mechanism of vitD, experiments were repeated in presence of the specific VDR ligands ZK159222 and ZK191784. CONCLUSIONS: The results of this study demonstrate that vitD can induce a significant increase in endothelial NO production. VitD interaction with VDR caused the phosphorylation of p38, AKT and ERK leading to eNOS activation.

Levosimendan modulates programmed forms of cell death through K(ATP) channels and nitric oxide.

Levosimendan exerts cardioprotection through mitochondrial K(ATP) (mitoK(ATP)) channels opening. In addition, intracoronary levosimendan was found to modulate programmed forms of cell death by nitric oxide (NO) involvement. The aim of this study was to examine the role of mitoK(ATP) channels and NO in the effects of levosimendan on apoptosis/autophagy. In H9c2 cells treated with hydrogen peroxide apoptosis/autophagy, survival signaling, cell viability, mitochondrial membrane potential, and permeability transition pore opening were analyzed through Western blot and colorimetric and fluorescence assays. Pretreatment of H9c2 cells with levosimendan was able to counteract the oxidative injuries caused by hydrogen peroxide. The effects of levosimendan were potentiated by diazoxide and were similar to those elicited by the autophagic activator rapamycin. The autophagic inhibitor 3-methyladenine reduced the effects of levosimendan, whereas after the pan-caspases inhibitor N-Acetyl-Asp-Glu-Val-Asp-al (Z-VAD.FMK), cell survival and autophagy in response to levosimendan increased. Both the mitoK(ATP) channels inhibition and the NO synthase blocking attenuated the cardioprotection elicited by levosimendan. The results have shown that levosimendan protects H9c2 cells against oxidative injuries through the modulation of the interplay between autophagy and apoptosis and the activation of survival signaling. The mitoK(ATP) channels and NO may be involved in such cardioprotection through interference with mitochondrial functioning.

Management of acute cardiac failure by intracoronary administration of levosimendan.

Acute cardiac failure caused by myocardial infarction or inadequate cardioprotection during heart surgery is associated with increased mortality and morbidity. Levosimendan is a new drug used in heart failure though it is limited by the systemic hypotension, which develops with intravenous administration. Intracoronary (IC) administration however should affect systemic circulation less while maintaining the beneficial cardiac effects of the drug. We herewith report the results from the first such clinical series. Levosimendan was administered IC in 33 consecutive patients who developed cardiogenic shock during heart surgery and were unable to wean off cardiopulmonary bypass despite maximal support. Preadministration/postadministration coronary graft flows, hemodynamic parameters, left ventricular function, and metabolic requirements were measured and compared. Levosimendan significantly increased graft flows and improved hemodynamic parameters. Systolic blood pressure (93 ± 26.4 vs. 106 ± 18.2 mm Hg, P < 0.05) and cardiac index (2.0 ± 0.5 vs. 3.1 ± 0.2, P < 0.001) were increased, whereas systemic vascular resistance (1470.7 ± 114 vs. 1195.8 ± 112, P < 0.01) was reduced. Better myocardial perfusion improved metabolic requirements, with myocardial oxygen extraction and glucose uptake increasing by 72% and 74%, respectively, whereas lactate production was reduced by 64%. Echocardiography demonstrated additional ventricular segment recruitment. Therefore, IC Levosimendan administration in acute heart failure is safe and efficacious producing improved cardiac function without significant detrimental hypotension.

Intracoronary melatonin increases coronary blood flow and cardiac function through ß-adrenoreceptors, MT1/MT2 receptors, and nitric oxide in anesthetized pigs.

Melatonin is involved in the regulation of the cardiovascular system through the modulation of sympathetic function and the nitric oxide (NO)-related pathway and interaction with MT1/MT2 receptors. However, information regarding its direct actions on coronary blood flow and cardiac function is scarce. This study therefore determined the primary in vivo effect of melatonin on cardiac function and perfusion and the involvement of the autonomic nervous system, MT1/MT2 receptors, and NO. In 35 pigs, melatonin infused into the coronary artery at 70 pg for each mL/min of coronary blood flow while preventing changes in heart rate and arterial pressure increased coronary blood flow, dP/dt(max), segmental shortening, and cardiac output by about 12%, 14%, 8%, and 23% of control values (P < 0.05), respectively. These effects were accompanied by an increase in coronary NO release of about 46% (P < 0.05) of control values. The aforementioned responses were graded in a further five pigs. Moreover, the blockade of muscarinic cholinoreceptors (n = 5) and α-adrenoreceptors (n = 5) did not abolish the observed responses to melatonin. After ß(1)-adrenoreceptors blocking (n = 5), melatonin failed to affect cardiac function, whereas ß(2)-adrenoreceptors (n = 5) and NO synthase inhibition (n = 5) prevented the coronary response and the effect of melatonin on NO release. Finally, all effects were prevented by MT1/MT2 receptor inhibitors (n = 10). In conclusion, melatonin primarily increased coronary blood flow and cardiac function through the involvement of MT1/MT2 receptors, ß-adrenoreceptors, and NO release. These findings add new information about the mechanisms through which melatonin physiologically modulates cardiovascular function and exerts cardioprotective effects.

ProspeCtive study to evaluate efficacy, safety and tOlerability of dietary supplemeNT of Curcumin (BCM95) in subjects with Active relapsing MultIple Sclerosis treated with subcutaNeous Interferon beta 1a 44 mcg TIW (CONTAIN): A randomized, controlled trial.

BACKGROUND: multiple sclerosis (MS) is a complex disease sustained by several pathogenic mechanisms. As such, combination therapy strategies, targeting a range of disease mechanisms, might represent the ideal therapeutic approach. Here we investigated the efficacy of curcumin, a naturally occurring poly-phenolic phytochemical with potent anti-inflammatory and antioxidant properties, in subjects under treatment with IFN ß-1a, to test the effects of this combination therapy on clinical and MRI parameters of inflammation and neurodegeneration in relapsing MS (RMS). METHODS: eighty active RMS were prospectively enrolled, randomized (1:1) to either the IFN-curcumin or the IFN-placebo group and followed up longitudinally with clinical and MRI assessments for 24 months. Primary endpoint was the efficacy of curcumin versus placebo as add-on therapy on new/enlarging T2 lesions in RMS subjects under treatment with subcutaneous IFN ß-1a 44 mcg TIW. Efficacy on clinical parameters (relapses and disability progression), other MRI parameters of inflammation (T1 Gd-enhancing lesions, combined unique active-CUA lesions) and neurodegeneration (T1-hypointense lesions, grey matter loss and white matter microstructural damage) as well as safety and tolerability of curcumin were explored as secondary endpoints. RESULTS: ten subjects dropped out from the study by month 12 (6 in the IFN-curcumin group and 4 in the IFN-placebo group), and 27 by month 24 (11 in the IFN-curcumin group and 16 in the IFN-placebo group). Although no between-group difference was present in terms of proportion of subjects free from new/enlarging T2 lesions, a lower proportion of patients with CUA lesions was noted at month 12 in the IFN-curcumin group in comparison with the IFN-placebo group (7.5% vs 17.5%, χ² test p= 0.0167). This result was not confirmed at month 24. The statistical analysis failed to reveal any difference between the two treatment groups - IFN-curcumin and IFN-placebo - in terms of relapses, disability progression, other MRI metrics of inflammation and MRI changes suggestive of ongoing neurodegeneration. No difference in the rate and nature of adverse events was observed between the two treatment groups. CONCLUSION: Although the study drop-out rate was too high to allow definite conclusions, our findings suggest that curcumin might add to IFN ß-1a efficacy on radiological signs of inflammation in MS, while it did not seem to exert any neuroprotective effect as assessed by clinical and MRI parameters. (NCT01514370).

Modulation of calcium movements by urocortin II in endothelial cells.

BACKGROUND: In endothelial cells urocortin II has recently been found to activate nitric oxide synthase through cAMP-dependent and Ca(2+)-related pathway. AIM: The present study was therefore planned to determine the mechanisms of urocortin II effect on Ca(2+) movements. METHODS: In Fura-2 loaded porcine aortic endothelial cells (PAE), the effects of urocortin II on [Ca(2+)]c were analyzed and compared with those of various K(+) channels agonists/antagonists. RESULTS: In Fura-2 loaded PAE, urocortin II promoted a transient increase of [Ca(2+)]c mainly originating from an intracellular pool sensitive to thapsigargin and slightly from the extracellular space. In addition, urocortin II caused the hyperpolarization of plasma membrane through the opening of K(+) channels, which contributed to the increased [Ca(2+)]c. These effects were abolished by the corticotropin releasing factor receptors (CRFR2) blocker, the adenylyl cyclase and Ca(2+)-calmodulin-kinase (CaMKII) inhibitors and by blockers of K(+) channels. In addition, in PAE cultured in Na(+)-free medium or loaded with the plasma-membrane Ca(2+) pump inhibitor the urocortin II-evoked Ca(2+) transient was slower. CONCLUSION: The results obtained show that urocortin II affects intracellular Ca(2+) homeostasis in PAE by both promoting a discharge of intracellular pool and by interfering with the operation of store-dependent channels through CRFR2-cAMP-CaMKII related signalling and K(+) channels opening.

Atorvastatin combined to interferon to verify the efficacy (ACTIVE) in relapsing-remitting active multiple sclerosis patients: a longitudinal controlled trial of combination therapy.

A large body of evidence suggests that, besides their cholesterol-lowering effect, statins exert anti-inflammatory action. Consequently, statins may have therapeutic potential in immune-mediated disorders such as multiple sclerosis. Our objectives were to determine safety, tolerability and efficacy of low-dose atorvastatin plus high-dose interferon beta-1a in multiple sclerosis patients responding poorly to interferon beta-1a alone. Relapsing-remitting multiple sclerosis patients, aged 18-50 years, with contrast-enhanced lesions or relapses while on therapy with interferon beta-1a 44 microg (three times weekly) for 12 months, were randomized to combination therapy (interferon + atorvastatin 20 mg per day; group A) or interferon alone (group B) for 24 months. Patients underwent blood analysis and clinical assessment with the Expanded Disability Status Scale every 3 months, and brain gadolinium-enhanced magnetic resonance imaging at screening, and 12 and 24 months thereafter. Primary outcome measure was contrast-enhanced lesion number. Secondary outcome measures were number of relapses, EDSS variation and safety laboratory data. Forty-five patients were randomized to group A (n = 21) or B (n = 24). At 24 months, group A had significantly fewer contrast-enhanced lesions versus baseline (p = 0.007) and significantly fewer relapses versus the two pre-randomization years (p < 0.001). At survival analysis, the risk for a 1-point EDSS increase was slightly higher in group B than in group A (p = 0.053). Low-dose atorvastatin may be beneficial, as add-on therapy, in poor responders to high-dose interferon beta-1a alone.

Modulation of programmed forms of cell death by intracoronary levosimendan during regional myocardial ischemia in anesthetized pigs.

PURPOSE: Powerful mediators of programmed cell death, such as apoptosis and autophagy, can contribute to myocyte cell loss during pathological cardiac conditions. Levosimendan has been shown to exert beneficial hemodynamic effects in presence of global myocardial ischemia and heart failure through vasodilatation and increase of cardiac contractility. Recently, the intracoronary administration of a bolus levosimendan was found to exert favourable cardiac anti-stunning effects without lowering arterial pressure, which limits the use of levosimendan mainly in coronary artery disease. Here we tested whether the intracoronary administration of levosimendan can beneficially modulate programmed cell death in acute regional myocardial ischemia. METHODS: Acute regional myocardial ischemia was induced in 20 anaesthetized pigs and intracoronary levosimendan 15 min bolus administration was started 4 h afterwards. The effects of levosimendan on coronary blood flow and cardiac function were evaluated and myocardial biopsies were examined for criteria of autophagy and apoptosis. RESULTS: The administration of levosimendan caused a significant increase of coronary blood flow (p < 0.05) in absence of changes in cardiac function. Moreover, levosimendan prevented the down-regulation of the anti-apoptotic gene, Bcl-2, and the up-regulation of the apoptotic markers Bax and cytochrome c, which resulted in a reduced expression of TUNEL fragmented nuclei (p < 0.05). Furthermore, levosimendan maintained Beclin 1 at 4 h and potentiated LC3 II expression, these results being consistent with autophagy activation. CONCLUSIONS: Such effects of intracoronary levosimendan bolus administration during regional myocardial ischemia indicate the occurrence of cardio-protection by modulation of programmed form of cell death.

Urocortin II induces nitric oxide production through cAMP and Ca2+ related pathways in endothelial cells.

BACKGROUND: Urocortin II has previously been shown in anesthetized pigs to increase coronary blood flow through activation of the endothelial nitric oxide synthase (eNOS) pathway and involvement of the subtype 2 of corticotropin releasing factor receptors (CRFR2). However, little information has been available regarding the intracellular signalling through these receptors and leading to the release of nitric oxide (NO). AIM: The present study was therefore planned to determine the mechanism involved in such signalling. METHODS: In porcine aortic endothelial cells (PAE) the effects of urocortin II on NO production and ERK, Akt, p38 and eNOS phosphorylation were examined in absence or presence of the adenylyl cyclase agonist forskolin and antagonist 2'5' dideoxyadenosine, the Ca2+ ionophore A23187, the Ca2+-calmodulin-kinase inhibitor KN93, the CRFR2 blocker astressin 2B and of the protein kinases specific inhibitors UO126, wortmannin and SB203580. RESULTS: Urocortin II caused a significant increase of NO production, which was amplified by forskolin and A23187 (P <0.05). All effects of urocortin II were abolished by l-NAME, 2'5' dideoxyadenosine, KN93, astressin 2B and by pre-treatment of cells with UO126, wortmannin and SB203580. Western Blot analysis confirmed the involvement of ERK, Akt and p38 in the eNOS activation. CONCLUSION: In PAE urocortin II interaction with CRFR2 caused a cAMP-dependent and Ca2+-related phoshorylation of ERK, Akt and p38 leading to eNOS activation.

The Impact of a New "Inverted Arch" Prosthetic Annuloplasty Ring on the Mitral Valve's 3-D Motion: An Experimental Ex-Vivo Study.

This experimental study aimed to evaluate the ex-vivo three-dimensional (3-D) motion of the Inverted Arch Ring (IAR), an innovative new design concept for a flexible incomplete annuloplasty prosthesis with an incorporated stabilizing rigid arch that can be used in correcting mitral valve regurgitation. Twenty explanted porcine hearts were placed in a circulation simulation system. Ultrasonometry transducers implanted in the mitral annulus were used to measure the 3-D valvular motion during a simulated cardiac cycle. Annular distance measurements were recorded and compared in each heart before and after the implantation of the IAR prosthesis at pressures corresponding to mid-systole and mid-diastole. Distances measured in mid-systole and mid-diastole demonstrated no significant differences in annular motion or in valve areas either prior to or after IAR implantation. Therefore, the results of this study confirm the minimal effects exerted by the IAR prosthesis on the mitral valve's 3-D motion during a simulated cardiac cycle.