Francisco J.B. Lima

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.

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.

Cytoprotective effect of 1-nitro-2-phenylethane in mice pancreatic acinar cells subjected to taurocholate: Putative role of guanylyl cyclase-derived 8-nitro-cyclic-GMP

The nitroderivative 1-nitro-2-phenylethane (NPE) was recently described as a compound possessing heme-dependent soluble guanylyl cyclase (sGC) stimulating properties in vascular smooth muscle cells. In this study, we tested such pharmacological property of NPE in mice pancreatic acinar cells subjected to the bile salt taurocholate, a type of pathological stimulus that simulates pancreatitis. Here, isolated acinar cells were treated with NPE in order to assess the role of sGC on the detrimental effects induced by taurocholate. NPE reduced taurocholate-elicited Ca(2+) overload, production of reactive oxygen species (ROS), apoptosis, necrosis, and exerted a protective effect against mitochondrial membrane potential (ΔΨm) dissipation. These NPE-induced effects were abolished by pretreatment with ODQ and KT 5823, and after the blockade of nitric oxide (NO) synthase with l-NAME, inhibitors of key components of the sGC pathway. Contrarily to cGMP that alone increased ΔΨm collapse and cell damage, the cytoprotective effect of NPE on ΔΨm and cell necrosis was almost reproduced by 8-nitro-cGMP, a second messenger generated by sGC under oxidative stress conditions. In conclusion, putative sGC stimulation with NPE reveals its cytoprotective profile on pancreatic cells subjected to taurocholate. Moreover, ROS and NO conjunctly appear to drive sGC activity in pancreatic acinar cells to implement an adaptive mechanism in response to oxidative and Ca(2+) stress through 8-nitro-cGMPsynthesis.

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.

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.

Contractile profile of esophageal and gastric fundus strips in experimental doxorubicin-induced esophageal atresia

Esophageal atresia (EA) is characterized by esophageal and gastric motility changes secondary to developmental and postsurgical damage. This study evaluated the in vitro contractile profile of the distal esophagus and gastric fundus in an experimental model of EA induced by doxorubicin (DOXO). Wistar pregnant rats received DOXO 2.2 mg/kg on the 8th and 9th gestational days. On day 21.5, fetuses were collected, sacrificed, and divided into groups: control, DOXO without EA (DOXO-EA), and DOXO with EA (DOXO+EA). Strips from the distal esophagus and gastric fundus were mounted on a wire myograph and isolated organ-bath system, respectively, and subjected to increasing concentrations of carbamylcholine chloride (carbachol, CCh). The isolated esophagus was also stimulated with increasing concentrations of KCl. In esophagus, the concentration-effect curves were reduced in response to CCh in the DOXO+EA and DOXO-EA groups compared to the control group (P<0.05). The maximum effect values (Emax) for DOXO+EA and DOXO-EA were significantly lower than control (P<0.05), but the half-maximal effective concentration (EC50) values were not significantly different when the three groups were compared (P>0.05). In response to KCl, the distal esophagus samples in the three groups were not statistically different with regard to Emax or EC50 values (P>0.05). No significant difference was noted for EC50 or Emax values in fundic strips stimulated with CCh (P>0.05). In conclusion, exposure of dams to DOXO during gestation inhibited the contractile behavior of esophageal strips from offspring in response to CCh but not KCl, regardless of EA induction. The gastric fundus of DOXO-exposed offspring did not have altered contractile responsiveness to cholinergic stimulation.

Gastric contractility in experimental gastroschisis

BACKGROUND/PURPOSE The mechanism of fetal gastric dilation in gastroschisis is controversial. This study was designed to characterize changes in the contractile profile of strips of stomach from rats following experimental gastroschisis. METHODS Pregnant Wistar rats were operated on day 18.5. Fetuses were divided into three groups: gastroschisis (G), sham (S), and control (C). On day 21.5, gastric fundus and antrum strips were obtained and suspended to a force transducer connected to a digital data acquisition system. They were submitted to increasing concentrations of carbachol (CCh) and weighed at the end of each procedure. Frequency and amplitude of each contraction were evaluated. RESULTS Under basal conditions, spontaneous oscillatory contractions of antrum and fundus strips of G, S, and C were similar (P>0.05; ANOVA). However, cumulative concentrations of CCh (0.01-100 μM) produced different effects in all groups and were characterized by a significant increase in amplitude and frequency of spontaneous contractions in antral smooth muscle and a sustained increase in tonus in fundic strips. Upon analysis, no significant difference in frequency or amplitude was noted in antral tissues comparing C to G and to S (P>0.05). No significant contractility difference was noted in fundic smooth muscle (comparing all groups, P>0.05), with the CCh-induced curve following a typical sigmoidal format, dependent on increasing concentrations (P<0.001). CONCLUSIONS Gastric contractile responses to CCh are preserved in experimental gastroschisis. These results do not support the theory that gastric dilation occurs secondary to intestinal inflammation alone.