The influence of L-NAME on iNOS expression and markers of oxidative stress in allergen-induced airway hyperreactivity.

Nitric oxide (NO) effects in airways are influenced by the activity of NO-synthase isoforms and NO metabolism. Inducible NO-synthase (iNOS), which produces large amounts of NO, is active during the inflammatory process. NO quickly reacts, producing reactive oxygen species (ROS). In this study we attempted to detect the expression of iNOS and markers of ROS in the airway hyperreactivity (AHR) condition. The study was performed in guinea pigs, divided into four groups. Two groups were treated with the non-selective inhibitor of NO-synthase L-NAME. The other two groups were used as controls. Exhaled NO was monitored in vivo, AHR was assessed both in vivo and in vitro, and the expression of iNOS in lung homogenate, and oxidative stress markers were measured in the venous blood. L-NAME significantly affected the AHR only in in vitro condition, blocked the expression of iNOS in control but not in sensitized animals, and decreased the level of exhaled NO. The results concerning the oxidative stress markers are equivocal. The study confirmed that NO is involved in the regulation of AHR; the effects being mediated via iNOS and ROS activity.

L-arginine supplementation and experimental airway hyperreactivity.

The interest in L-arginine metabolism was triggered primarily by the discovery of nitric oxide (NO) synthesis in mammals and its remarkable biological roles. The real role of L-arginine in the airway hyperreactivity (AHR) has not been established yet. Therefore, we studied whether supplementation of L-arginine can influence the experimental AHR evoked by two different triggers - allergen and exogenous irritant (toluene vapours). Male TRIK strain guinea pigs were used in the study. We used two patterns of pretreatment with L-arginine in vivo, short- and long-term, in a dose of 300 mg/kg administered i.p., after which we studied reactivity of airway smooth muscles in vitro. Pretreatment with L-arginine for 3 days decreased the airway smooth muscle reactivity induced by toluene vapour, whereas pretreatment for 17 days was without any additional effect on smooth muscle reactivity. The short-term pretreatment in ovalbumin-induced hyperreactivity caused an increase in airway smooth muscle reactivity to lower concentrations of both bronchoconstrictors. On the other side, this pretreatment significantly decreased smooth muscle reactivity to high concentrations of both bronchoconstrictors. Supplementation of L-arginine resulted in a modification of the airway smooth muscle response. The effect of supplementation was different depending on the AHR trigger, airway region and pretreatment duration. The results also underscore the importance of an optimal L-arginine level for the control of bronchial tone.

Arginase in the airways hyperreactivity.

BACKGROUND: The interest of arginase action is increasing because limitation of L-arginine bioavailability by arginase for NO synthesis via constitutive NOS can contribute to airway hyperreactivity. OBJECTIVES: We investigated the effect of intervention in the arginase activity in guinea pig model of experimental ovalbumin-induced airway hyperreactivity. METHODS: We analysed the response of tracheal and lung tissue smooth muscle strips to histamine or acetylcholine after in vitro administration of arginase in a dose of 75 UI or after administration of the non-selective inhibitor of arginase N(omega)-hydroxy-L-arginine (NOHA) in a dose of 5 and 10 micromol. We used as well as the incubation of strips with the precursor of NO synthesis L-arginine in a dose of 10(-4) mol/l together with NOHA. RESULTS: We did not find any significant differences in the reactivity of tracheal and lung tissue smooth muscle if we applied arginase in a dose of 75 UI in vitro. NOHA in a dose of 5 a 10 micromol induced the decrease of tracheal and lung tissue smooth muscle reactivity overall. The decrease of the contraction amplitude was dose-dependent. The supplementation of NO synthesis precursor L-arginine in a dose of 10(-4) mol/l together with NOHA intensified the decrease of the airways reactivity induced by an arginase inhibition. CONCLUSION: The results suggest that arginase is involved in the control of airways bronchomotoric tone and therefore modulation of arginase activity could be a useful tool for airway smooth muscle tone control in clinical conditions (Fig. 7, Ref. 33). Full Text (Free, PDF) www.bmj.sk.

Relation of L-arginine to airway hyperreactivity.

The deficiency or the decrease in the bioavailability in basic substrate for nitric oxide synthesis - L-arginine can be one of factors contributing to the airway hyperreactivity. We studied the influence of L-arginine supplementation on the experimental airway hyperreactivity induced in guinea pigs by exposure to toluene vapours. L-arginine was administered before exposure in a dose of 300 mg/kg b.w. intraperitoneally during 3 or 17 days. After that the airway reactivity changes to histamine or acetylcholine were studied in in vitro conditions. In addition to that the tissue strips from exposed animals were incubated with L-arginine in concentration 10(-4) mol/l. The administration of L-arginine during 3 days decreased the airway reactivity increased by irritant exposure. We recorded the decrease in the airway reactivity in animals with bronchial hyperreactivity after incubation of tissue strips with L-arginine, too. The pre-treatment of animals with L-arginine during 17 days did not affect the airway smooth muscle reactivity in larger extent. The exogenous administration of L-arginine resulted in a protective effect under the conditions of experimental airway hyperreactivity. The effect of supplementation was different depending on airway level and pre-treatment duration. The results refer to the importance of optimal L-arginine level for the control of bronchomotoric tone.

Effect of NO-synthase and arginase inhibition in airway hyperreactivity.

BACKGROUND: The decreased L-arginine bioavailability, the basic substrate for nitric oxide synthesis, can be one of the factors contributing to the airways hyperreactivity. OBJECTIVES: We investigated the effect of various inhibitors of the enzyme activities utilizing L-arginine in a guinea pig model of experimental ovalbumin-induced airway hyperreactivity. METHODS: We used the in vivo pre-treatment with non-specific inhibitor of NO synthase No-nitro-L-arginine metylester (L-NAME) and relatively specific inhibitor of inducible NO synthase--aminoguanidine. Inhibitors were administered in one-shot (on the 14th day, 30 minutes before the inhalation of ovalbumin) or in a long-time regime (during the whole period of sensibilization by ovalbumin--14 days). We administered the inhibitor of arginase Nomega-hydroxy-L-arginine (NOHA) to the tracheal and lung tissue smooth muscle strips from sensibilized animals. RESULTS: We observed an increase in the tracheal smooth muscle response to histamine in animals that received an inhalation dose of L-NAME (40 mg/kg b.w.) or aminoguanidine (50 mg/kg b.w.) 30 minutes before the inhalation of ovalbumin but did not evoke any significant difference in the reactivity of lung tissue smooth muscle. Tracheal smooth muscle responded with enhanced contraction amplitude to histamine after chronic pre-treatment with L-NAME or aminoguanidine. The inhibition of arginase with NOHA in vitro decreased the tracheal and lung tissue smooth muscle reactivity to histamine. CONCLUSION: The results suggest that NO-synthase isoforms as well as arginase are involved in the production of NO and in the control of bronchomotoric tonus (Fig. 4, Tab. 2, Ref. 31). Full Text (Free, PDF) www.bmj.sk.

Effect of nitric oxide synthases inhibitors on exogenous irritant-induced bronchial hyper-reactivity in guinea pigs.

Nitric oxide (NO) is an important endogenous mediator involved in many biological functions in both physiological and pathological conditions. Many of studies suggest that high level of NO may play a role in the pathogenesis of various diseases including respiratory diseases with bronchial hyper-reactivity (BHR). The aim of our study was to examine the relationship between NO production and BHR. The reactivity of tracheal and lung tissue smooth muscle to histamine and acetylcholine was measured in vitro in male guinea pigs pre-treated with NO synthase (NOS) inhibitors. The drugs were administered in vivo during either 3 or 17 days. Furthermore, the animals were exposed in vivo to the toluene vapours after administration of agents. NOS inhibitors showed mainly beneficial effect in the presented study. They decreased the hyper-reactivity of the tracheal and lung tissue smooth muscle evoked by toluene. The decrease was dependent on the duration of their administration and on the type of inhibitor. Short-term administration of inhibitors was more effective than long-term one. A more significant effect was recorded after the pre-treatment with non-selective inhibitor L-NAME. The results showed possible participation of constitutive forms of NOS in the BHR.

Interaction between nitric oxide and prostanoids in the respiratory system.

BACKGROUND: Prostaglandins and nitric oxide are important mediators of different physiological and pathophysiological processes. So far, is not characterized clearly their relationship in the conditions of airways hyperreactivity. OBJECTIVES: We tried to detect the relationship of interaction NOS-COX in conditions of exogenous irritant-induced experimental bronchial hyperreactivity. METHODS: Male guinea pigs were used in the experiment. Animals received agent that inhibits COX activity--diclofenac in a dose of 10 mg/kg i.m. or direct NO donor--molsidomine in a dose of 2 mg/kg i.p. Agents were administered singly (10 days) or in combination (last 3 days). Then animals were exposed to the toluene vapours for two hours over each of three consecutive days to provoke hyperreactivity. Then we recorded the reactivity changes to cumulative doses of histamine or acetylcholine (10(-8)-10(-3) mol/I). RESULTS: The administration of NO donor (10 days) and consecutive COX inhibition (3 days) increased the reactivity of both observed preparations in comparison to agents administered single. COX inhibition during 10 days and consecutive treatment with NO donor (3 days) evoked different changes of tracheal smooth muscle and lung tissue smooth muscle response but had more beneficial effect on the airways reactivity on the whole. CONCLUSION: It is possible to suppose some participation of both followed enzymatic systems and theirs interaction in our experimental conditions since airways reactivity was affected the by used agents (Fig. 7, Ref. 32).

Inhibition of guanylyl cyclase in the airways hyperreactivity.

BACKGROUND: The majority of nitric oxide (NO) effects in the respiratory system are mediated via the stimulation of soluble guanylyl cyclase with subsequent generation of the second messenger--cyclic guanosine monophosphate (cGMP). OBJECTIVES: We were interested in the effect of non-selective soluble guanylyl cyclase inhibitor--methylene blue on the exogenous irritant-induced bronchial hyperreactivity. METHODS: Male guinea pigs were used in the experiment. The animals received non-selective soluble guanylyl cyclase inhibitor--methylene blue in a dose of 50 or 100 mg/kg b.w. 30 minutes before inhalation of the exogenous irritant--toluene vapours. The toluene exposition lasted three consecutive days during two hours in in vivo conditions. The monitoring of tracheal and lung tissue strips reactivity changes was carried out in in vitro conditions. The brochoconstrictor mediators histamine and acetylcholine in the cumulative doses (10(-8)-10(-3) mol/l) were used in the experiment. RESULTS: The methylene blue pretreatment induced the decrease of tracheal and lung tissue smooth muscle contraction amplitude increased by exogenous irritant--toluene. We recorded different smooth muscle response depending on the doses of inhibitor. Methylene blue in a dose of 50 mg/kg b.w. affected mainly tracheal smooth muscle, in a dose of 100 mg/kg b.w. mainly the lung tissue. CONCLUSION: The interaction between nitric oxide and soluble guanylyl cyclase can be important for bronchial reactivity changes. The changes depended on the dose of inhibitor and on the type of respiratory system tissue (trachea, lung). We can summarise that changes of the airways reactivity are not only evoked by NO/cGMP pathway but probably by any other mechanisms (Fig. 5, Ref. 26).

Mucolytics and antioxidant activity.

We investigated effects of the mucolytics ambroxol and N-acetylcysteine on airways reactivity evoked by histamine in guinea pigs exposed to toluene vapors. We did not find significant changes in reactivity of tracheal smooth muscle from animals treated with mucolytics compared to the control group. However, the administration of ambroxol and N-acetylcysteine caused a significant decrease in lung tissue reactivity. The effect of ambroxol was more pronounced after intraperitoneal injection than after inhalation, while N-acetylcysteine was only effective after inhalation. The protective effects of mucolytics in the lung tissue may be due to their antioxidant activity together with other mechanisms.

Molecular pathways of endoplasmic reticulum dysfunctions: possible cause of cell death in the nervous system.

This review summarizes recent information on the role of calcium in the process of neuronal injury with special attention to the role of calcium stores in the endoplasmic reticulum (ER). Experimental results present evidence that ER is the site of complex processes such as calcium storage, synthesis and folding of proteins and cell response to stress. ER function is impaired in many acute and chronic diseases of the brain which in turn induce calcium store depletion and conserved stress responses. Understanding the mechanisms leading to ER dysfunction may lead to recognition of neuronal protection strategies.

Effects of hyperoxia and allergic airway inflammation on cough reflex intensity in guinea pigs.

Toxic influence of high oxygen concentration on pulmonary function and structures has been known for many years. However, the influence of high oxygen concentration breathing on defensive respiratory reflexes is still not clear. In our previous experiments, we found an inhibitory effect of 100 % oxygen breathing on cough reflex intensity in healthy guinea pigs. The present study was designed to detect the effects of hyperoxia on cough reflex in guinea pigs with allergic airway inflammation. In the first phase of our experiment, the animals were sensitized with ovalbumin. Thirty-two sensitized animals were used in two separate experiments according to oxygen concentration breathing: 100 % or 50 % oxygen for 60 h continuously. In each experiment, one group of animals was exposed to hyperoxia, another to ambient air. The cough reflex was induced both by aerosol of citric acid before sensitization, then in sensitized animals at 24 h and 60 h of exposition to oxygen/air in awake animals, and by mechanical stimulation of airway mucosa in anesthetized animals just after the end of the experiment. In contrast to 50 % oxygen, 100 % oxygen breathing leads to significant decrease in chemically induced cough in guinea pigs with allergic inflammation. No significant changes were present in cough induced by mechanical stimulation of airways.

Objective assessment of cough suppressants under normal and pathological experimental conditions.

The influences of the antitussive activity of glaucine were studied in 56 non-anaesthetized cats under normal and pathological conditions. Cough was induced by mechanical stimulation of the airways with a nylon fibre. The authors found that if glaucine was administered at a dose of 5.0, 7.5 and 10.0 mg/kg b.w., i.p., it evoked statistically significant suppression on single cough components. After inflammation of the airways was induced with unsoluted croton oil, no decrease in antitussive activity of glaucine could be observed, according to the number of cough efforts, frequency, intensity of maximal cough effort, and intensity of cough attack during expiration. Glaucine used under such conditions was not found to be powerful enough to suppress either the intensity of maximal cough effort or the intensity of cough attack during inspiration. The antitussive effect of glaucine was stronger under pathological conditions (Staphylococcus-induced inflammation). The antitussive effect of glaucine was approximately the same as with codeine if administered in equal doses.

Time course of ischemia/reperfusion-induced oxidative modification of neural proteins in rat forebrain.

Time course of oxidative modification of forebrain neural proteins was investigated in the rat model of global and partial cerebral ischemia/reperfusion. Animals were subjected to 4-vessel occlusion for 15 min (global ischemia). After the end of ischemia and at different reperfusion times (2, 24 and 48 h), lipoperoxidation-dependent and direct oxidative modification neural protein markers were measured in the forebrain total membrane fraction (tissue homogenate). Ischemia itself causes significant changes only in levels of tryptophan and bityrosine fluorescence when compared to controls. All tested parameters of protein modification altered significantly and were maximal at later reperfusion stage. Content of carbonyl group in re-flow period steadily increased and culminated at 48 h of reperfusion. The highest increase in the fluorescence of bityrosines was detected after 24 h of reperfusion and was statistically significant to both sham operated and ischemic groups. The changes in fluorescence intensity of tryptophan decreased during a reperfusion time dependent manner. Formation of lysine conjugates with lipoperoxidation end-products significantly increased only at later stages of reperfusion. Total forebrain membranes from animals subjected to 3-vessel occlusion model to 15 min (partial ischemia) show no altered content of oxidatively modified proteins compared to controls. Restoration of blood flow for 24 h significantly decreased only fluorescence of aromatic tryptophan. Partial forebrain ischemia/reperfusion resulted in no detectable significant changes in oxidative products formation in extracerebral tissues (liver and kidney) homogenates. Our results suggest that global ischemia/reperfusion initiates both the lipoperoxidation-dependent and direct oxidative modifications of neural proteins. The findings support the view that spatial and temporal injury at later stages of ischemic insult at least partially involves oxidative stress-induced amino acid modification. The results might have important implications for the prospective post-ischemic antioxidant therapy.

[Antitussive action of extracts and polysaccharides of marsh mallow (Althea officinalis L., var. robusta)]. / Antitussive Wirkung des Extraktes und der Polysaccharide aus Eibisch (Althaea officinalis L., var. robusta).

The complex extract and the polysaccharide isolated from the roots of marsh mallow were tested for antitussive activity in unanaesthetized cats of both sexes. Cough was elicited by mechanical stimulation of laryngopharyngeal and tracheobronchial mucous area of the respiratory system with a Nylon fibre (diameter 0.35 mm). Cough was evaluated on the basis of the changes in lateral tracheal pressure. The polysaccharide and the complex extract were administered p.o. in a dose of 50 and 100 mg/kg b.w., respectively. The efficiency of the mentioned compounds was compared with the cough-suppressing effect of drugs belonging to the non-narcotic antitussics. The results of the experiments showed that administration of the polysaccharide led to a statistically significant decrease of the number of cough efforts both from laryngopharyngeal and tracheobronchial areas of the the respiratory system. The polysaccharide in a dose of 50 mg/kg b.w. was as effective in inhibition of the cough reflex as Sirupus Althaeae in a dose of 1000 mg/kg b.w. and more effective than prenoxdiazine in a dose of 30 mg/kg b.w. However, the cough-suppressing effect of the polysaccharide was lower than that of dropropizine. The extract was less effective than the polysaccharide.

Effects of irritants on airways reactivity.

The authors observed reactivity changes of airways smooth muscles after exposure of experimental animals to toluene and hyperoxia. The pretreatment with antioxidants did not prevent these changes.

Nitric oxide and airway reactivity.

BACKGROUND: Nitric oxide is a neurotransmitter of the inhibitory nonadrenergic noncholinergic mediation in the respiratory system. Its participation in the regulation of airways functions is determined by its level in the organism. MAIN PURPOSE: We examined participation of nitric oxide in the changes of the airway reactivity evoked by toluene exposure as the source of the free radicals. The changes of nitric oxide level in the organism were evoked by administration of its indirect donor isosorbide dinitrate. Thiol groups were provided by administration of antioxidative mucolytic N-acetylcysteine. METHODS: Used drugs--isosorbide dinitrate (5 mg/kg b.w.) and N-acetylcysteine (300 mg/kg b.w.) were administered intraperitoneally or by inhalation 30 minutes before each exposure to the toluene vapours. The control group was not treated with drugs. After toluene exposure (2 hours in each of 3 consecutive days) tracheal and lung strips smooth muscle reactivity to histamine was observed under in "in vitro" conditions. RESULTS: The administration of isosorbide dinitrate decreased especially the lung strip smooth muscle reactivity to histamine. We revealed more expressive effect of the pretreatment with intraperitoneally administered isosorbide dinitrate in the comparison with inhalation. Simultaneous pretreatment with N-acetylcysteine intensified beneficial effect of isosorbide dinitrate probably by increasing of the intracellullary level of thiols. CONCLUSION: In our experimental conditions possible participation of nitric oxide in changes of airways smooth muscle reactivity after exposure to the toluene follows from results, as well as the importance of thiol groups for the activity of its indirect donors. (Fig. 6, Tab. 3, Ref. 35.)

[Antioxidants in the respiratory system]. / Antioxidanty v respiracnom systéme.

BACKGROUND: Reactive oxygen species can participate in the airways reactivity changes after oxidants. The authors have observed an increase in airways reactivity after an exposure to toluene in guinea-pigs and cats in previous experiments. There literature data provide information on the prevention or the delay of free radical damage by antioxidants. MAIN PURPOSE: The aim of our study was to evaluate the effect of nonenzymatic antioxidants--vitamin C and vitamin E on the airways reactivity changes after the exposure to toluene vapours. METHODS: After a one-month-lasting pretreatment with 500 mg/kg/day vitamin C and 50 mg/kg/day vitamin E the guinea-pigs were exposed to toluene for 3 days 2 hours. Then the reactivity of trachea and lung strip smooth muscle to histamine was evaluated. RESULTS: The pretreatment with vitamin C did not evoke statistical significant changes of trachea and lung strip smooth muscles reactivity when compared with the control group. The pretreatment with vitamin E produced a statistically non-significant decrease in trachea smooth muscle reactivity, but an increase in contraction amplitude of lung strip smooth muscle. Trachea was without expressive histological changes. The lung showed granulomatous inflammation with lymphocytes and eosinophils. SUMMARY: Antioxidants in used doses did not prevent the reactivity changes evoked by toluene exposure. (Fig. 2, Tab. 4, Ref. 14.)

[Mechanisms of the effect of oxidants on the respiratory system]. / K mechanizmu pôsobenia oxidantov na dýchací systém.

BACKGROUND: It is known that oxidants may evoke changes of respiratory tract functions. The precise mechanisms of these changes are yet unknown. MAIN PURPOSE: In this study possible participation of eicosanoids, cytochrome P-450 and reactive oxygen species in the changes of airways reactivity evoked by toluene exposure as the source of free radicals was followed up by an indirect method. METHODS: Used drugs--naproxen (50 mg/kg b.w.), cimetidine (50 mg/kg b.w.) and N-acetylcysteine (300 mg/kg b.w.) were administered in two doses (first 30 minutes before exposure to toluene, second six hours after first dose). After exposure to toluene (2 hours in each of 3 consecutive days) was followed up reactivity of tracheal and lung smooth muscle to histamine in "in vitro" conditions. The studied substances were not administered in the control group of animals. RESULTS: In pretreated animals exposed to toluene the administration of naproxen, cimetidine and N-acetylcysteine does not provoke pronounced changes of tracheal smooth muscle reactivity compared to control group. More pronounced effect of these drugs with decrease contraction amplitude was detected on lung smooth muscle reactivity. CONCLUSION: According to our results it is not possible to determine the precise mechanisms which participate in changes of airways reactivity. There are probably multifactorial in nature.

[Functional and morphological changes in the respiratory organs of cats after long-term exposure to pure normobaric oxygen]. / Funkcné a morfologické zmeny dýchacích orgánov maciek dlhodobo exponovaných cistému normobarickému kyslíku.

BACKGROUND: Pulmonary oxygen toxicity is very well known and proved. The influence of hyperoxia on the respiratory reflexes is not known till now. MAIN PURPOSE: To ascertain if long-lasting breathing of pure normobaric oxygen (PNO) alter respiratory reflexes. METHODS: 34 adult cats of both sexes, weighing 2.5-4.0 kg, were used in two experiments. In the first experiment 16 animals with inserted chronic tracheal cannula (CTC) were used. In the second experiment 18 animals without CTC were employed. Part of animals in both experiments was exposed to PNO (day by day for 2 weeks, 10 h daily), remaining animals were exposed to room air under the same conditions. Side tracheal pressure was recorded in unanesthetized animals of the first experiment. The second experiment was performed in anaesthetised animals (Pentobarbital Spofa, 35 mg/kg, i.p.). Oesophageal pressure and blood pressure in femoral artery were recorded. Cough reflex, sneezing and aspiration reflexes were induced by mechanical stimulation of airway mucosa. Pulmonary chemoreflex was elicited by i.v. administration of 50 micrograms phenyl biguanid. Hering-Breuer inflation reflex was induced by lung inflation with pressure of 1 kPa. Reactivity of tracheal and pulmonary smooth muscle to histamine were measured in vitro. Differences in recorded parameters between animals exposed to PNO, and to room air, were tested by Mann-Whitney-Wilcoxon test and by Student's t-test. When p < 0.05, the differences were recognized as significant. RESULTS: Significant decreasing of the expiratory parameters of the cough induced from laryngopharyngeal mucosa, inhibition of sneezing, and inhibition of aspiration reflex, were found in animals exposed to PNO. Relaxing reaction of tracheal smooth muscle of control animals to histamine was reversed to contraction in animals exposed to PNO. Morphological changes of the respiratory tract induced by influence of oxygen were found CONCLUSION: Long-lasting breathing of PNO induced changes of respiratory reactions elicited mainly from upper airway.(Fig. 6, Tab. 3, Ref. 22)

[Airway hyperreactivity]. / Hyperreaktivita dýchacích ciest.

The airway hyperreactivity (AHR) is a symptom occurring in various diseases of the respiratory system. It is defined as an abnormal bronchoconstriction response to a different spectrum of biological, chemical and pharmacological stimuli. Pathogenesis of airway hyperreactivity is not well understood. The available literature data shows that in the AHR pathogenesis is important not only genetic predisposition or influence of combination environmental and genetic factors, but also the presence and activity of various inflammatory mediators and other endogenous factors (growth factors, nuclear transcription factors). In this process is also important role of neural regulation and release of pro-inflammatory neurotransmitters. Our aim was to provide a comprehensive overview of knowledge about the symptoms--from the risk factors and pathogenesis through the clinical importance to the need for better understanding new options in therapeutic interventions.