Levamisole Impairs Vascular Function by Blocking α-Adrenergic Receptors and Reducing NO Bioavailability in Rabbit Renal Artery.

Levamisole is an anthelmintic drug restricted to veterinary use but is currently detected as the most widely used cocaine cutting agent in European countries. Levamisole-adulterated cocaine has been linked to acute kidney injury, marked by a decrease in glomerular filtration rate, which involves reduced renal blood flow, but data on the alteration of renovascular response produced by levamisole are scarce. Renal arteries were isolated from healthy rabbits and used for isometric tension recording in organ baths and protein analysis. We provide evidence that depending on its concentration, levamisole modulates renovascular tone by acting as a non-selective α-adrenergic receptor blocker and down-regulates α1-adrenoceptor expression. Furthermore, levamisole impairs the endothelium-dependent relaxation induced by acetylcholine without modifying endothelial nitric oxide synthase (eNOS) expression. However, exposure to superoxide dismutase (SOD) partially prevents the impairment of ACh-induced relaxation by levamisole. This response is consistent with a down-regulation of SOD1 and an up-regulation of NADPH oxidase 4 (Nox4), suggesting that endothelial NO loss is due to increased local oxidative stress. Our findings demonstrate that levamisole can interfere with renal blood flow and the coordinated response to a vasodilator stimulus, which could worsen the deleterious consequences of cocaine use.

Sodium valproate treatment reverses endothelial dysfunction in aorta from rabbits with acute myocardial infarction.

Sodium valproate (VPA), a histone deacetylase (HDAC) inhibitor, could be a promising candidate to treat acute myocardial infarction (AMI). In this study, AMI was induced in New Zealand White rabbits by occluding the left circumflex coronary artery for 1 h, followed by reperfusion. The animals were distributed into three experimental groups: the sham-operated group (SHAM), the AMI group and the AMI + VPA group (AMI treated with VPA 500 mg/kg/day). After 5 weeks, abdominal aorta was removed and used for isometric recording of tension in organ baths or protein expression by Western blot, and plasma for the determination of nitrate/nitrite (NOx) levels by colorimetric assay. Our results indicated that AMI induced a reduction of the endothelium-dependent response to acetylcholine without modifying the endothelium-independent response to sodium nitroprusside, leading to endothelial dysfunction. VPA treatment reversed AMI-induced endothelial dysfunction and even increased NO sensitivity in vascular smooth muscle. This response was consistent with an antioxidant effect of VPA, as it was able to reverse the superoxide dismutase 1 (SOD 1) down-regulation induced by AMI. Our experiments also ruled out that the VPA mechanism was related to eNOS, iNOS, sGC and arginase expression or changes in NOx plasma levels. Therefore, we conclude that VPA improves vasodilation by increasing NO bioavailability, likely due to its antioxidant effect. Since endothelial dysfunction was closely related to AMI, VPA treatment could increase aortic blood flow, making it a potential agent in reperfusion therapy that can prevent the vascular damage.

Improvement of Vascular Insulin Sensitivity by Ranolazine.

Ranolazine (RN) is a drug used in the treatment of chronic coronary ischemia. Different clinical trials have shown that RN behaves as an anti-diabetic drug by lowering blood glucose and glycosylated hemoglobin (HbA1c) levels. However, RN has not been shown to improve insulin (IN) sensitivity. Our study investigates the possible facilitating effects of RN on the actions of IN in the rabbit aorta. IN induced vasodilation of the abdominal aorta in a concentration-dependent manner, and this dilatory effect was due to the phosphorylation of endothelial nitric oxide synthase (eNOS) and the formation of nitric oxide (NO). On the other hand, IN facilitated the vasodilator effects of acetylcholine but not the vasodilation induced by sodium nitroprusside. RN facilitated all the vasodilatory effects of IN. In addition, IN decreased the vasoconstrictor effects of adrenergic nerve stimulation and exogenous noradrenaline. Both effects were in turn facilitated by RN. The joint effect of RN with IN induced a significant increase in the ratio of p-eNOS/eNOS and pAKT/AKT. In conclusion, RN facilitated the vasodilator effects of IN, both direct and induced, on the adrenergic system. Therefore, RN increases vascular sensitivity to IN, thus decreasing tissue resistance to the hormone, a key mechanism in the development of type II diabetes.

Protective action of ultrasound-guided electrolysis technique on the muscle damage induced by notexin in rats.

It is known that exercise can be one of the causes of muscular damage. In recent times, physiotherapists and medical professionals have been employing USGET techniques to stimulate muscle recovery to improve its performance after the injury. We pretend to analyse if the Ultrasound-guided electrolysis (USGET) technique could reduce muscle damage, inflammation, and pain in the present study. Female Wistar rats were assigned to one of three different groups: control (C), notexin (NOT) and notexin with USGET (electrolysis at 6mA) (NOT+USGET). We used the USGT technique, based on electrical stimulation with a continuous current of 4 pulses at an intensity of 6 mA for 5 seconds, conveyed to the muscle. The response was tested with motor function tests. In these tests, we could observe an increase in time and foot faults when crossing a beam in the NOT group compared to C group rats. On the other hand, a significant decrease in both variables was detected in the NOT+USGET compared to the NOT group. Muscle power was measured with a grip strength test, obtaining far better performances in NOT+USGET rats when compared to NOT rats. Moreover, the USGET technique prevented the increase of pro-inflammatory proteins IL-6 and chemokines CCL3 (Chemokine (C-C motif) ligand 3), CCL4 (Chemokine (C-C motif) ligand 4), and CCL5 (Chemokine (C-C motif) ligand 5) with their receptor CCR5 (C-C chemokine receptor type 5), induced by notexin in the quadriceps. At the same time, the study evidenced a decrease in both CCR8 (C-C chemokine receptor type 5,) and NF-ᴋB (nuclear factor- ᴋB) expressions after USGET treatment. On the other hand, we obtained evidence that demonstrated anti-inflammatory properties of the USGET technique, thus being the increase in IL-10 (Interleukin 10) and IL-13 (Interleukin 13) in the NOT+USGET group compared to the NOT group. Furthermore, when applying NSGET after damage, an increase in anti-inflammatory mediators and reduction of pro-inflammatory mediators, which, overall, promoted muscle regeneration, was observed. These results support the idea that the NSGET technique improves muscle recovery after toxic damages, which would justify its employment.

Facilitation of Insulin Effects by Ranolazine in Astrocytes in Primary Culture.

Ranolazine (Rn) is a drug used to treat persistent chronic coronary ischemia. It has also been shown to have therapeutic benefits on the central nervous system and an anti-diabetic effect by lowering blood glucose levels; however, no effects of Rn on cellular sensitivity to insulin (Ins) have been demonstrated yet. The present study aimed to investigate the permissive effects of Rn on the actions of Ins in astrocytes in primary culture. Ins (10-8 M), Rn (10-6 M), and Ins + Rn (10-8 M and 10-6 M, respectively) were added to astrocytes for 24 h. In comparison to control cells, Rn and/or Ins caused modifications in cell viability and proliferation. Rn increased protein expression of Cu/Zn-SOD and the pro-inflammatory protein COX-2 was upregulated by Ins. On the contrary, no significant changes were found in the protein expression of NF-κB and IκB. The presence of Rn produced an increase in p-ERK protein and a significant decrease in COX-2 protein expression. Furthermore, Rn significantly increased the effects of Ins on the expression of p-AKT, p-eNOS, p-ERK, Mn-SOD, and PPAR-γ. In addition, Rn + Ins produced a significant decrease in COX-2 expression. In conclusion, Rn facilitated the effects of insulin on the p-AKT, p-eNOS, p-ERK, Mn-SOD, and PPAR-γ signaling pathways, as well as on the anti-inflammatory and antioxidant effects of the hormone.

Deleterious effects of levamisole, a cocaine adulterant, in rabbit aorta.

Levamisole, a veterinary anthelmintic drug, is one of the most widely used and dangerous cocaine adulterants. Like cocaine, levamisole acutely blocks noradrenaline reuptake but with much less potency, although its vascular effects are not well known. In this study, we evaluated the vascular effects of levamisole and cocaine in rabbit aortic rings used for isometric recording of tension in organ baths and protein expression by western blot. Our results indicated that levamisole (10-5-10-3 M) induced a concentration-dependent relaxation in rings precontracted with noradrenaline (10-7-3 × 10-7 M). Furthermore, it reduced the contractile response to phenylephrine (10-9-3 × 10-5 M) that was not modified by cocaine (10-5-10-4 M), and reduced α1-adrenergic receptor expression. Levamisole (10-6-10-4 M) produced a potentiation of the electrical field stimulation that was not further enhanced by the combination of both drugs. However, high concentrations of levamisole (10-3 M) abolished adrenergic neurotransmission whether administered alone or with cocaine (10-4 M). In addition, levamisole (10-5-10-3 M) also decreased endothelium-dependent relaxation to acetylcholine that was not further impaired by cocaine (10-4 M), and that was partially reversed by superoxide dismutase (SOD, 200 U/ml). These results demonstrate that levamisole has a dual effect on the adrenergic system, and its effects are independent of the presence of cocaine. At lower concentrations, it enhances the contractile sympathetic response by blocking presynaptic α2-adrenergic receptors, while at high concentrations, the effect of the antagonism of α1-adrenergic receptor prevails. In addition, levamisole induces endothelial dysfunction by reducing NO bioavailability, and this effect could be in part mediated by oxidative stress.

PPARγ as an indicator of vascular function in an experimental model of metabolic syndrome in rabbits.

BACKGROUND AND AIMS: Underlying mechanisms associated with vascular dysfunction in metabolic syndrome (MetS) remain unclear and can even vary from one vascular bed to another. METHODS: In this study, MetS was induced by a high-fat, high-sucrose diet, and after 28 weeks, aorta and renal arteries were removed and used for isometric recording of tension in organ baths, protein expression by Western blot, and histological analysis to assess the presence of atherosclerosis. RESULTS: MetS induced a mild hypertension, pre-diabetes, central obesity and dyslipidaemia. Our results indicated that MetS did not change the contractile response in either the aorta or renal artery. Conversely, vasodilation was affected in both arteries in a different way. The aorta from MetS showed vascular dysfunction, including lower response to acetylcholine and sodium nitroprusside, while the renal artery from MetS presented a preserved relaxation to acetylcholine and an increased sensitivity to sodium nitroprusside. We did not find vascular oxidative stress in the aorta from MetS, but we found a significant decrease in PPARγ, phospho-Akt (p-Akt) and phospho-eNOS (p-eNOS) protein expression. On the other hand, we found oxidative stress in the renal artery from MetS, and PPARγ, Akt and p-Akt were overexpressed. No evidence of atherosclerosis was found in arteries from MetS. CONCLUSIONS: MetS affects vascular function differently depending on the vessel. In the aorta, it decreases both the vasodilation and the expression of the PPARγ/Akt/eNOS pathway, while in the renal artery, it increases the expression of PPARγ/Akt signalling pathway without decreasing the vasodilation.

Inflammatory Chemokines Expression Variations and Their Receptors in APP/PS1 Mice.

BACKGROUND: In Alzheimer's disease (AD), an increase in inflammation is distinctive. Amyloid precursor protein plus presenilin-1 (APP/PS1 mice) is a model for this illness. Chemokines secreted by central nervous system (CNS) cells could play multiple important roles in AD. Data looking for the chemokines involved in inflammatory mechanisms are lacking. To understand the changes that occur in the inflammation process in AD, it is necessary to improve strategies to act on specific inflammatory targets. OBJECTIVE: Chemokines and their receptors involved in phagocytosis, demyelination, chemotaxis, and coagulation were the objective of our study. METHODS: Female APPswe/PS1 double-transgenic mice (B6C3-Tg) were used and cortex brain from 20-22-month-old mice obtained and used to quantify chemokines and chemokine receptors expression using RT-PCR technique. RESULTS: Significant inflammatory changes were detected in APP/PS1 compared to wild type mice. CCR1, CCR3, CCR4, and CCR9 were elevated, and CCR2 were decreased compared with wild type mice. Their ligands CCL7, CCL11, CCL17, CCL22, CCL25, and CXCL4 showed an increase expression; however, changes were not observed in CCL2 in APP/PS1 compared to wild type mice. CONCLUSION: This change in expression could explain the differences between AD patients and elderly people without this illness. This would provide a new strategy for the treatment of AD, with the possibility to act in specific inflammatory targets.

Action of low doses of Aspirin in Inflammation and Oxidative Stress induced by aß1-42 on Astrocytes in primary culture.

Aspirin has been used as anti-inflammatory and anti-aggregate for decades but the precise mechanism(s) of action after the presence of the toxic peptide Aß1-42 in cultured astrocytes remains poorly resolved. Here we use low-doses of aspirin (10-7 M) in astrocytes in primary culture in presence or absence of Aß1-42 toxic peptide. We noted an increase of cell viability and proliferation with or without Aß1-42 peptide presence in aspirin treated cells. In addition, a decrease in apoptosis, determined by Caspase 3 activity and the expression of Cyt c and Smac/Diablo, were detected. Also, aspirin diminished necrosis process (LDH levels), pro-inflammatory mediators (IL-ß and TNF-α) and NF-ᴋB protein expression, increasing anti-inflammatory PPAR-γ protein expression, preventing Aß1-42 toxic effects. Aspirin inhibited COX-2 and iNOS without changes in COX-1 expression, increasing anti-oxidant protein (Cu/Zn-SOD and Mn-SOD) expression in presence or absence of Aß1-42. Taken together, our results show that aspirin, at low doses increases cell viability by decreasing inflammation and oxidative stress, preventing the deleterious effects of the Aß1-42 peptide on astrocytes in primary culture. The use of low doses of aspirin may be more suitable for Alzheimer's disease.

Relaxant and antiadrenergic effects of ranolazine in human saphenous vein.

OBJECTIVES: Ranolazine improves vascular function in animal models. We evaluate the effects of ranolazine on vascular function and adrenergic response in human saphenous vein. METHODS: Rings from 53 patients undergoing coronary artery bypass grafting were mounted in organ baths. Concentration-response curves to ranolazine were constructed in rings precontracted with phenylephrine, endothelin-1, vasopressin, KCl and the thromboxane A2 analogue U-46619. In rings precontracted with phenylephrine, relaxation to ranolazine was tested in the absence and presence of endothelial factors inhibitors, K+ channel blockers and verapamil. The effects of ranolazine on frequency-response and concentration-response curves to phenylephrine were performed in the absence and presence of endothelial factors inhibitors and K+ channel blockers. Endothelial nitric oxide synthase, α1 adrenergic receptor and large conductance Ca2+-activated K+ channel protein expressions were measured by Western blotting. RESULTS: Ranolazine (10-9-10-4 M) produced a concentration-dependent relaxation only in rings precontracted with phenylephrine that was reduced by endothelial denudation, NG-nitro-l-arginine methyl ester (10-4 M), charybdotoxin (10-7 M) and verapamil (10-6 M). Ranolazine diminished adrenergic contractions induced by electrical field stimulation (2-4 Hz) and phenylephrine (10-9-10-5 M) that were prevented by tetraethylammonium (10-3 M) and charybdotoxin (10-7 M). Ranolazine significantly decreased α1 adrenergic receptor and increased large conductance Ca2+-activated K+ channel protein expression in the saphenous vein. CONCLUSIONS: Ranolazine diminishes the adrenergic vasoconstriction, acting as α1 antagonist, and by increasing large conductance Ca2+-activated K+ channel involvement. The relaxant effects of ranolazine are partially mediated by endothelial nitric oxide, large conductance Ca2+-activated K+ channels and the blockade of voltage-dependent Ca2+ channels.

The Role of Chemokines in Alzheimer's Disease.

OBJECTIVE: The most common multifactorial neurodegenerative disorder occurring in old age is Alzheimer's disease. The neuropathological hallmarks of that disorder are amyloid plaques with the presence of ß -amyloid aggregates, intraneuronal tau protein tangles, and chronic inflammation. Brain cells such as microglia and astrocytes are inflammatory cells associated with Alzheimer's disease and involved in the production of inflammatory mediators, such as cytokines and chemokines. Chemokines consist of a large family of protein mediators with low molecular weight, which able to control the migration and residence of all immune cells. In pathological conditions, such as Alzheimer's disease, chemokines contribute to the inflammatory response by recruiting T cells and controlling microglia/ macrophages activation. METHODS: The present study focuses on the role that chemokines and their receptors play in Alzheimer's disease and in processes such as inflammation and oxidative stress. RESULTS: Chemokines are important mediators in AD and inflammation. They promote Aß deposition and TAU hyperphosphorylation aggravating and increasing the progression of AD. Moreover, they affect the processing of senile plaques and produce abnormal TAU phosphorylation. CONCLUSION: There is no cure for AD but the therapeutic potential of chemokines to control the development of the disease may be a field of study to consider in the future.

Function of Glia in Aging and the Brain Diseases.

Microglia cells during aging, neurodegeneration and neuroinflammation show different morphological and transcriptional profiles (related to axonal direction and cell adhesion). Furthermore, expressions of the receptors on the surface and actin formation compared to young are also different. This review delves into the role of glia during aging and the development of the diseases. The susceptibility of different regions of the brain to disease are linked to the overstimulation of signals related to the immune system during aging, as well as the damaging impact of these cascades on the functionality of different populations of microglia present in each region of the brain. Furthermore, a decrease in microglial phagocytosis has been related to many diseases and also has been detected during aging. In this paper we also describe the role of glia in different illness, such as AD, ALS, pain related disorders, cancer, developmental disorders and the problems produced by opening of the blood brain barrier. Future studies will clarify many points planted by this review.

Targeting Early Atherosclerosis: A Focus on Oxidative Stress and Inflammation.

Atherosclerosis is a chronic vascular inflammatory disease associated to oxidative stress and endothelial dysfunction. Oxidation of low-density lipoprotein (LDL) cholesterol is one of the key factors for the development of atherosclerosis. Nonoxidized LDL have a low affinity for macrophages, so they are not themselves a risk factor. However, lowering LDL levels is a common clinical practice to reduce oxidation and the risk of major events in patients with cardiovascular diseases (CVD). Atherosclerosis starts with dysfunctional changes in the endothelium induced by disturbed shear stress which can lead to endothelial and platelet activation, adhesion of monocytes on the activated endothelium, and differentiation into proinflammatory macrophages, which increase the uptake of oxidized LDL (oxLDL) and turn into foam cells, exacerbating the inflammatory signalling. The atherosclerotic process is accelerated by a myriad of factors, such as the release of inflammatory chemokines and cytokines, the generation of reactive oxygen species (ROS), growth factors, and the proliferation of vascular smooth muscle cells. Inflammation and immunity are key factors for the development and complications of atherosclerosis, and therefore, the whole atherosclerotic process is a target for diagnosis and treatment. In this review, we focus on early stages of the disease and we address both biomarkers and therapeutic approaches currently available and under research.

Changes in Chemokines and Chemokine Receptors Expression in a Mouse Model of Alzheimer's Disease.

The amyloid precursor protein plus presenilin-1 (APP/PS1) mice are a frequently-used model for Alzheimer's disease studies (AD). However, the data relevant to which proteins are involved in inflammatory mechanism are not sufficiently well-studied using the AD mouse model. Using behavioral studies, quantitative RT-PCR and Western-blot techniques, significant findings were determined by the expression of proteins involved in inflammation comparing APP/PS1 and Wild type mice. Increased GFAP expression could be associated with the elevation in number of reactive astrocytes. IL-3 is involved in inflammation and ABDF1 intervenes normally in the transport across cell membranes and both were found up-regulated in APP/PS1 mice compared to Wild type mice. Furthermore, CCR5 expression was decreased and both CCL3 and CCL4 chemokines were highly expressed indicating a possible gliosis and probably an increase in chemotaxis from lymphocytes and T cell generation. We also noted for the first time, a CCR8 increase expression with diminution of its CCL1 chemokine, both normally involved in protection from bacterial infection and demyelination. Control of inflammatory proteins will be the next step in understanding the progression of AD and also in determining the mechanisms that can develop in this disease.

Chronic exercise impairs nitric oxide pathway in rabbit carotid and femoral arteries.

KEY POINTS: Some of the beneficial effects of exercise in preventing vascular related diseases are mediated by the enhancement of endothelial function where the role of nitric oxide (NO) is well documented, although the relevance of calcium activated potassium channels is not fully understood. The impact of oxidative stress induced by training on endothelial function remains to be clarified. By evaluating different endothelial vasodilator pathways on two vascular beds in a rabbit model of chronic exercise, we found a decreased NO bioavailability and endothelial nitric oxide synthase expression in both carotid and femoral arteries. Physical training induced carotid endothelial dysfunction as a result of an increase in oxidative stress and a reduction in superoxide dismutase expression. In the femoral artery, the lower production of NO was counteracted by an increased participation of large conductance calcium activated potassium channels, preventing endothelial dysfunction. ABSTRACT: The present study aimed to evaluate the effects of chronic exercise on vasodilator response in two different arteries. Rings of carotid and femoral arteries from control and trained rabbits were suspended in organ baths for isometric recording of tension. Endothelial nitric oxide synthase (eNOS), Cu/Zn and Mn-superoxide dismutase (SOD), and large conductance calcium activated potassium (BKCa) channel protein expression were measured by western blotting. In the carotid artery, training reduced the relaxation to ACh (10-9 to 3 × 10-6  m) that was reversed by N-acetylcysteine (10-3  m). l-NAME (10-4  m) reduced the relaxation to ACh in both groups, although the effect was lower in the trained group (in mean ± SEM, 39 ± 2% vs. 28 ± 3%). Physical training did not modify the relaxation to ACh in femoral arteries, although the response to l-NAME was lower in the trained group (in mean ± SEM, 41 ± 5% vs. 17 ± 2%). Charybdotoxin (10-7  m) plus apamin (10-6  m) further reduced the maximal relaxation to ACh only in the trained group. The remaining relaxation in both carotid and femoral arteries was abolished by KCl (2 × 10-2  m) and BaCl2 (3 × 10-6  m) plus ouabain (10-4  m) in both groups. Physical training decreased eNOS expression in both carotid and femoral arteries and Cu/Zn and Mn-SOD expression only in the carotid artery. BKCa channels were overexpressed in the trained group in the femoral artery. In conclusion, chronic exercise induces endothelial dysfunction in the carotid artery as a result of oxidative stress. In the femoral artery, it modifies the vasodilator pathways, enhancing the participation of BKCa channels, thus compensating for the impairment of NO-mediated vasodilatation.

Effects of photodynamic therapy in periodontal treatment: A randomized, controlled clinical trial.

AIM: To evaluate the effects of photodynamic therapy (PDT) in the nonsurgical treatment of chronic periodontitis. MATERIALS AND METHODS: A randomized, single-blind, controlled, parallel-group clinical trial was performed. Sixty patients were enrolled: 20 healthy controls and 40 patients with periodontitis. The 40 patients were randomized for scaling and root planing (SRP) or SRP + PDT. Periodontal (plaque index, probing depth, clinical recession, clinical attachment level, bleeding on probing and gingival crevicular fluid volume, corresponding to 381 versus 428 critical sites), microbiological (Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Prevotella intermedia and Campylobacter rectus presence, 18 versus 19 samples) and biochemical (interleukin (IL)-1ß, IL-6 and tumour necrosis factor (TNF)-α, receptor activator of nuclear factor-kappaB ligand (RANK-L) and osteoprotegerin (OPG) levels, 18 versus 19 samples) parameters were recorded. RESULTS: Within each group, significant improvements were found for clinical parameters, though without significant differences between groups. RANK-L was significantly decreased at week 13 in the SRP + PDT group compared with the SRP group. SRP + PDT, but not SRP alone, significantly reduced the abundance of A. actinomycetemcomitans. CONCLUSIONS: Except for a significant decrease in the pathogenic burden of A. actinomycetemcomitans, coadjuvant PDT resulted in no additional improvement compared with SRP alone in patients diagnosed with moderate-to-advanced chronic periodontitis.

Bifidobacterium pseudocatenulatum CECT 7765 supplementation restores altered vascular function in an experimental model of obese mice.

Aims.Bifidobacterium pseudocatenulatum CECT 7765 improves metabolic and immunological altered functions in high fat fed mice, however little is known about the effects of potential probiotics on vascular reactivity. The aim of the present study was to investigate the effects of a potential probiotic strain, Bifidobacterium pseudocatenulatum CECT 7765, on vascular response in obese mice. Methods. Aorta samples were obtained from mice, which were divided into three groups: a control group, receiving a standard diet; an obese group, receiving a high-fat diet; and an obese group receiving high-fat diet and a daily dose of B. pseudocatenulatum CECT 7765 by oral gavage. Aortic rings were suspended in organ baths for isometric recording of tension. mRNA expression of eNOS was evaluated by real-time polymerase chain reaction. Results. Contractions induced by KCl, noradrenaline and thromboxane analogue were 33%, 30% and 45% lower respectively in aortic rings from obese mice. Bifidobacteria administration reversed this effect. eNOS inhibition increased the response to noradrenaline in the three groups with a significant lower magnitude in aortic rings from obese mice receiving bifidobacteria supplement. Acetylcholine caused a greater vasodilation in aorta from obese group (46±3% for control and 69±4% for obese group; p<0.05) and bifidobacteria reversed it (57±5%). Response to sodium nitroprusside was displaced 2.9 times to the left in a parallel manner in obese group. Relaxation to sodium nitroprusside remained unchanged in the bifidobacteria fed group. There was about five-fold decreased mRNA expression of eNOS in aortic segments from the group receiving bifidobacteria. Conclusion.Bifidobacterium pseudocatenulatum CECT 7765 restores the obesity-induced altered vascular function mainly by reducing nitric oxide release.

Neuronal Effects of Sugammadex in combination with Rocuronium or Vecuronium.

Rocuronium (ROC) and Vecuronium (VEC) are the most currently used steroidal non-depolarizing neuromuscular blocking (MNB) agents. Sugammadex (SUG) rapidly reverses steroidal NMB agents after anaesthesia. The present study was conducted in order to evaluate neuronal effects of SUG alone and in combination with both ROC and VEC. Using MTT, CASP-3 activity and Western-blot we determined the toxicity of SUG, ROC or VEC in neurons in primary culture. SUG induces apoptosis/necrosis in neurons in primary culture and increases cytochrome C (CytC), apoptosis-inducing factor (AIF), Smac/Diablo and Caspase 3 (CASP-3) protein expression. Our results also demonstrated that both ROC and VEC prevent these SUG effects. The protective role of both ROC and VEC could be explained by the fact that SUG encapsulates NMB drugs. In BBB impaired conditions it would be desirable to control SUG doses to prevent the excess of free SUG in plasma that may induce neuronal damage. A balance between SUG, ROC or VEC would be necessary to prevent the risk of cell damage.

Effects of Ranolazine on Astrocytes and Neurons in Primary Culture.

Ranolazine (Rn) is an antianginal agent used for the treatment of chronic angina pectoris when angina is not adequately controlled by other drugs. Rn also acts in the central nervous system and it has been proposed for the treatment of pain and epileptic disorders. Under the hypothesis that ranolazine could act as a neuroprotective drug, we studied its effects on astrocytes and neurons in primary culture. We incubated rat astrocytes and neurons in primary cultures for 24 hours with Rn (10-7, 10-6 and 10-5 M). Cell viability and proliferation were measured using trypan blue exclusion assay, MTT conversion assay and LDH release assay. Apoptosis was determined by Caspase 3 activity assay. The effects of Rn on pro-inflammatory mediators IL-ß and TNF-α was determined by ELISA technique, and protein expression levels of Smac/Diablo, PPAR-γ, Mn-SOD and Cu/Zn-SOD by western blot technique. In cultured astrocytes, Rn significantly increased cell viability and proliferation at any concentration tested, and decreased LDH leakage, Smac/Diablo expression and Caspase 3 activity indicating less cell death. Rn also increased anti-inflammatory PPAR-γ protein expression and reduced pro-inflammatory proteins IL-1 ß and TNFα levels. Furthermore, antioxidant proteins Cu/Zn-SOD and Mn-SOD significantly increased after Rn addition in cultured astrocytes. Conversely, Rn did not exert any effect on cultured neurons. In conclusion, Rn could act as a neuroprotective drug in the central nervous system by promoting astrocyte viability, preventing necrosis and apoptosis, inhibiting inflammatory phenomena and inducing anti-inflammatory and antioxidant agents.

WIN 55,212-2, agonist of cannabinoid receptors, prevents amyloid ß1-42 effects on astrocytes in primary culture.

Alzheimer's disease (AD), a neurodegenerative illness involving synaptic dysfunction with extracellular accumulation of Aß1-42 toxic peptide, glial activation, inflammatory response and oxidative stress, can lead to neuronal death. Endogenous cannabinoid system is implicated in physiological and physiopathological events in central nervous system (CNS), and changes in this system are related to many human diseases, including AD. However, studies on the effects of cannabinoids on astrocytes functions are scarce. In primary cultured astrocytes we studied cellular viability using MTT assay. Inflammatory and oxidative stress mediators were determined by ELISA and Western-blot techniques both in the presence and absence of Aß1-42 peptide. Effects of WIN 55,212-2 (a synthetic cannabinoid) on cell viability, inflammatory mediators and oxidative stress were also determined. Aß1-42 diminished astrocytes viability, increased TNF-α and IL-1ß levels and p-65, COX-2 and iNOS protein expression while decreased PPAR-γ and antioxidant enzyme Cu/Zn SOD. WIN 55,212-2 pretreatment prevents all effects elicited by Aß1-42. Furthermore, cannabinoid WIN 55,212-2 also increased cell viability and PPAR-γ expression in control astrocytes. In conclusion cannabinoid WIN 55,212-2 increases cell viability and anti-inflammatory response in cultured astrocytes. Moreover, WIN 55,212-2 increases expression of anti-oxidant Cu/Zn SOD and is able to prevent inflammation induced by Aß1-42 in cultured astrocytes. Further studies would be needed to assess the possible beneficial effects of cannabinoids in Alzheimer's disease patients.