Preventive effects of a nutraceutical mixture of berberine, citrus and apple extracts on metabolic disturbances in Zucker fatty rats.

BACKGROUND: The prevention of obesity represents a major health and socio-economic challenge. Nutraceuticals are regularly highlighted for their beneficial effects in preventing the metabolic disturbances associated with obesity. However, few studies have described the combined action of nutraceutical mixtures combining polyphenols with alkaloids. OBJECTIVE: The aim of this study was to evaluate the effects of long-term dietary supplementation with a mixture of Berberine, Citrus and Apple extracts (BCA) in the primary prevention of obesity and its metabolic and vascular complications in the obese Zucker rat, a spontaneous model of genetic obesity and insulin resistance. METHODS: Sixteen 8-week-old obese Zucker male rats were randomly divided into two groups: all rats received oral gavage daily either with water, untreated obese (U-ObZ) or BCA (BCA-ObZ) mixture for thirteen weeks. Morphological and metabolic parameters were measured along the study. Cumulative concentration-response curves to insulin, acetylcholine and phenylephrine were determined on isolated thoracic aorta. Colon permeability measurements were performed using the Ussing chamber technique. Fecal samples collected at the beginning and the end of the protocol were used as a template for amplification of the V3-V4 region of the 16S rDNA genes. RESULTS: BCA supplementation reduced weight gain (p<0.05) and food intake (p<0.05) in the BCA-ObZ group rats compared to the U-ObZ group rats. It also improved glucose tolerance (p<0.001) and decreased fasting insulin and Homeostasis model assessment index (p<0.05). Through ex vivo experiments, the BCA mixture enhanced significantly aortic insulin relaxation (p<0.01), reduced α1-adrenoceptor-mediated vasoconstriction (p<0.01), and decreased distal colon permeability. Moreover, short-chain fatty acid producers such as Bacteroides, Blautia, and Akkermansia were found to be increased by the BCA mixture supplementation. CONCLUSION: The results showed that a 13-week-supplementation with BCA mixture prevented weight gain and improved glucose metabolism in obese Zucker rats. We also demonstrated that BCA supplementation improved vascular function, colonic barrier permeability and gut microbiota profile.

Effects of chronic mirabegron treatment on metabolic and cardiovascular parameters as well as on atherosclerotic lesions of WHHL rabbits with high-fructose high-fat diet-induced insulin resistance.

BACKGROUND AND AIM: Metabolic syndrome (MetS) is a global health and economic burden. Finding a suitable pharmacological approach for managing this syndrome is crucial. We explored the therapeutic potential of mirabegron (MIR), a ß3-adrenergic receptor agonist, as a repurposed agent for the treatment of MetS and its cardiovascular consequences. METHODS: Thirty Watanabe heritable hyperlipidemic rabbits (WHHL) were divided into 3 groups: control, high-fructose high-fat diet (HFFD) and HFFD + MIR that received a chow diet, HFFD and HFFD along with MIR treatment, respectively. The protocol lasted for 12 weeks, during which weight and abdominal circumference were monitored; plasma fasting levels of lipids, glucose and insulin were measured and an intravenous glucose tolerance test (IVGTT) was performed. Homeostasis model assessment of insulin resistance (HOMA-IR) was calculated. Cardiac function was assessed using in-vivo and ex-vivo approaches. Vascular reactivity was estimated via isolated carotid arteries method. Aortic atherosclerosis was evaluated using histological and immuno-histochemical techniques. RESULTS: In contrast to the HFFD group, MIR-treated rabbits showed fasting insulin, HOMA-IR and TG levels stabilization and exhibited improved cardiac inotropy and lusitropy, while on the other hand, displayed aggravated atheroma plaque development. CONCLUSION: Long-term treatment with MIR prevented the increase in TG levels and the establishment of IR and enhanced the cardiac function of a rabbit animal model of MetS with combined dyslipidemia and IR.

Long-term high-fructose high-fat diet feeding elicits insulin resistance, exacerbates dyslipidemia and induces gut microbiota dysbiosis in WHHL rabbits.

The metabolic syndrome (MetS) has become a global public health burden due to its link to cardiovascular disease and diabetes mellitus. The present study was designed to characterize the metabolic and cardiovascular disturbances, as well as changes in gut microbiota associated with high-fructose high-fat diet (HFFD)-induced MetS in Watanabe heritable hyperlipidemic (WHHL) rabbits. Twenty-one Watanabe rabbits were assigned to a control (n = 9) and HFFD (n = 12) groups, receiving a chow diet and a HFFD, respectively. During a 12-weeks protocol, morphological parameters were monitored; plasma fasting levels of lipids, glucose and insulin were measured and a glucose tolerance test (GTT) was performed. HOMA-IR was calculated. Cardiac function and vascular reactivity were evaluated using the Langendorff isolated heart and isolated carotid arteries methods, respectively. 16S rRNA sequencing of stool samples was used to determine gut microbial composition and abundance. HFFD-fed Watanabe rabbits exhibited increased fasting insulin (p < 0.03, 12th week vs. Baseline), HOMA-IR (p < 0.03 vs. Control), area under the curve of the GTT (p < 0.02 vs. Control), triglycerides (p < 0.05, 12th week vs. Baseline), TC (p < 0.01 vs. Control), LDL-C (p < 0.001 vs. Control). The HFFD group also displayed a significant decrease in intestinal microbial richness, evenness and diversity (FDR < 0.001, FDR < 0.0001, FDR < 0.01, respectively vs. Control group) and an increase in its Firmicutes/Bacteroidetes ratio (R = 3.39 in control vs. R = 28.24 in the HFFD group) indicating a shift in intestinal microbial composition and diversity. Our results suggest that HFFD induces insulin resistance and gut microbiota dysbiosis and accentuates dyslipidemia; and that, when subjected to HFFD, Watanabe rabbits might become a potential diet-induced MetS animal models with two main features, dyslipidemia and insulin resistance.

Soluble guanylate cyclase chronic stimulation effects on cardiovascular reactivity in cafeteria diet-induced rat model of metabolic syndrome.

Metabolic syndrome is linked to an increased risk of cardiovascular complications by a mechanism involving mainly decreased nitric oxide (NO) bioavailability and impaired NO-soluble guanylate cyclase (sGC)- cyclic guanosine monophosphate (cGMP) signalling (NO-sGC-cGMP). To further develop this scientific point, this study aimed to investigate the effects of long-term treatment with BAY 41-2272 (a sGC stimulator) on cardiovascular reactivity of spontaneously hypertensive rats (SHR) as a model of metabolic syndrome. SHR were randomly divided into 3 groups: control group, cafeteria diet (CD)-fed group and CD-fed group treated daily with BAY 41-2272 (5 mg/kg) by gastric gavage for 12 weeks. In vivo measurements of body weight, abdominal circumference, blood pressure and glucose tolerance test were performed. At the end of the feeding period, ex vivo cumulative concentration-response curves were performed on isolated perfused heart (isoproterenol (0.1 nM - 1 µM)) and thoracic aorta (phenylephrine (1 nM-10 µM), acetylcholine (1 nM-10 µM), and sodium nitroprusside (SNP) (0.1 nM-0.1 µM)). We showed that chronic CD feeding induced abdominal obesity, hypertriglyceridemia, glucose intolerance and exacerbated arterial hypertension in SHR. Compared to control group, CD-fed group showed a decrease in ß-adrenoceptor-induced cardiac inotropy, in coronary perfusion pressure and in aortic contraction to phenylephrine. While relaxing effects of acetylcholine and SNP were unchanged. BAY 41-2272 long-term treatment markedly prevented arterial hypertension development and glucose intolerance, enhanced the α1-adrenoceptor-induced vasoconstriction, and restored cardiac inotropy and coronary vasodilation. These findings suggest that BAY 41-2272 may be a potential novel drug for preventing metabolic and cardiovascular complications of metabolic syndrome.

Sildenafil citrate long-term treatment effects on cardiovascular reactivity in a SHR experimental model of metabolic syndrome.

Much evidence indicates that metabolic syndrome is strongly correlated with a decrease in nitric oxide and an increase in oxidative stress leading to cardiovascular alterations. In recent years, gut microbiota has emerged as a new contributor to the metabolic syndrome establishment and associated cardiovascular diseases, but the underlying mechanisms remain unclear. We hypothesized that a positive modulation of cyclic guanosine monophosphate (cGMP) pathway, through phosphodiesterase type 5 (PDE5) inhibition could prevent cardiovascular alterations and gut dysbiosis that may be associated to metabolic syndrome. Spontaneously hypertensive rats (SHR) were randomly divided into 4 groups: control, cafeteria diet (CD) and sildenafil citrate treated groups (5mg/kg per os) were given either a CD or a standard chow diet for 10 weeks. Body weight, arterial blood pressure and glucose tolerance test were monitored. At the 10th week, cardiac inotropy and coronary perfusion pressure were evaluated on isolated heart according to Langendorff method. Cumulative concentration response curves to phenylephrine and acetylcholine were determined on thoracic aorta rings for vascular reactivity evaluation. Faecal samples were collected for the gut microbiota analysis. Compared to the control group, CD-fed rats showed a significant increase in body weight gain, arterial blood pressure and were glucose intolerant. This group showed also a decrease in ß-adrenoceptor-induced cardiac inotropy and coronary vasodilation. Gut microbiota analysis revealed a significant reduction in the abundance of Lactobocillus spp in cafeteria diet-fed rats when compared to the control ones. Sildenafil citrate long-term treatment decreased weight gain and arterial blood pressure, improved coronary vasodilation and reduced α1-adrenoceptor-induced vasoconstriction in CD group. However, it did not reverse gut dysbiosis induced by chronic CD feeding. These results suggest that cGMP pathway targeting may be a potential therapeutic strategy for the management of the metabolic syndrome and associated cardiovascular disorders.

Long-term atorvastatin treatment decreases heart maximal oxygen consumption and its vulnerability to in vitro oxidative stress in Watanabe heritable hyperlipidemic rabbit.

Statins are currently used in prevention of cardiovascular diseases in high-risk populations, and could be considered in primary prevention. However, few studies are available on the long-term effects of low doses of statins, especially on mitochondrial function and reactive oxygen species (ROS) metabolism at cardiac level. This study aimed to determine potential effects of a long-term atorvastatin treatment, at low-dose concentration, on the myocardium mitochondrial respiration. Thirty-four Watanabe rabbits were treated or not with atorvastatin (2.5 mg·kg-1·day-1) from the age of 3 to 12 months. Every 3 months, proton leak, basal (V0), and maximal (Vmax) mitochondrial respiration on cardiac permeabilized fibers were measured. Additionally, the vulnerability to ROS, cardiac enzymatic antioxidant defenses, and oxidative damage (lipoperoxidation) were analyzed. Proton leak increased over the duration of the experiment (up to 60% from Vmax at 12 months). Moreover, the statin treatment induced a decrease of Vmax and a decrease of ROS susceptibility of cardiac mitochondria. However, the lipoperoxidation and the antioxidant defenses were not dependent on the presence of statin treatment, or on its duration. This is the first study showing a protective effect of long-term statins treatment against the ROS susceptibility in the cardiac muscle.

The relaxant effect of the Montivipera bornmuelleri snake venom on vascular contractility.

Molecular richness of snake venoms is an important source of proteins and toxins with potent effects on the cardiovascular system. The alteration of the vascular system in the victim after a venomous snake bite is usually expressed by a significant decrease in blood pressure. Therefore, exploring snake venom to extract and characterize its biomolecules is of considerable medical interest, and formed the basis of this study. We assessed the potential of the venom of Montivipera bornmuelleri, a viper from Lebanon, to induce relaxant effect on isolated Wistar rat aorta via several mechanisms of action. The overall hypotensive effect of Montivipera bornmuelleri venom results from its synergetic action on different channels for the reduction of blood pressure. By actions of its metalloproteinases and phospholipase A2, the venom may induce the production of nitric oxide acting accordingly a vasodilator effect. It could act on the voltage-dependent potassium channels and/or the L-type calcium channels, inhibiting angiotensin converting enzyme and/or inhibiting the α1-adrenoceptors. This work demonstrates vasorelaxant effect of the Montivipera bornmuelleri venom acting on different pathways, reducing blood pressure.

Effect of nebivolol treatment during pregnancy on the intrauterine fetal growth, mortality and pup postnatal development in the l-NAME-induced hypertensive rats.

The present study was carried out to evaluate the effect of nebivolol vs. bisoprolol treatment on the intrauterine fetal growth, mortality and postnatal development in Nω-Nitro-l-arginine methyl ester hydrochloride (l-NAME)-induced hypertensive rats. Hypertension was induced in normotensive pregnant Wistar rats by daily administration of l-NAME (100mg/kg/day, in the drinking water) for the period of pregnancy. After 9 days of l-NAME treatment, rats with systolic and diastolic blood pressure (SBP and DBP) more than 140/90mmHg were considered hypertensive. Then, some of them were treated from day 11 to day 18 of pregnancy with nebivolol (8mg/kg/day) or bisoprolol (10mg/kg/day) via oral gavage. SBP, DBP and heart rate (HR) were re-evaluated by tail cuff method on day 19 of pregnancy and morphometrical or histological studies were performed on day 20. In addition, the mortality and postnatal development of newborn pups were assessed in all groups. The l-NAME administration during pregnancy induced an increase in SBP and DBP while HR did not change. Nebivolol or bisoprolol treatment completely prevented the elevation of SBP and DBP induced by l-NAME with a reduction in HR in pregnant and non-pregnant rats. The intra-uterine fetal growth and the postnatal development of newborn rats in nebivolol-treated hypertensive group were significantly lower vs. control and higher vs. bisoprolol-treated group with a higher mortality in the both types of treatments vs. control rats. The nebivolol and bisoprolol administration produce adverse effects on fetal growth and postnatal development, that limits their therapeutic use in females during pregnancy.

Prospective pilot study to detect dogs with non food-induced canine atopic dermatitis using Fourier transform infrared spectroscopy.

BACKGROUND: The diagnosis of canine atopic dermatitis (CAD) remains challenging due to the lack of a simple biomarker or metabolic profile. In human medicine, Fourier transform infrared spectroscopy (FTIR) is an analytical technique used for several diseases. It requires a small amount of sample and allows the identification of structural moieties of biomolecules on the basis of their infrared absorption, with limited sample pretreatment. HYPOTHESIS/OBJECTIVES: The aim of the study was to evaluate the diagnostic value of FTIR. ANIMALS: Three groups were tested: 21 dogs with non food-induced CAD (NFICAD), 16 dogs with inflammatory conditions of various origins but without allergic dermatoses (OD) and 10 healthy dogs (H). METHODS: Peripheral blood was collected and spectra were acquired with a FTIR spectrophotometer. A principal component analysis (PCA) was performed on the full wavenumber spectra (4000-600/cm), followed by a Fisher discriminant analysis (DA) to assess the differences between the three groups. RESULTS: The PCA followed by the DA of whole spectra showed significant differences between the three groups. These results suggest that by using the FTIR method, dogs with NFICAD can be differentiated from healthy dogs and dogs with nonallergic inflammation. There was no overlap between the spectral data of the three groups indicating that NFICAD dogs were correctly segregated from the H and OD groups. CONCLUSIONS: A study on a larger cohort including common pruritic skin diseases is necessary to confirm these initial results and the relevance of this diagnostic technique.

A non-hypocholesterolemic atorvastatin treatment improves vessel elasticity by acting on elastin composition in WHHL rabbits.

BACKGROUND AND AIMS: Statins are prescribed for their preventative effects within atherosclerosis development. To our knowledge, no study focusing on very low-dose (non-hypolipidemic effect) and long-term atorvastatin treatment in vivo was available. Our aim was to assess the effect of such atorvastatin treatment on the mechanical and functional characteristics of arteries in the context of primary prevention. METHODS: An atorvastatin treatment (2.5 mg/kg/day) was tested against controls on 34 male 3 to 12 month-old WHHL rabbits. No effect on total cholesterol, triglycerides, HDL or LDL was observed. The arterial stiffness was evaluated on vigil animals by pulse wave velocity (PWV) measurement. Then, in vitro measurements were made to evaluate (1) the endothelial and vascular smooth muscle function, (2) the elasticity of the arterial wall and (3) the composition in collagen and elastin in the aorta. RESULTS: The PWV increasing observed with age in control group was canceled by treatment, creating a significance difference between groups at 12 months (5.17 ± 0.50 vs 2.14 ± 0.34 m s(-1) in control and treated groups respectively). Vasoreactivity modifications can't explain this result but maintain of elasticity with treatment in large arteries was confirm by a static tensile test. A first possible explanation is the change of wall composition with treatment, validated by the percentage of elastin at 12 months, 4.4% lower in the control group compared to the treated group (p < 0.05). CONCLUSIONS: This study shows that a non-hypocholesterolemic statin treatment could improve vessel elasticity in the atherosclerotic WHHL model. The great novelty of this work is the vessel wall composition changing associated. This first approach in animal opens the reflection on the use of these low doses in humans. This could be interesting in the context of arterial stiffening with aging, non-hyperlipidemic atherosclerosis or with cholesterol reduce by another therapy or lifestyle modification.

Change in vascular smooth muscle response to 5-HT due to short- or long-term endothelial denudation of the bovine digital vein.

Several chronic progressive vascular diseases, such as laminitis, show vasocontractile dysfunction that might evolve into reperfusion injury and/or vessel structural remodelling, which may be traced back to aberrant endothelial function. In the present study, the vasomotor responses of bovine digital veins (BDVs) to 5-hydroxytryptamine (5-HT) were investigated in blood vessels, with and without endothelium present, and in samples deprived of endothelium before or after overnight incubation in tissue culture medium, to evaluate the effects of short- and long-term endothelial damage on vascular smooth muscle (VSM) reactivity. No significant effects were observed in the blood vessels tested immediately after the removal of endothelium. In contrast, a significant increase in VSM reactivity to 5-HT was seen in vessels incubated without endothelium. This long-term change in smooth muscle reactivity was prevented by exposure to the nitric oxide (NO) donor nitroprusside (P < 0.01), suggesting that the long-term lack of inhibitory control exerted by endothelium-derived NO is involved in increased VSM reactivity. The RhoA/ROCK pathway inhibitor fasudil reduced VSM hyper-contractility to ~65% (P < 0.001), the superoxide dismutase-mimetic tempol normalised the vascular response and the non-selective COX-inhibitor indomethacin exerted a moderate inhibitory effect (P < 0.05). Thus, over-activation of the RhoA/ROCK pathway and production of reactive oxygen species could account for VSM hyper-reactivity, triggered by long-term endothelium-deprivation in BDVs, suggesting that these biochemical mechanisms are potential targets for controlling the progressive vasocontractile dysfunction of digital veins in animals affected with laminitis.

Effect of nebivolol treatment during pregnancy on the genital circulation, fetal growth and postnatal development in the Wistar rat.

The aim of study was to evaluate the effects of nebivolol, a cardioselective beta-1 adrenergic receptor blocker of the third generation with vasodilatory properties, vs. bisoprolol on the genital circulation, uterine vasculature, fetal growth and postnatal development in pregnant Wistar rats. Non invasive measurements of systolic and diastolic blood pressure (SBP and DBP) and heart rate (HR), and invasive measurement of genital blood flow (GBF) were taken in pregnant rats, by tail cuff and transonic probe methods respectively, after an oral treatment by gastric gavage with nebivolol (8mg/kg/day) or bisoprolol (10mg/kg/day) from day 11 to day 18 of pregnancy. Other morphometrical and histological measurements were performed on the ovarian and uterine arteries to evaluate the effect of nebivolol on the uterine vasculature. Furthermore, postnatal mortality and pup growth were recorded. The data demonstrated that nebivolol (compared with bisoprolol) induced a significant decrease in SBP, HR and GBF while DBP remained unchanged. Moreover, nebivolol increased the diameter and the length of ovarian and uterine arteries and the number of uterine artery segmental branches. The results also showed that the body weight gain of newborns in the nebivolol group was significantly lower vs. bisoprolol and vs. control with a higher mortality rate. The nebivolol action is not only limited to its favorable hemodynamic effects represented by a decrease in blood pressure, but it also produces adverse effects on fetal growth and postnatal development that may limit its therapeutic use in females during pregnancy.

Increased numbers of peripheral blood CD34+ cells in dogs with canine atopic dermatitis.

BACKGROUND: The bone marrow may be involved in human atopic diseases, as shown by the release of CD34+ cells into the peripheral blood. HYPOTHESIS/OBJECTIVES: The aim was to determine the numbers of CD34+ cells in atopic dogs. ANIMALS: The following three groups of dogs were studied: 27 dogs with nonfood-induced atopic dermatitis (NFICAD); 16 dogs with nonallergic inflammatory diseases; and 13 healthy control dogs. METHODS: Dogs with NFICAD were selected after fulfilment of Favrot's criteria and exclusion of other pruritic dermatoses, including flea infestation and adverse reaction to foods. The Canine Atopic Dermatitis Extent and Severity Index (CADESI)-03 and a Visual Analog Scale (VAS) score for pruritus were used to quantify clinical signs. A phycoerythrin-conjugated anticanine CD34 antibody was used to stain peripheral blood CD34+ cells, and these were enumerated using a flow cytometer. The CD34+ cell counts were compared between groups and tested (in the NFICAD group) for correlation with the severity of clinical signs. RESULTS: The numbers of peripheral CD34+ cells in dogs with NFICAD (median 1.7) were statistically higher than in dogs with other nonallergic inflammatory diseases (median 1.0; P = 0.01) and healthy control dogs (median 0.9; P = 0.009). In dogs with NFICAD, there was no correlation between CD34+ cell numbers and CADESI-03 scores or owner-assessed pruritus (VAS score). CONCLUSIONS AND CLINICAL IMPORTANCE: The results of this study suggest the possible involvement of CD34+ cells in dogs with NFICAD. The role of CD34+ cells in the aetiopathogenesis of canine atopic dermatitis remains to be determined.

Equine digital veins are more sensitive to superoxide anions than digital arteries.

This work was designed to investigate (i) the effect of superoxide dismutase (SOD) inhibition on endothelial function and (ii) the free radical-induced endothelial dysfunction in equine digital veins (EDVs) and equine digital arteries (EDAs) isolated from healthy horses. EDV and EDA rings were suspended in a 5 ml organ bath containing Krebs solution. After a 60 min equilibration period, EDV and EDA rings were contracted with phenylephrine. Then, cumulative concentration-response curves (CCRCs) to acetylcholine were performed. In both EDVs and EDAs, acetylcholine (1 nM to 10 µM) produced concentration-dependent relaxation. We investigated the influence of SOD inhibition by diethyldithiocarbamate (DETC; 100 µM), a CuZnSOD inhibitor, on EDAs and EDVs relaxant responses to acetylcholine. Acetylcholine -mediated relaxation was impaired by DETC only in EDVs. SOD activity assayed by a xanthine-xanthine oxidase method was higher in EDAs compared with EDVs (P<0.05). CCRCs to acetylcholine established in the presence of pyrogallol (30 µM) or homocysteine (20 µM), two superoxide anions generating systems showed that in both EDVs and EDAs, the acetylcholine-mediated relaxation was significantly impaired by pyrogallol and homocysteine. This impairment was more pronounced in EDVs than in EDAs. Moreover, the pyrogallol-induced impairment of acetylcholine-mediated relaxation was potentiated by DETC to a greater extent in EDVs. We concluded that due to the lower activity of SOD, EDVs are more sensitive to superoxide anions than EDAs. So, any alteration of superoxide anions metabolism is likely to have a more important impact on venous rather than arterial relaxation.

Protein content analysis and antimicrobial activity of the crude venom of Montivipera bornmuelleri; a viper from Lebanon.

Viperidae snakes venoms represent a source of efficient bioactive components that have already led to the development of several new drugs. In this work, we analyzed the protein content of the Montivipera bornmuelleri crude venom using LC-ESI-MS, sephadex G-75 gel filtration and SDS-PAGE and demonstrated the presence of proteins with molecular masses corresponding to metalloprotease III, serine-protease and PLA2 in three fractions collected after gel filtration. Equally, we examined the antimicrobial effect of the venom that showed an important potency, as bactericidal agent, based on MBC and MIC values obtained, against Staphylococcus aureus and Morganella morganii bacteria. However, no activity was registered against Enterococcus faecalis, being the most resistant bacteria, neither against Aspergillus flavus and Penicillium digitatum fungal. Furthermore, on eleven other bacterial strains and the Candida albicans fungus, the venom has shown an intermediate efficacy by slightly reducing the growth. Our data concerning the Montivipera bornmuelleri venom give evidence of a rich and complex content aiding the exploration of new bioactive molecules for biopharmaceuticals purposes.

Celiprolol induces ß(3)-adrenoceptors-dependent relaxation in isolated porcine coronary arteries.

In porcine coronary arteries (PCAs), celiprolol, a selective ß(1)-adrenoceptors antagonist, induces vasodilatation by an endothelium- and nitric oxide (NO)-dependent pathway. However, the mechanisms of that vascular effect have not been precisely established. ß(3)-Adrenoceptors have been shown to be involved in the relaxation per se of various vascular beds, including coronary vessels. Thus, we evaluated (i) the presence of ß(3)-adrenoceptors in the PCA and (ii) their role in celiprolol-induced vasodilatation. PCA rings were placed in organ baths and preconstricted with KCl. All experiments were performed in the presence of nadolol (a ß(1)/ß(2)-adrenoceptor antagonist). Cumulative concentration-response curves to SR 58611A and ICI 215001 (2 ß(3)-adrenoceptor agonists) and to celiprolol were constructed. We also used semiquantitative reverse transcription - polymerase chain reaction, which clearly showed the presence of ß(3)-adrenoceptor transcripts. SR 58611A, ICI 215001, and celiprolol induced concentration-dependent relaxations in PCA rings. SR 58611A-induced relaxation was almost abolished after removal of endothelium or pretreatment with L-NAME (a NO synthase inhibitor). The vasorelaxations induced by SR 58611A and celiprolol were inhibited in the presence of SR 59230A and L-748337 (2 selective ß(3)-adrenoceptor antagonists). We showed (i) that PCAs possess functional ß(3)-adrenoceptors mediating endothelium- and NO-dependent relaxation, and (ii) that celiprolol exerts a ß(3)-adrenoceptor agonistic activity in this vascular bed.

Vasorelaxant effect of propentofylline in isolated equine digital veins.

We evaluated the vasorelaxant effect of propentofylline (PPF), a methylxanthine derivative, and its mechanism of action in equine digital veins (EDVs). Cumulative concentration-response curves to PPF (1 nM-300 µM) were recorded in phenylephrine-precontracted EDV rings under different experimental conditions. PPF-induced relaxation was partially inhibited by endothelium removal, but was unaltered by CGS-15943 (an adenosine receptor antagonist; 3 µM). PPF-induced relaxation was partially inhibited in the presence of L-NAME (a nitric oxide (NO) synthase inhibitor; 100 µM), ODQ (an inhibitor of soluble guanylyl cyclase; 30 µM) or Rp-8-Br-PET-cGMP-S (a protein kinase G inhibitor; 3 µM). It was not modified by indomethacin (a non-selective cyclooxygenase (COX) inhibitor; 10 µM), and was slightly potentiated by H-89 (a protein kinase A inhibitor; 2 µM). In endothelium-intact EDVs, PPF-induced relaxation was associated with a 2.4- and 24.1-fold increase in the tissue cGMP and cAMP content respectively. PPF (100 µM) did not shift the concentration-response curve to phenylephrine (1 nM-300 µM) but reduced the maximal effect. To investigate whether PPF can affect cAMP- and cGMP-induced relaxations, relaxation curves to forskolin (an activator of adenylate cyclase) and to sodium nitroprusside (SNP, a NO donor) were recorded in EDV rings pretreated with PPF (100 µM). PPF only slightly potentiated the forskolin-induced relaxation without affecting the SNP-induced relaxation. We demonstrated that PPF-induced relaxation in EDVs is partially endothelium-dependent. The PPF-induced relaxation partially occurred via NO release and both cAMP and cGMP generation, through COX-independent mechanisms but could also result from the inhibition of cAMP-phosphodiesterase activity for the highest concentrations.

Characterization of pruritus in canine atopic dermatitis, flea bite hypersensitivity and flea infestation and its role in diagnosis.

BACKGROUND: In dogs, flea infestation (FI), flea bite hypersensitivity (FBH) and canine atopic dermatitis (CAD) have been mainly characterized by their lesions but never by their pruritus. In clinical practice, many of these dogs exhibit only pruritus. HYPOTHESIS/OBJECTIVES: The purpose of this study was to evaluate the characteristics of pruritus in these dermatoses and their potential usefulness for diagnosis. ANIMALS: Dogs included were selected from the Oniris clinical data. Cases were selected in which the dogs had only one of the three dermatoses diagnosed. The diagnosis of CAD was based on Prélaud's criteria and positive intradermal tests except flea; for FBH by compatible clinical signs and a response to an intradermal test with flea allergen; and for FI by the presence of fleas. Moreover, in each group, other primary pruritic skin diseases were excluded. METHODS: Location, behavioural manifestations, seasonality and quantification of the pruritus were evaluated. The statistical analysis used chi-squared test with a P-value <0.05. RESULTS: Three hundred and forty-six dogs were analysed, 91 with CAD, 110 FI and 145 FBH. The period (season) of onset was not statistically different either for each dermatosis or among the three dermatoses. Some locations were highly specific for one dermatosis as follows: ventral abdomen/medial surface of thigh (chewing) and radius/carpus/tibia/tarsus (chewing) in FI; back/dorsolumbar area (chewing) and tail (chewing) in FBH; and paws (chewing/licking) and face/neck (rubbing) in CAD. CONCLUSIONS AND CLINICAL IMPORTANCE: Some features of pruritus could be suggestive of the causal disease, with possible diagnostic value in pruritic dogs.

Autoantibodies against cardiac ß(1)-adrenoceptor do not affect the low-affinity state ß(1)-adrenoceptor-mediated inotropy in rat cardiomyocytes.

Circulating autoantibodies directed against the 2nd extracellular loop (EL-2) of ß(1)-adrenoceptors (ß(1)-AABs) have been detected in the serum of patients with various cardiovascular pathologies. ß(1)-AABs induce agonistic, positive inotropic effects via ß(1)-adrenoceptors (ß(1)ARs). In the mammalian heart, ß(1)-AR can exist in 2 distinct activated configurations (the so-called high- and low-affinity states). The aim of the present study was to investigate whether the action of ß(1)-AAB is dependent on the affinity state of ß(1)AR in isolated ventricular cardiomyocytes of adult Wistar rats. Immunoglobulin G (IgG) containing ß(1)-AAB obtained from animals immunized with a peptide corresponding to the EL-2 of human ß(1)-AR, caused a dose-dependent increase in cell shortening. Isoproterenol-induced inotropy was significantly reduced in cardiomyocytes that had been preincubated with IgG containing ß(1)-AAB and in cardiomyocytes isolated from immunized rats. The negative effects of preincubation with IgG containing ß(1)-AAB on the response to isoproterenol was inhibited in the presence of bisoprolol. CGP 12177A and pindolol-induced inotropy was not affected by IgG preincubation or immunization. No detectable inotropic effect of cell shortening was obtained with IgG containing ß(1)-AAB in the presence of propranolol and 3-isobutyl-1-methylxanthine. The present study demonstrates that ß(1)-AABs have no agonist/antagonist-like effects upon low-affinity state ß(1)-ARs. This result indicates that ß(1)-AABs recognize and stabilize the high-affinity state, but are unable to stabilize and (or) induce the low-affinity state receptor.

Evaluation of the role of superoxide anions in endotoxin-induced impairment of beta-adrenoceptor-mediated vasodilation in equine digital veins.

OBJECTIVE: To investigate the role of superoxide anions in the lipopolysaccharide (LPS)-induced impairment of beta-adrenoceptor-mediated equine digital vein (EDV) vasodilation. SAMPLE POPULATION: EDVs isolated from forelimbs of 24 healthy adult horses. PROCEDURES: Endothelium-intact or endothelium-denuded EDV rings were incubated with or without LPS (10 microg/mL) of Escherichia coli (O55:B5) for 4 hours. Cumulative concentration-relaxation curves resulting from administration of isoprenaline, a nonselective beta-adrenoceptor agonist, or from administration of SR 58611A, a selective beta(3)-adrenoceptor agonist, were recorded in phenylephrine-preconstricted EDVs in the absence or the presence of superoxide dismutase (200 U/mL). Isoprenaline-induced relaxation was also evaluated with or without the cyclooxygenase inhibitors indomethacin (10 microM) and NS-398 (10 microM). RESULTS: Isoprenaline and SR 58611A induced concentration-dependent relaxation of EDV rings, which was inhibited by LPS exposure. Superoxide dismutase abolished the inhibitory effect of LPS on the isoprenaline- and SR 58611A-mediated relaxation. Pretreatment of the LPS-treated EDVs with indomethacin or NS-398 restored the isoprenaline-mediated relaxation and abolished the LPS-induced impairment to a similar extent as superoxide dismutase. CONCLUSIONS AND CLINICAL RELEVANCE: Results supported a role of superoxide anions in the LPS-induced impairment of beta-adrenoceptor-mediated EDV vasodilation. The LPS-induced oxidative stress in EDVs may contribute to vascular dysfunctions associated with laminitis in horses.