Teresinha Silva de Brito

Inhaled 1,8-Cineole Reduces Inflammatory Parameters in Airways of Ovalbumin-Challenged Guinea Pigs

Eucalyptol, also known as 1,8-cineole, is a monoterpene traditionally used to treat respiratory disorders due to its secretolytic properties. In addition to its myorelaxant effects, it also has anti-inflammatory actions in vitro. In this study, we aimed to evaluate the efficacy of acute treatment with 1,8-cineole on reducing airway inflammatory parameters. Ovalbumin (OVA)-sensitized guinea pigs were submitted to antigenic challenge (OVA) with or without pre-treatment with a single dose of 1,8-cineole administered by inhalation. Airway inflammatory parameters were reduced or absent in 1,8-cineole-treated animals as compared with untreated guinea pigs. Acute treatment with 1,8-cineole impaired the development of airway hyperresponsiveness to carbachol in isolated tracheal rings. Levels of the pro-inflammatory cytokines TNFα and IL-1β was lower in bronchoalveolar lavage fluid (BALF) of 1,8-cineol-treated guinea pigs than in untreated animals. 1,8-Cineole impaired the OVA-induced increase of the myeloperoxidase activity in BALF. 1,8-Cineole also prevented the reduction of the mucociliary clearance induced by the antigen presentation. The present investigation provides evidence that inhaled 1,8-cineole prevents hyperresponsiveness and inhibits inflammation in airways of ovalbumin-challenged guinea pigs.

1-Nitro-2-phenylethane, the main constituent of the essential oil of Aniba canelilla, elicits a vago-vagal bradycardiac and depressor reflex in normotensive rats

Previously, it was shown that intravenous (i.v.) treatment with the essential oil of Aniba canelilla (EOAC) elicited a hypotensive response that is due to active vascular relaxation rather than to the withdrawal of sympathetic tone. The present study investigated mechanisms underlying the cardiovascular responses to 1-nitro-2-phenylethane, the main constituent of the EOAC. In pentobarbital-anesthetized normotensive rats, 1-nitro-2-phenylethane (1-10mg/kg, i.v.) elicited dose-dependent hypotensive and bradycardiac effects which were characterized in two periods (phases 1 and 2). The first rapid component (phase 1) evoked by 1-nitro-2-phenylethane (10mg/kg) was fully abolished by bilateral vagotomy, perineural treatment of both cervical vagus nerves with capsaicin (250 microg/ml) and was absent after left ventricle injection. However, pretreatment with capsazepine (1mg/kg, i.v.) or ondansetron (30 microg/kg, i.v.) did not alter phase 1 of the cardiovascular responses to 1-nitro-2-phenylethane (10mg/kg, i.v.). In conscious rats, 1-nitro-2-phenylethane (1-10mg/kg, i.v.) evoked rapid hypotensive and bradycardiac (phase 1) effects that were fully abolished by methylatropine (1mg/kg, i.v.). It is concluded that 1-nitro-2-phenylethane induces a vago-vagal bradycardiac and depressor reflex (phase 1) that apparently results from the stimulation of vagal pulmonary rather than cardiac C-fiber afferents. The transduction mechanism of the 1-nitro-2-phenylethane excitation of C-fiber endings is not fully understood and does not appear to involve activation of either Vanilloid TPRV(1) or 5-HT(3) receptors. The phase 2 hypotensive response to 1-nitro-2-phenylethane seems to result, at least in part, from a direct vasodilatory effect since 1-nitro-2-phenylethane (1-300 microg/ml) induced a concentration-dependent reduction of phenylephrine-induced contraction in rat endothelium-containing aorta preparations.

Phytochemical study guided by the myorelaxant activity of the crude extract, fractions and constituent from stem bark of Hymenaea courbaril L.

ETHNOPHARMACOLOGICAL RELEVANCE Hymenaea courbaril L. (Caesalpinoideae) is used in Brazilian folk medicine to treat anemia, kidney problems, sore throat and other dysfunctions of the respiratory system, such as bronchitis and asthma, although such properties are yet to be scientifically validated. AIM OF THE STUDY In order to give a scientific basis to support the traditional use of Hymenaea courbaril, this study was designed to evaluate antioxidant, myorelaxant and anti-inflammatory properties of the ethanol extract from stem bark and its fractions. The myorelaxant effect of astilbin, a flavonoid isolated from the bioactive ethyl acetate fraction (EAF), has also been evaluated. MATERIAL AND METHODS In the present study ethanol extract from stem bark (EEHC) and fractions were analyzed using bioassay-guided fractionation. The following activities were investigated: antioxidant by 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay, myorelaxant on rat tracheal smooth muscle, and anti-inflammatory using ovalbumin-induced leukocytosis and airway hyperresponsiveness in rats. RESULTS The results of the present investigation show that the whole extract of Hymenaea courbaril and some of its fractions strongly scavenged DPPH radical. The extract showed myorelaxant activity on rat trachea, being EAF its highest efficient fraction. Bio-guided study allowed the isolation of astilbin, a well-known flavonoid. The activity induced by this compound indicates that it may be partly responsible for the myorelaxant effect of EAF. EAF reduced contractions that depended on divalent cation inflow through voltage-operated Ca(2+) channels (VOCCs) or receptor-operated Ca(2+) channels (ROCCs), but it was more potent to inhibit VOCC- than ROCC-dependent contraction induced by Ca(2+) addition in ACh-enriched Ca(2+)-free medium. Oral pretreatment of antigen-challenged animals with EAF prevented airway hyperresponsiveness on KCl-induced contraction and reduced the number of total white cells, particularly eosinophils and neutrophils in bronchoalveolar lavage. CONCLUSIONS This study provided scientific basis that Hymenaea courbaril presents potential antioxidant, myorelaxant and anti-inflammatory actions, which support its use in folk medicine to treat inflammatory airway diseases.

The essential oil of Eucalyptus tereticornis, and its constituents α- and β-pinene, potentiate acetylcholine-induced contractions in isolated rat trachea

The effects of the essential oil of Eucalyptus tereticornis (EOET), especially the effects of its constituents alpha- and beta-pinene, were studied on rat trachea in vitro. In tracheal rings, EOET, alpha- or beta-pinene potentiated the contractions induced by acetylcholine (ACh). Contractions induced by K(+) (60mM) were also potentiated by alpha- and beta-pinene, but were reduced by EOET. Our findings show that EOET has myorelaxant effects on rat airways, but potentiates ACh-induced contractions. Monoterpenes alpha- and beta-pinene are involved in its potentiating actions, but are not responsible for its myorelaxant effects. A putative inhibition of the acetylcholinesterase enzyme is involved.

Inhibitory effect of 1,8-cineole on guinea-pig airway challenged with ovalbumin involves a preferential action on electromechanical coupling.

1 1,8‐Cineole is a terpenoid constituent of essential oils with anti‐inflammatory properties. It reduces the neural excitability, functions as an antinociceptive agent and has myorelaxant actions in guinea‐pig airways. The aim of the present study was to investigate the mechanism underlying the myorelaxant effects of 1,8‐cineole in guinea‐pig isolated trachea from either naïve guinea‐pigs or ovalbumin (OVA)‐sensitized animals subjected to antigenic challenge. 2 Isometric recordings were made of the tone of isolated tracheal rings. Rings with an intact epithelium relaxed beyond basal tone in the presence of 1,8‐cineole (6.5 × 10−6 to 2 × 10−2 mol/L) in a concentration‐dependent manner (P < 0.001, anova) with a pD2 value of 2.23 (95% confidence interval 2.10–2.37). Removal of the epithelium or pretreatment of intact tissue for 15 min with 50 µmol/L NG‐nitro‐l‐arginine methyl ester, 5 mmol/L tetraethylammonium, 0.5 µmol/L tetrodotoxin or 5 µmol/L propranolol did not alter the potency (pD2) or the maximal myorelaxant effect (Emax) of 1,8‐cineole. 3 1,8‐Cineole also significantly decreased the Schultz‐Dale contraction induced by OVA, mainly in preparations from OVA‐sensitized animals submitted to antigen challenge. 1,8‐Cineole decreased tracheal hyperresponsiveness to KCl and carbachol caused by antigen challenge and almost abolished the concentration–response curves to KCl, whereas it had little effect on the concentration–response curves to carbachol. Under Ca2+‐free conditions and in the presence of 10−4 mol/L acetylcholine, neither 1,8‐cineole (6.5 × 10−3 mol/L) nor verapamil (1 × 10−5 mol/L) affected Ca2+‐induced contractions, but they almost abolished Ba2+‐induced contractions. 4 In conclusion, the findings of the present study show that 1,8‐cineole is a tracheal myorelaxant that acts preferentially on contractile responses elicited electromechanically.

Sildenafil decreases rat tracheal hyperresponsiveness to carbachol and changes canonical transient receptor potential gene expression after antigen challenge

Inhibition of type-5 phosphodiesterase by sildenafil decreases capacitative Ca2+ entry mediated by transient receptor potential proteins (TRPs) in the pulmonary artery. These families of channels, especially the canonical TRP (TRPC) subfamily, may be involved in the development of bronchial hyperresponsiveness, a hallmark of asthma. In the present study, we evaluated i) the effects of sildenafil on tracheal rings of rats subjected to antigen challenge, ii) whether the extent of TRPC gene expression may be modified by antigen challenge, and iii) whether inhibition of type-5 phosphodiesterase (PDE5) may alter TRPC gene expression after antigen challenge. Sildenafil (0.1 µM to 0.6 mM) fully relaxed carbachol-induced contractions in isolated tracheal rings prepared from naive male Wistar rats (250-300 g) by activating the NO-cGMP-K+ channel pathway. Rats sensitized to antigen by intraperitoneal injections of ovalbumin were subjected to antigen challenge by ovalbumin inhalation, and their tracheal rings were used to study the effects of sildenafil, which more effectively inhibited contractions induced by either carbachol (10 µM) or extracellular Ca2+ restoration after thapsigargin (1 µM) treatment. Antigen challenge increased the expression of the TRPC1 and TRPC4 genes but not the expression of the TRPC5 and TRPC6 genes. Applied before the antigen challenge, sildenafil increased the gene expression, which was evaluated by RT-PCR, of TRPC1 and TRPC6, decreased TRPC5 expression, and was inert against TRPC4. Thus, we conclude that PDE5 inhibition is involved in the development of an airway hyperresponsive phenotype in rats after antigen challenge by altering TRPC gene expression.

Metabolic acidosis aggravates experimental acute kidney injury

AIMS Ischemia/reperfusion (I/R) injury and metabolic acidosis (MA) are two critical conditions that may simultaneously occur in clinical practice. The result of this combination can be harmful to the kidneys, but this issue has not been thoroughly investigated. The present study evaluated the influence of low systemic pH on various parameters of kidney function in rats that were subjected to an experimental model of renal I/R injury. MAIN METHODS Metabolic acidosis was induced in male Wistar rats by ingesting ammonium chloride (NH4Cl) in tap water, beginning 2 days before ischemic insult and maintained during the entire study. Ischemia/reperfusion was induced by clamping both renal arteries for 45 min, followed by 48 h of reperfusion. Four groups were studied: control (subjected to sham surgery, n=8), I/R (n=8), metabolic acidosis (MA; 0.28 M NH4Cl solution and sham surgery, n=6), and MA+I/R (0.28 M NH4Cl solution plus I/R, n=9). KEY FINDINGS Compared with I/R rats, MA+I/R rats exhibited higher mortality (50 vs. 11%, p=0.03), significant reductions of blood pH, plasma bicarbonate (pBic), and standard base excess (SBE), with a severe decline in the glomerular filtration rate and tubular function. Microscopic tubular injury signals were detected. Immunofluorescence revealed that the combination of MA and I/R markedly increased nuclear factor κB (NF-κB) and heme-oxygenase 1 (HO-1), but it did not interfere with the decrease in endothelial nitric oxide synthase (eNOS) expression that was caused by I/R injury. SIGNIFICANCE Acute ischemic kidney injury is exacerbated by acidic conditions.

Antispasmodic and myorelaxant effects of the flavoring agent methyl cinnamate in gut: Potential inhibition of tyrosine kinase

Methyl cinnamate (MC) is a safe flavoring agent useful to food industry. Although chemically analog to tyrosine kinase inhibitors, there is little information regarding its biological actions. Here, we aimed at assessing the MC effects on gastrointestinal contractility and the putative involvement of tyrosine kinase in the mediation of these effects. Isometric contractions were recorded in rat isolated strips from stomach, duodenum and colon segments. In gastric strips, MC (3-3000 µM) showed antispasmodic effects against carbachol-induced contractions, which remained unchanged by either l-NAME or tetraethylammonium pretreatment and occurred with potency similar to that obtained against contractions evoked by potassium or U-46619. In colon strips, MC was four times more potent than in gastric ones. MC and the positive control genistein inhibited phasic contractions induced by acetylcholine in Ca2+-free medium, an effect fully prevented by sodium orthovanadate. Both MC and genistein decreased the spontaneous contractions of duodenal strips and shortened the time necessary for gastric fundic tissues to reach 50% of maximal relaxation. In freshly isolated colon myocytes, MC decreased the basal levels of cytoplasmic Ca2+, but not the potassium-elicited cytoplasmic Ca2+ elevation. Colon strips obtained from rats subjected to intracolonic acetic acid instillation showed reduced contractility to potassium, which was partially recovered in MC-treated rats. Inhibitory effect of nifedipine against cholinergic contractions, blunted in acetic acid-induced colitis, was also recovered in MC-treated rats. In conclusion, MC inhibited the gastrointestinal contractility with a probable involvement of tyrosine kinase pathways. In vivo, it was effective to prevent the deleterious effects of colitis resulting from acetic acid injury.

Vasorelaxation induced by methyl cinnamate, the major constituent of the essential oil of Ocimum micranthum, in rat isolated aorta

The aim of the present study was to investigate the vascular effects of the E‐isomer of methyl cinnamate (E‐MC) in rat isolated aortic rings and the putative mechanisms underlying these effects. At 1–3000 μmol/L, E‐MC concentration‐dependently relaxed endothelium‐intact aortic preparations that had been precontracted with phenylephrine (PHE; 1 μmol/L), with an IC50 value (geometric mean) of 877.6 μmol/L (95% confidence interval (CI) 784.1–982.2 μmol/L). These vasorelaxant effects of E‐MC remained unchanged after removal of the vascular endothelium (IC50 725.5 μmol/L; 95% CI 546.4–963.6 μmol/L) and pretreatment with 100 μmol/L NG‐nitro‐l‐arginine methyl ester (IC50 749.0 μmol/L; 95% CI 557.8–1005.7 μmol/L) or 10 μmol/L 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one (IC50 837.2 μmol/L; 95% CI 511.4–1370.5 μmol/L). Over the concentration range 1–3000 μmol/L, E‐MC relaxed K+‐induced contractions in mesenteric artery preparations (IC50 314.5 μmol/L; 95% CI 141.9–697.0 μmol/L) with greater potency than in aortic preparations (IC50 1144.7 μmol/L; 95% CI 823.2–1591.9 μmol/L). In the presence of a saturating contractile concentration of K+ (150 mmol/L) in Ca2+‐containing medium combined with 3 μmol/L PHE, 1000 μmol/L E‐MC only partially reversed the contractile response. In contrast, under similar conditions, E‐MC nearly fully relaxed PHE‐induced contractions in aortic rings in a Ba2+‐containing medium. In preparations that were maintained under Ca2+‐free conditions, 600 and 1000 μmol/L E‐MC significantly reduced the contractions induced by exogenous Ca2+ or Ba2+ in KCl‐precontracted preparations, but not in PHE‐precontracted preparations (in the presence of 1 μmol/L verapamil). In addition, E‐MC (1–3000 μmol/L) concentration‐dependently relaxed the contractions induced by 2 mmol/L sodium orthovanadate. Based on these observations, E‐MC‐induced endothelium‐independent vasorelaxant effects appear to be preferentially mediated by inhibition of plasmalemmal Ca2+ influx through voltage‐dependent Ca2+ channels. However, the involvement of a myogenic mechanism in the effects of E‐MC is also possible.

Renal- and calcium-dependent vascular effects of Polybia paulista wasp venom

In the present study, the effects of Polybia paulista venom (PPV) on renal and vascular tissues were investigated. Isolated kidneys perfused with PPV (1 and 3 μg/mL) had increased perfusion pressure, renal vascular resistance, urinary flow, and glomerular filtration rate; and reduced sodium tubular transport. Histological evaluation demonstrated deposits of proteins in Bowmans space and tubular lumen, and focal areas of necrosis. The venom promoted a cytotoxic effect on Madin-Darby canine kidney (MDCK) cells. A significant increase in lactic dehydrogenase levels was observed in response to venom exposure. In isolated mesenteric vascular beds, pressure and vascular resistance augmented in a dose-dependent manner. PPV increased the contractility of aortic rings maintained under basal tension. This contractile response was inhibited when preparations were maintained in Ca2+-free medium. Likewise, verapamil, a voltage-gated calcium channel blocker, also inhibited the contractile response. In this study, phentolamine, a blocker of α-adrenergic receptor blocker, significantly reduced the contractile effect of PPV in the aortic ring. In conclusion, PPV produced nephrotoxicity, which suggests a direct effect on necrotic cellular death in renal tubule cells. The vascular contractile effect of PPV appears to involve calcium influx through voltage-gated calcium channels via adrenergic regulation.