Tracheal Smooth Muscle Research Articles

Study of the interaction of glutamatergic and nitrergic signalling in conditions of the experimental airways hyperreactivity

BackgroundGlutamatergic and nitrergic system participate in the control of respiratory system functions. It is only little information regarding a possible interaction of both systems in the airways hyperractivity. We investigated the effect of agents modulating the activity of these systems on the experimental ovalbumin-induced airways hyperreactivity as well as on the changes of exhaled nitric oxide (eNO) levels. MethodsWe used the agonists of NMDA receptors – N-methyl-D-aspartic acid (NMDA) and monosodium glutamate (MSG), selective competitive antagonist (DL-2-amino-5-phosphonovaleric acid – AP-5) and selective non-competitive antagonist (dizocilpine – MK-801) of these receptors. We used also non-specific inhibitor of NO synthases Nω-nitro-L-arginine methyl ester (L-NAME). The airways responsiveness to histamine or acetylcholine was evaluated under in vitro conditions. ResultsNMDA administration caused the increase of tracheal smooth muscle response in ovalbumin-induced hyperreactivity to acetylcholine. The effect of MSG was less pronounced. MK-801 as well as AP-5 provoked the decrease of reactivity mainly to acetylcholine in tracheal smooth muscle. We recorded the changes in eNO levels. The activation of NMDA receptor with NMDA or MSG increased eNO levels. The inhibition of NO synthase with L-NAME caused the fall of eNO levels. MK-801 shows (within the group) the more expressive effect in the eNO levels during sensitization than AP-5 group. ConclusionThe results confirm the possibility of NMDA receptors participation in the experimental airways hyperreactivity.

Focal Adhesion Kinase Regulates Collagen I–Induced Airway Smooth Muscle Phenotype Switching

Increased extracellular matrix (ECM) deposition and airway smooth muscle (ASM) mass are major contributors to airway remodeling in asthma. Recently, we demonstrated that the ECM protein collagen I, which is increased surrounding asthmatic ASM, induces a proliferative, hypocontractile ASM phenotype. Little is known, however, about the signaling pathways involved. Using bovine tracheal smooth muscle, we investigated the role of focal adhesion kinase (FAK) and downstream signaling pathways in collagen I-induced ASM phenotype modulation. Phosphorylation of FAK was increased during adhesion to both uncoated and collagen I-coated culture dishes, without differences between these matrices. Nor were any differences found in cellular adhesion. Inhibition of FAK activity by overexpression of the FAK deletion mutants FAT (focal adhesion targeting domain) and FRNK (FAK-related nonkinase) attenuated adhesion. After attachment, FAK phosphorylation increased in a time-dependent manner in cells cultured on collagen I, whereas no activation was found on an uncoated plastic matrix. In addition, collagen I increased in a time- and concentration-dependent manner the cell proliferation, which was fully inhibited by FAT and FRNK. Similarly, the specific pharmacologic FAK inhibitor PF-573228 [6-((4-((3-(methanesulfonyl)benzyl)amino)-5-trifluoromethylpyrimidin-2-yl) amino)-3,4-dihydro-1H-quinolin-2-one] as well as specific inhibitors of p38 mitogen-activated protein kinase (MAPK) and Src also fully inhibited collagen I-induced proliferation, whereas partial inhibition was observed by inhibition of phosphatidylinositol-3-kinase (PI3-kinase) and mitogen-activated protein kinase kinase (MEK). The inhibition of cell proliferation by these inhibitors was associated with attenuation of the collagen I-induced hypocontractility. Collectively, the results indicate that induction of a proliferative, hypocontractile ASM phenotype by collagen I is mediated by FAK and downstream signaling pathways.

Reactive oxygen species induce a Ca2+-spark increase in sensitized murine airway smooth muscle cells

The level of reactive oxygen species (ROS) and the activity of spontaneous, transient, localized Ca2+ increases (known as Ca2+ sparks) in tracheal smooth muscle cells (TSMCs) in an experimental allergic asthma mouse model has not yet been investigated. We used laser confocal microscopy and fluorescent dyes to measure ROS levels and Ca2+ sparks, and we found that both events were significantly increased in TSMCs obtained from ovalbumin (OVA)-sensitized/-challenged mice compared with control mice. ROS levels began to increase in TSMCs after the first OVA challenge, and this increase was sustained. However, this elevation and Ca2+-spark increase was abolished after the administration of the ROS scavenger N-acetylcysteine amide (NACA) for 5days. Furthermore, a similar inhibition was also observed following the direct perfusion of NACA into cells isolated from the (OVA)-sensitized mice that were not treated with NACA. Moreover, we used 0.1-mM caffeine treatment to increase the Ca2+ sparks in single TSMCs and observed cell shortening. In addition, we did not find increases in the mRNA levels of ryanodine (RyRs) and inositol 1,4,5-trisphosphate (IP3Rs) receptors in the tracheal smooth muscle cells of (OVA)-sensitized mice compared with controls. We concluded that ROS and Ca2+ sparks increased in (OVA)-sensitized TSMCs. We found that ROS induces Ca2+ sparks, and increased Ca2+ sparks resulted in the contraction of (OVA)-sensitized TSMCs, resulting in the generation of airway hyperresponsiveness (AHR). This effect may represent a novel mechanism for AHR pathogenesis and might provide insight into new methods for the clinical prevention and treatment of asthma and asthmatic AHR.

Nicotinic Acid Adenine Dinucleotide Phosphate (NAADP) Is a Second Messenger in Muscarinic Receptor-induced Contraction of Guinea Pig Trachea

Nicotinic acid adenine dinucleotide phosphate (NAADP) is increasingly being demonstrated to be involved in calcium signaling in many cell types and species. Although it has been shown to play a role in smooth muscle cell contraction in several tissues, nothing is known about its possible role in tracheal smooth muscle, a muscle type that is clinically relevant to asthma. To determine whether NAADP functions as a second messenger in tracheal smooth muscle contraction, we used the criteria set out by Sutherland for a molecule to be designated a second messenger. We report that NAADP satisfies all five criteria as follows. First, the NAADP antagonist Ned-19 inhibited contractions in tracheal rings and calcium increases in isolated smooth muscle cells induced by the muscarinic agonist carbachol. Second, NAADP increased cytosolic calcium in isolated cells when microinjected and was blocked by Ned-19. Third, tracheal homogenates could synthesize NAADP by base exchange from exogenous NADP and nicotinic acid and metabolize exogenous NAADP to nicotinic acid adenine dinucleotide by a 2'-phosphatase. Fourth, carbachol induced a rapid and transient increase in endogenous NAADP levels. Fifth, tracheal homogenates contained NAADP-binding sites of high affinity. Taken together, these data demonstrate that NAADP functions as a second messenger in tracheal smooth muscle, and therefore, steps in the NAADP signaling pathway might provide possible new drug targets.

Carbon Monoxide Induces Dose‐Dependent Relaxation of Airway Smooth Muscle via cGMP Signaling Pathway

In present study we investigated the effect of endogenous carbon monoxide (CO) on relaxation of rat airway smooth muscle (ASM). Airway segments were excised from 28 day‐old rats. In pre‐constricted tracheal smooth muscle (TSM), electrical field stimulation (EFS) was applied to induce relaxation in absence or presence of heme oxigenase (HO) substrate ‐ hematin (10 μM and 100 μM); HO inhibitor tin protoporphyrin‐IX (SnPP‐IX; 100 μM); nitric oxide (NO) synthase (NOS) inhibitor Nω‐nitro‐L‐arginine methyl ester (L‐NAME; 100 μM); soluble guanylyl cyclase inhibitor 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one (ODQ; 10 μM) and the SnPP‐IX + L‐NAME. All experiments were performed in presence of isoproterenol (100 μM) and indomethacin. Hematin increased relaxant responses of ASM in a dose‐dependent manner. SnPP‐IX and L‐NAME, alone or combined significantly reduced relaxant responses induced by EFS in control and hematin‐preincubated tissues. ODQ also reduced relaxant responses to the level obtained by combining SnPP‐IX and L‐NAME.According to the results we conclude that in rat TSM, CO induces dose‐dependent relaxation and nondrenergic noncholinergic (NANC) inhibitory responses use CO and NO as main molecules to induce relaxation via cGMP signaling pathway.

Phosphorylation of Kv7 potassium channels by protein kinase C in human airway smooth muscle cells

In previous studies, bronchoconstrictor agonists inhibited Kv7 potassium currents in guinea pig airway smooth muscle cells (ASMCs) and the selective Kv7 channel blocker XE991 induced severe bronchoconstriction in human precision cut lung slices (PCLS). However, the mechanism of regulation of airway Kv7 channels is unknown. In the present study, we investigated protein kinase C (PKC)‐induced phosphorylation and inhibition of Kv7 channels. The ability of PKC activator phorbol 12‐myristate 13‐acetate (PMA) to induce bronchoconstriction was examined using human PCLS. We found that PMA induced robust constriction of human airways. Immunoprecipitation and western blot analysis were used to detect phosphorylation of Kv7.5 channels in response to histamine or PMA using human trachealis smooth muscle cells (HTSMCs) infected with a FLAG‐tagged human Kv7.5 adenoviral vector. Both treatments caused increased FLAG‐Kv7.5 phosphorylation. Histamine inhibited Kv7.5 currents in HTSMCs but that effect was abolished when cells were pretreated with the PKC inhibitor calphostin C. Our findings suggest a role of PKC‐induced phosphorylation in the suppression of Kv7 channels in ASMCs. (Funded by Loyola University Chicago.)

Role of large-conductance Ca2+-activated K+ channels and protein kinase G in ketamine-induced isolated tracheal smooth muscle relaxation in rats with asthma

Objective To investigate the role of large-conductance Ga2+-activated K+ (BKCa) channels and protein kinase G (PKG) in ketamine-induced isolated tracheal smooth muscle relaxation in rats with asthma.Methods Healthy Sprague-Dawley rats,weighing 250-300 g,were used in this study.Asthma was induced with egg albumin.Thirty-six tracheal rings of 15 rats in which asthma model was successfully established were randomly divided into 3 groups (n =12 each):ketamine treatment group (group AK),IBTX (BKCa channel blocker) +ketamine treatment group (group AKI),and KT-58232 (PKG inhibitor) + ketamine treatment group (group AKK).Tracheal rings were suspended in an organ bath filled with oxygenated Kreb's solution at 36.5-37.5 ℃.In group AK,the tracheal rings were precontracted with acetyleholine 0.1 mmol/L,and the rings were then exposed to ketamine 0.4 g/L for 15 min.In group AKI,before acetyleholine and ketamine were added to the solution,the rings were pretreated with IBTX 3μmol/L for 30 min.In group AKK,before acetyleholine and ketamine were added to the solution,the rings were pretreated with KT-5823 2μmol/L for 30 min.The tension of rings was measured by using a force-displacement transducer.Results The amplitude of relaxation of isolated tracheal smooth muscle was significantly decreased in groups AKI and AKK as compared with group AK (P ＜ 0.05).Conclusion Ketamine induces isolated tracheal smooth muscle relaxation through activating BKCa channels and PKG signaling pathway in rats with asthma. Key words: Large-conductance calcium-activated potassium channels ; Protein kinases ; Ketamine ; Myocytes, smooth muscle; Trachea; Asthma

Increased mechanical strain imposed on murine lungs during ventilation in vivo depresses airway responsiveness and activation of protein kinase Akt

Continuous positive airway pressure (CPAP) administered to tracheostomized rabbits and ferrets for 4 days or 2 wk suppresses bronchial reactivity in vivo and suppresses airway reactivity in lobes and tracheal segments isolated from these animals. In vitro studies of canine tracheal smooth muscle tissues indicate that mechanical loading suppresses the activation of the growth regulatory kinase, Akt, and that Akt is a negative regulator of smooth muscle differentiation. The transduction of mechanical signals in the tracheal tissues in vitro is mediated by integrin-associated adhesion complexes. To determine whether airway responsiveness and Akt activation are modulated by mechanical loads applied for short time periods to the airways of living animals in vivo, mice were mechanically ventilated for 2 h with high (5 cmH2O) or low (0-1 cmH2O) positive end-expiratory pressure (PEEP) and then ventilated at low PEEP for 30 min. Ventilation of mice with PEEP in vivo for 2 h depressed airway responsiveness to methacholine measured in vivo subsequent to the PEEP treatment. Airway narrowing in vitro in intraparenchymal airways in isolated lung slices and contractile responses of isolated tracheal segments in vitro were suppressed for at least 6 h subsequent to the in vivo exposure to PEEP. Tracheal segments isolated from high PEEP-treated mice exhibited significantly lower levels of Akt activation than tracheae from low PEEP-treated mice. The results indicate that mechanical loads imposed in vivo result in physiological and biochemical changes in the airway tissues after a relatively short 2-h period of in vivo loading.

Efficacy of fexofenadine in isolated rat tracheas

Histamine is an important chemical mediator in both nasal and bronchial inflammation in patients with allergic rhinitis and asthma. The effect of histamine receptor-1 antagonists on nasal mucosa in vivo is well known, however, the effect on tracheal smooth muscle has rarely been explored. The purpose of this study was to determine the effects of fexofenadine on isolated tracheal smooth muscle in vitro. Six tracheal strips were used for each experiment, and one untreated strip served as a control. We examined the effectiveness of fexofenadine on isolated rat tracheal smooth muscle by testing the effect on: 1) tracheal smooth muscle resting tension; 2) contraction caused by 10E-6 M methacholine as a parasympathetic mimetic; and 3) electrically induced tracheal smooth muscle contractions. The results indicated that addition of methacholine caused the trachea to contract in a dose-dependent manner. The addition of fexofenadine at a dose of 10E-4 M elicited a significant relaxation response compared to 10E-6 M methacholine-induced contraction. There were no detectable changes in the peak tension of electrical field stimulation-induced contractions in the fexofenadine group. High concentrations of fexofenadine had an anti-cholinergic effect. In addition to diminishing histamine-mediated allergic symptoms, fexofenadine may have a potentially therapeutic implication in alleviating asthma-related symptoms due to reducing methacholine-induced contractions of tracheal smooth muscle though these aspects were not studied.

The effect of hydroethanol extract of Achillea millefolium on β-adrenoceptors of guinea pig tracheal smooth muscle

Different pharmacological effects of Achillea millefolium including its relaxant effect on smooth muscle have been shown previously. In the present study the stimulatory effect of the plant extract on β-adrenoceptor of tracheal muscle was examined in order to investigate one possible mechanism for its observed relaxant effect. Effect of three concentrations of hydroethanol extract, 10 nM propranolol, and saline on β-adrenoceptor was tested in two experimental groups including; nonincubated tracheal smooth muscles (group 1) and incubated tracheal smooth muscle with chlorpheniramine (group 2). Concentration response curves to isoprenaline were performed in precontracted tracheal smooth muscle in the presence of the extract, propranolol and saline. Values of EC50 and CR-1 were measured. Leftward shifts in isoprenaline curves were observed in the presence of medium and high concentrations of the extract compared with saline in both groups. The values of EC50 obtained in the presence of medium and high concentrations of the extract only in group 1 were nonsignificantly lower than that of saline. The values of CR-1 obtained in the presence of all concentrations of the extract in both groups were negative and significantly different with that of propranolol. The results indicated a small stimulatory effect of the extract on ß2-adrenoceptors.

The effect of hydro-ethanolic extract of Achillea millefolium on muscarinic receptors of guinea pig tracheal smooth muscle

Objective:To investigate one possible mechanism for the observed relaxant effect of A. millefolium (Achillea millefolium), in the present study the inhibitory effect of the extract of this plant on muscarinic receptors was examined.Materials and Methods:The effects of three concentrations of aqueous-ethanolic extract, 10 nM atropine, and saline on muscarinic receptors were tested in three conditions: In non incubated tracheal smooth muscle (group 1), tracheal chain incubated with propranolol and chlorpheniramine (group 2), and the one incubated with propranolol (group 3).Results:The EC50 obtained in the presence of all three concentrations of the extract were significantly higher compared to saline in groups 2 and 3 (P < 0.001and P < 0.01 in group 2 and 3 respectively). The EC50 obtained in the presence of all concentrations of the extract in group 2 were significantly improved compared to groups 1 and 3 (P < 0.05 to P < 0.001). The maximum responses to methacholine in presence of only the higher concentration of the extract (0.8mg/ml) was significantly lower than that of saline in groups 1 (P < 0.05). There was neither significant difference between slopes of methacholine-response curves obtained in the presence of different concentrations of the extract and that of saline nor between the three groups. The values of (CR-1), obtained in the presence of all concentrations of the extract, were significantly lower compared to atropine in the first group but were not significantly different in other groups. The values of (CR-1) obtained in the presence of all concentrations of the extract were significantly improved in groups 2, compared to groups 1 and 3 (P < 0.05 to P < 0.001).Conclusion:These results showed an inhibitory effect for the extract of A. millefolium on muscarinic receptors of tracheal smooth muscle. A histamine (H1) receptor blockade was also suggested for the extract.

Evaluation of Thioperamide Effects Using Rat's Trachea Model

ObjectivesThioperamide is used as an antagonist to the histamine H3 receptor. During administration of the drug, the trachea may be affected via nasal or oral inhalation. This study was to determine the effects of thioperamide on the trachea of rats in vitro.MethodsWe tested the effectiveness of thioperamide on isolated rat trachea submersed in Kreb's solution in a muscle bath. Changes in tracheal contractility in response to the application of parasympathetic mimetic agents were measured. The following assessments of thioperamide were performed: 1) effect on tracheal smooth muscle resting tension; 2) effect on contraction caused by 10-6 M methacholine as a parasympathetic mimetic; 3) effect of the drug on electrically-induced tracheal smooth muscle contractions.ResultsThioperamide induced a significant relaxation response at a preparation concentration up to 10-4 M. The drug also inhibited the electrical field stimulation induced spike contraction. However, thioperamide alone had a minimal effect on the basal tension of the trachea at increasing concentrations.ConclusionThe study indicated that high concentrations of thioperamide might actually antagonize cholinergic receptors and block parasympathetic function of the trachea.

Bronchodilatory effect of ethanolic extract of the leaves of Nyctanthes arbortristis

Background:Nyctanthes arbortristis has been used in traditional medicine for the treatment of asthma and cough.Aim:In this study, bronchodilatory effect of ethanolic extract of the N. arbortristis was investigated under in vitro conditions. The concentration–response curve of the tracheal smooth muscle (TSM) to histamine was recorded in presence or absence of ethanolic extract and Nω-nitro-l-arginine methyl ester (L-NAME). Dose–response effect of ethanolic extract on pre-constricted tissues was investigated. The ethanolic extract inhibited the histamine-induced maximum contractile responses of TSM (P < 0.001). Ethanolic extract also cause dose-dependent relaxation of TSM. These effects were reversed by L-NAME. Phytochemical analysis showed the presence of typical plant constituents. These results suggest the possible use of extract of the leaves of N. arbortristis as a bronchodilator in therapeutic treatment of asthma.

Effects of selenoprotein P on the contraction and relaxation of the airway smooth muscle

Selenoprotein P (SeP) not only represents the major selenoprotein in plasma, but also provides more than 50% of the total plasma selenium. However, there is no report concerning the direct action of selenium or selenium-containing compounds on the contraction and relaxation of the airway smooth muscle. Therefore, we investigated the effects of SeP and sodium selenite (SS) on the indirectly induced contraction and relaxation of the cat bronchi, and gel contraction of cultured bovine tracheal smooth muscle cells (BTSMC) induced by ATP. In the present results, SeP or SS suppressed the amplitude of twitch-like contractions of cat bronchiole without affecting the non-adrenergic and non-cholinergic (NANC) relaxations evoked by electrical field stimulation. SeP also suppressed the ATP-induced gel contraction of BTSMC. These results suggest that SeP suppresses the amplitude of twitch-like contraction of cat bronchiole by acting directly on the bronchiolar smooth muscle.

Spasmolytic effect of novel antagonists of the adenosine A2 receptor in isolated guinea pig tracheal smooth muscle

We have investigated the relaxation effect of nine novel adenosine A2B antagonists on precontracted guinea pig tracheas by carbachol (10μM). Isometric contractions of isolated guinea pig tracheas were recorded at 2g resting tension; and carbachol dose-response curves were performed. EC50 and IC50 values were identified by plotting cumulative concentration-response curve and pD2 values were calculated. Theophylline was able to relax carbachol precontracted tracheas on 10−3 M conc. The relaxant effect of 1×10−4M compound 5 (ARR-1) was found highly significant as compared to theophylline (P<0.001), whereas compound 6 (ARR-2) showed equipotent effect in a concentration dependent manner. However, other compounds were not able to attenuate contractions induced by carbachol significantly as compared to theophylline. Compound 5 (ARR-1) may be an important new compound in testing the current hypotheses proposing a therapeutic role for an adenosine antagonist in asthma. This effect might be attributed to inhibition of cAMP through adenyl cyclase enzyme. The results exclude an important contribution of adenosine A2B receptors.

Trans-Caryophyllene, a Natural Sesquiterpene, Causes Tracheal Smooth Muscle Relaxation through Blockade of Voltage-Dependent Ca2+ Channels

trans-Caryophyllene is a major component in the essential oils of various species of medicinal plants used in popular medicine in Brazil. It belongs to the chemical class of the sesquiterpenes and has been the subject of a number of studies. Here, we evaluated the effects of this compound in airway smooth muscle. The biological activities of trans-caryophyllene were examined in isolated bath organs to investigate the effect in basal tonus. Electromechanical and pharmacomechanical couplings were evaluated through the responses to K+ depolarization and exposure to acetylcholine (ACh), respectively. Isolated cells of rat tracheal smooth muscle were used to investigate trans-caryophyllene effects on voltage-dependent Ca2+ channels by using the whole-cell voltage-clamp configuration of the patch-clamp technique. trans-Caryophyllene showed more efficiency in the blockade of electromechanical excitation-contraction coupling while it has only minor inhibitory effect on pharmacomechanical coupling. Epithelium removal does not modify tracheal smooth muscle response elicited by trans-caryophyllene in the pharmacomechanical coupling. Under Ca2+-free conditions, pre-exposure to trans-caryophyllene did not reduce the contraction induced by ACh in isolated rat tracheal smooth muscle, regardless of the presence of intact epithelium. In the whole-cell configuration, trans-caryophyllene (3 mM), inhibited the inward Ba2+ current (IBa) to approximately 50% of control levels. Altogether, our results demonstrate that trans-caryophyllene has anti-spasmodic activity on rat tracheal smooth muscle which could be explained, at least in part, by the voltage-dependent Ca2+ channels blockade.

P46 The impact of hydrogen sulfide on ovalbumin-sensitized canine trachealis smooth muscle

Background Hydrogen sulfide has been found to relax vascular smooth muscle, to function as a neuroprotectant molecule in the CNS in an extensive corpus of literature. However, very little has been reported regarding the effect of H2S on airway smooth muscle. Just as relaxation of vascular smooth muscle would be a benefit for hypertensive subjects, relaxation of airways smooth muscle would be helpful for asthmatic subjects. Methods Conventional force–velocity properties of the sensitized canine trachealis smooth muscle were measured with a muscle servo-motor system. Electrical field stimulation was provided to muscle strips in vitro. Results In the sensitized trachealis H2S reduced both the shortening and the velocity of shortening of the strip. The reduction in shortening was due to prolongation of the preceding relaxation. Analysis of the derivatives of shortening indicated that cross-bridge activity under H2S increased spontaneously during relaxation. Biochemical studies showed a spontaneous increase in both myosin light-chain phosphorylation and intracellular calcium as well as a decrease in myosin light-chain phosphatase activity. Further, both endogenous H2S synthesizing enzymes were reduced. Conclusion These results suggest a potential therapeutic pathway for asthma relief.

The achilles heel in melatonin: asthma.

Asthma is a clinical syndrome characterized by chronic airway inflammation, airway responsiveness, and expiratory airflow limitation. Nocturnal symptoms and decreases in lung function are common aspects of the asthma clinical syndrome. Nocturnal symptoms also appear to be associated with asthma-related mortality. In addition to its importance to the regulation of human circadian rhythms, an accumulating body of evidence also suggests that melatonin is also involved in the regulation of smooth muscle tone. For this reason, this study aimed to evaluate contraction and relaxation responses in tracheal smooth muscle rings obtained from rats treated with melatonin. Following administration of melatonin (50mg/kg/day) at the same time every day for 6 weeks, in vitro organ bath experiments were performed with rat tracheal preparations exposed to contractile (acetycholine and serotonin) and relaxant (theophylline and papaverine) agents. Melatonin treatment strengthened contraction responses, but did not affect relaxation responses in rat tracheal preparations. We think that melatonin might play a role in the pathogenesis of nocturnal asthma. Therefore, clinicians should be aware of the importance of melatonin to nocturnal exacerbation of asthma symptoms and alert asthmatic patients that use exogenous melatonin supplementation of its potential negative effects.

Quantitatively analyze composition principle of Ma Huang Tang by structural equation modeling

Ethnopharmacological relevanceIn Chinese classic formulas, Ma Huang Tang (MHT), composed of Ephedra, Cassia twig, Bitter apricot kernel and Prepared licorice, has been widely used to treat cold, influenza, acute bronchitis, bronchial asthma and other pulmonary diseases. However, there is no quantitative interpretation about composition principle of MHT as well as other Chinese compound prescriptions. This study was aimed using structural equation modeling (SEM) to decipher ‘monarch, minister, assistant and guide’ which is the unique and integrated composition principle of Chinese compound recipes, by taking MHT for instance. Materials and methodsSixteen prescriptions of different dose ratios were combined orthogonally from four herbal drugs of MHT, then their diaphoretic, antispasmodic and analgesic effects were assessed by the indicators of the rat sweating point number, the spasmolysis percentage of guinea pig trachea and the murine writhing number, respectively. Basing on SME, the systematology analysis method to complex causality, path diagrams for herbal drugs were drawn with the Amos software and the relationships of the four herbal ingredients and therapeutic effects were measured. ResultsSixteen recipes induced SD rats sweating, remitted spasm of guinea pig trachea smooth muscle, and relieved ICR mouse pain due to acetic acid in comparison with animal model group or normal control groups. Three different SME models were specified and the relevant relationship was analyzed. According to the results of measured standardized path coefficients, Ephedra exerts the greatest contribution to the integral potency, so it acts as the monarch drug in MHT; Cassia twig is slightly weakly effective than Ephedra, and has the most significant interaction with Ephedra, which shows that it is the minister drug; the direct effects of Bitter apricot kernel and Prepared licorice on the integral potency are non-significant, while these two drugs have very significant synergetic effect with Ephedra or Cassia twig, thus they can be interpreted as subordinate drugs to strengthen the therapeutical effects of the monarch and minister drugs; the higher interaction values of Bitter apricot kernel suggest that it is the assistant drug, and Prepared licorice is the guide drug with lower values. ConclusionSME can be used to quantitatively analyze the composition principle of Chinese compound prescriptions like MHT, which demystifies the ancient and classical system theory of traditional Chinese medicine from a totally new viewpoint.

Substance P Regulates Environmental Tobacco Smoke-Enhanced Tracheal Smooth Muscle Responsiveness in Mice

Environmental tobacco smoke (ETS) is an environmental trigger that leads to airway inflammation and airway hyperresponsiveness (AHR) in susceptible individuals and animals, but the underlying mechanism is not fully understood. Substance P (SP) release from sensory nerve fibers has been linked to AHR. The present experiments characterize the role of SP in tracheal smooth muscle on ETS-increased airway responses. The mice were exposed to either sidestream tobacco smoke (SS), a surrogate to ETS, or filtered air (FA) for 1 day or 5 consecutive days. Contractions of tracheal smooth muscle to SP and electrical field stimulation (EFS) were not significantly altered in 1 of day SS-exposed mice. However, 5 of days SS exposure significantly increased airway smooth muscle contractions to SP and EFS. Administration of CP-99994, an antagonist of the neurokinin (NK)1 receptor, attenuates the SS exposure-enhanced tracheal smooth muscle responses to EFS. Furthermore, the immunohistochemistry showed that SP nerve fibers were increased in tracheal smooth muscle after 5 of days SS exposure. These results suggest that the increased SP production may contribute to SS-enhanced smooth muscle responsiveness in mice trachea.