Early exposure to farm dust in an allergic airway inflammation rabbit model: Does it affect bronchial and cough hyperresponsiveness?

INTRODUCTION: Over the past 50 years, the prevalence of allergic respiratory diseases has been increasing. The Hygiene hypothesis explains this progression by the decrease in the bio-diversity of early microbial exposure. This study aims to evaluate the effect of early-life farm exposure on airway hyperresponsiveness and cough hypersensitivity in an allergic airway inflammation rabbit model. METHOD: A specific environment was applied to pregnant rabbits and their offspring until six weeks after birth. Rabbits were housed in a pathogen-free zone for the control group and a calf barn for the farm group. At the end of the specific environmental exposure, both groups were then housed in a conventional zone and then sensitized to ovalbumin. Ten days after sensitization, the rabbit pups received ovalbumin aerosols to provoke airway inflammation. Sensitization to ovalbumin was assessed by specific IgE assay. Cough sensitivity was assessed by mechanical stimulation of the trachea, and bronchial reactivity was assessed by methacholine challenge. The farm environment was characterized by endotoxin measurement. RESULTS: A total of 38 rabbit pups were included (18 in the farm group). Endotoxin levels in the farm environment varied from 30 to 1854 EU.m-3. There was no significant difference in specific IgE values to ovalbumin (p = 0.826) between the two groups. The mechanical threshold to elicit a cough did not differ between the two groups (p = 0.492). There was no difference in the number of cough (p = 0.270) or the intensity of ventilatory responses (p = 0.735). After adjusting for age and weight, there was no difference in respiratory resistance before and after methacholine challenge. CONCLUSION: Early exposure to the calf barn did not affect cough sensitivity or bronchial reactivity in ovalbumin-sensitized rabbits. These results suggest that not all farm environments protect against asthma and atopy. Continuous exposure to several sources of microbial diversity is probably needed.

Impact of Inhaled Corticosteroids on the Modulation of Respiratory Defensive Reflexes During Artificial Limb Exercise in Ovalbumin-Sensitized Rabbits.

INTRODUCTION: Cough is a major lower airway defense mechanism that can be triggered by exercise in asthma patients. Studies on cough reflex in experimental animal models revealed a decrease of cough reflex sensitivity during exercise in healthy animals, but a lack of desensitization in ovalbumin-sensitized rabbits. The aim of our study is to evaluate the impact of inhaled corticosteroids on cough reflex during artificial limb exercise in an animal model of eosinophilic airway inflammation. MATERIALS AND METHODS: Sixteen adult ovalbumin-sensitized rabbits were randomly divided into two groups. The "OVA-Corticoid" group (n = 8) received inhaled corticosteroids (budesonide; 1 mg/day during 2 consecutive days) while the "OVA-Control" (n = 8) group was exposed to saline nebulization. The sensitivity of defensive reflexes induced by direct mechanical stimulation of the trachea was studied in anesthetized animals, at rest and during artificial limb exercise. Cell count was performed on bronchoalveolar lavage fluid and middle lobe tissue sections to assess the level of eosinophilic inflammation. RESULTS: All rabbits were significantly sensitized but there was no difference in eosinophilic inflammation on bronchoalveolar lavage or tissue sections between the two groups. Artificial limb exercise resulted in a significant (p = 0.002) increase in minute ventilation by 30% (+ 209 mL.min-1, ± 102 mL/min-1), with no difference between the two groups. 322 mechanical tracheal stimulations were performed, 131 during exercise (40.7%) and 191 at rest (59.3%). Cough reflex was the main response encountered (46.9%), with a significant increase in cough reflex threshold during artificial limb exercise in the "OVA-Corticoid" group (p = 0.039). Cough reflex threshold remained unchanged in the "OVA-Control" group (p = 0.109). CONCLUSION: Inhaled corticosteroids are able to restore desensitization of the cough reflex during artificial limb exercise in an animal model of airway eosinophilic inflammation. Airway inflammation thus appears to be involved in the physiopathology of exercise-induced cough in this ovalbumin sensitized rabbit model. Inhaled anti-inflammatory treatments could have potential benefit for the management of exercise-induced cough in asthma patients.

Desensitization of the Cough Reflex Induced by Corticosteroids in Ovalbumin-Sensitized Rabbits During Artificial Limb Exercise.

INTRODUCTION: Cough is a major symptom frequently experienced during exercise, mainly in asthmatic patients. Inhaled glucocorticoids represent the keystone treatment in the management of asthma, but little is known about interactions between cough and exercise, especially in controlled patients. During exercise, cough reflex (CR) appears downregulated in healthy animal models whereas a lack of desensitization of CR has been shown in ovalbumin-sensitized animal models, mimicking asthmatic disease. AIMS AND OBJECTIVES: The goal of our study was to clarify the potential modulation of the CR induced by inhaled corticosteroids (CS) in ovalbumin (OVA) sensitized rabbits during artificial limb exercise. MATERIALS AND METHODS: Seventeen OVA sensitized rabbits were studied. Among them, 9 were treated with CS delivered intravenously (OVA-Corticoids). The ventilatory response to direct tracheal stimulation, performed at rest and during exercise, was determined to assess the incidence and the sensitivity of the CR. Broncho-alveolar lavage (BAL) and cell counts were performed to determine the level of airway inflammation. Exercise was mimicked by electrically induced hindlimb muscular contractions (EMC). RESULTS: Compared to rest values, EMC increased minute ventilation by 28% without any decrease in respiratory resistance (Rsr). Among 322 tracheal stimulations, 172 (53%) were performed at rest and 150 (47%) during exercise. The sensitivity of CR decreased during artificial limb exercise compared to baseline in OVA-Corticoids rabbits (p = 0.0313) while it remained unchanged in OVA rabbits (p = NS). CONCLUSION: Corticosteroids appear to restore the desensitization of the CR in OVA sensitized rabbits during artificial limb exercise, suggesting the potential role of airway inflammation in the pathophysiology of cough during exercise in asthmatics.

Modulation of protective reflex cough by acute immune driven inflammation of lower airways in anesthetized rabbits.

Chronic irritating cough in patients with allergic disorders may reflect behavioral or reflex response that is inappropriately matched to the stimulus present in the respiratory tract. Such dysregulated response is likely caused by sensory nerve damage driven by allergic mediators leading to cough hypersensitivity. Some indirect findings suggest that even acid-sensitive, capsaicin-insensitive A-δ fibers called "cough receptors" that are likely responsible for protective reflex cough may be modulated through immune driven inflammation. The aim of this study was to find out whether protective reflex cough is altered during acute allergic airway inflammation in rabbits sensitized to ovalbumin. In order to evaluate the effect of such inflammation exclusively on protective reflex cough, C-fiber mediated cough was silenced using general anesthesia. Cough provocation using citric acid inhalation and mechanical stimulation of trachea was realized in 16 ovalbumin (OVA) sensitized, anesthetized and tracheotomised rabbits 24h after OVA (OVA group, n = 9) or saline challenge (control group, n = 7). Number of coughs provoked by citric acid inhalation did not differ between OVA and control group (12,2 ±6,1 vs. 17,9 ± 6,9; p = 0.5). Allergic airway inflammation induced significant modulation of cough threshold (CT) to mechanical stimulus. Mechanically induced cough reflex in OVA group was either up-regulated (subgroup named "responders" CT: 50 msec (50-50); n = 5 p = 0.003) or down-regulated (subgroup named "non responders", CT: 1200 msec (1200-1200); n = 4 p = 0.001) when compared to control group (CT: 150 msec (75-525)). These results advocate that allergen may induce longer lasting changes of reflex cough pathway, leading to its up- or down-regulation. These findings may be of interest as they suggest that effective therapies for chronic cough in allergic patients should target sensitized component of both, reflex and behavioral cough.

Modulation of defensive airway reflexes during continuous positive airway pressure in the rabbit.

Lung hyperinflation may alter the pattern of ventilatory reflexes in chronic respiratory disorders. The aim of the study was to test the effect of continuous positive airway pressure (CPAP) on ventilatory responses to mechanical stimulation of the trachea.

Down-Regulation of Cough during Exercise Is Less Frequent in Healthy Children than Adults. Role of the Development and/or Atopy?

Cough is typically associated with physical activity in children with asthma, but the characteristics of the relationship between cough and exercise has not been established under physiological conditions. The aim of the study was to describe the effect of exercise on the reflex cough response elicited by a single breath of capsaicin in non-asthmatic children. A group of non-asthmatic adults was studied as reference. Thirty children and 29 adults were recruited. The cough reflex sensitivity to capsaicin was first determined to establish the dose that provokes 5 cough efforts (C5). The number of coughs elicited by C5 (NC5) was then compared at baseline and during a standardized submaximal treadmill exercise. Data are expressed as median (interquartile range). Children and adults showed a significant decrease in NC5 (respectively from 5.0 (4.0-6.0) to 2.5 (2.0-4.0), p < 0.0005 and from 6.0 (5.0-7.0) to 2.0 (0.0-3.0, p < 0.0005). During exercise, NC5 was observed to decrease in all adult subjects, but in only 24/30 children (80%, p = 0.02). A trend for a higher incidence of personal and familial atopy was observed in children that lacked cough down-regulation during exercise compared with other children. It is concluded that the cough reflex response to capsaicin is down regulated by exercise in both children and adults. The effect however is less consistently observed in the former. The difference may reflect maturation of descending inhibitory pathways of the cough reflex, but may also be associated to atopy. The data stress the importance of assessing the time relationship of cough and exercise in questionnaire studies of asthma.

Lack of desensitization of the cough reflex in ovalbumin-sensitized rabbits during exercise.

INTRODUCTION: Cough is a major symptom of asthma frequently experienced during exercise but little is known about interactions between cough and exercise. The goal of our study was to clarify the potential modulation of the cough reflex (CR) by exercise in a spontaneously breathing anaesthetized animal model of airway eosinophilic inflammation. MATERIALS & METHODS: Ten ovalbumin (OVA) sensitized adult rabbits and 8 controls were studied. The ventilatory response to direct tracheal stimulation, performed both at rest and during exercise was determined to quantify the incidence and the sensitivity of the CR. Broncho-alveolar lavages (BAL) and cell counts were performed to assess the level of the airway inflammation following OVA-induced sensitization. Exercise was mimicked by Electrically induced hindlimb Muscular Contractions (EMC). RESULTS: Among 494 tracheal stimulations, 261 were performed at rest and 233 at exercise. OVA challenges in sensitized rabbits caused a significant increase in the percentage of eosinophils (p = 0.008) in BAL. EMC increased minute ventilation by 36% and 35% in OVA and control rabbits respectively, compared to rest values. The sensitivity of the CR decreased during exercise compared to baseline in control rabbits (p = 0.0313) while it remained unchanged in OVA rabbits. CONCLUSION: The desensitization of the CR during exercise in control rabbits was abolished in OVA rabbits. The precise role of airway inflammation in this lack of CR desensitization needs to be further investigated but it might contribute to the exercise-induced cough in asthmatics.

Hemocyte responses of Dreissena polymorpha following a short-term in vivo exposure to titanium dioxide nanoparticles: preliminary investigations.

The widespread use of titanium-based nanoparticles and their environmental release may pose a significant risk to aquatic organisms within freshwater ecosystems. Suspension-feeder invertebrates like bivalve molluscs represent a unique target group for nanoparticle toxicology. The aim of this work was to investigate the short-term responses of Dreissena polymorpha hemocytes after in vivo exposure to titanium dioxide nanoparticles (TiO(2) NP). For this purpose, freshwater mussels were exposed to P25 TiO(2) NP at the concentrations of 0.1, 1, 5 and 25mg/L during 24h. Viability, phagocytosis activity and mitogen activated protein kinase (MAPK) phosphorylation level of ERK 1/2 and p38 in hemocytes extracted from exposed mussels were compared to those from control specimens. Results demonstrated an inhibition of the phagocytosis activity after exposure to TiO(2) NP at 0.1 and 1mg/L. Similar trends, albeit less pronounced, were reported for higher concentrations of NP. Transmission electron microscopy showed for the first time the internalization of TiO(2) NP into Dreissena polymorpha hemocytes. Besides, exposure to NP increased the ERK 1/2 phosphorylation levels in all treatments. Concerning the phosphorylation level of p38, only exposures to 5 and 25mg/L of NP induced significant p38 activation in comparison to that of the control. Finally, these short-term effects observed at environmentally relevant concentrations highlighted the need for further studies concerning ecotoxicological evaluation of nanoparticle release into an aquatic environment.

Ecotoxicological assessment of TiO2 byproducts on the earthworm Eisenia fetida.

The increasing production of nanomaterials will in turn increase the release of nanosized byproducts to the environment. The aim of this study was to evaluate the behaviour, uptake and ecotoxicity of TiO(2) byproducts in the earthworm Eisenia fetida. Worms were exposed to suspensions containing 0.1, 1 and 10 mg/L of byproducts for 24 h. Size of TiO(2) byproducts showed aggregation of particles up to 700 µm with laser diffraction. Only worms exposed at 10 mg/L showed bioaccumulation of titanium (ICP-AES), increasing expression of metallothionein and superoxide dismutase mRNA (Real-time PCR) and induction of apoptotic activity (Apostain and TUNEL). TiO(2) byproducts did not induce cytotoxicity on cœlomocytes, but a significant decrease of phagocytosis was observed starting from 0.1 mg/L. In conclusion, bioaccumulation of byproducts and their production of reactive oxygen species could be responsible for the alteration of the antioxidant system in worms.

Nanoparticle interactions with zinc and iron: implications for toxicology and inflammation.

BACKGROUND: Particulate air pollution (PM(10)) consists of a mixture of components, including nanoparticles and metals. Studies from our laboratory have demonstrated that transition metals can potentiate the ability of nanoparticles to induce lung inflammation and that the zinc content of PM(10) was largely responsible for their potential to induce inflammation. These results are also relevant to zinc-containing engineered nanoparticles. OBJECTIVES: To investigate the potential of ZnCl(2) and FeCl(3) to interact with nanoparticle carbon black in cell-free and biological systems to generate ROS, express pro-inflammatory mediators and cytotoxic ability. METHODS: ROS production was examined using DCFH-DA. J774 cells were treated for 4 h with 14 nm CB and/or ZnCl(2) before measuring TNF-alpha by ELISA. Cytoskeletal changes were investigated using confocal microscopy. Flow cytometry was used to examine apoptotic/necrotic cells and phagocytic ability. RESULTS: In a cell-free system the particles generated significant ROS, whereas ZnCl(2) did not. Treatment of cells with 100 microM ZnCl(2), but not FeCl(3), increased TNF-alpha. Treatment with 14 nm CB alone induced TNF-alpha, which was synergistically enhanced by ZnCl(2). No interactions were observed in cells treated with 14 nm CB and FeCl(3). Cytoskeletal changes were observed with increasing concentrations of ZnCl(2). These results were confirmed by flow cytometry indicating that ZnCl(2) induced markers of apoptosis and necrosis. The phagocytic ability of cells was also significantly decreased. Nanoparticle carbon black alone did not induce changes in apoptosis/necrosis or the phagocytosis activity of the cells. CONCLUSION: Despite an inability to induce ROS production, ZnCl(2) stimulated TNF-alpha production which was synergistically enhanced by 14 nm carbon black. The ability of zinc to induce morphological changes and cell death was not altered by nanoparticle treatment.

Influences of ozone exposure upon macrophage responsivity to N-formyl-methionyl-leucyl-phenylalanine: mobility and metabolic changes.

Alveolar macrophages represent one of the first lines of cell defence in the lungs. They employ several mechanisms, including phagocytosis and secretion of reactive oxygen and nitrogen species. fMLP, a formylated peptide of bacterial origin, is a potent inducer of phagocyte chemotaxis and is also involved in generating antimicrobial agents such as nitric oxide (NO) and hydrogen peroxide (H2O2). In this study we analysed the in vitro effects of fMLP on the mobility of the THP-1 cell line, which served as a model for alveolar macrophages. Cell mobility and cytotoxicity were also analysed after pre-exposures to an atmosphere polluted with ozone (0.03-0.5 ppm) followed by a fMLP treatment. Finally, the secreted molecules (H2O2 and NO) were measured after ozone exposures ranging from 5 to 30 min and fMLP action. Activation by fMLP alone induced cell movement, whereas pre-exposure to the ozone concentrations decreased it. Addition of fMLP had different effects on cytotoxicity, mobility and metabolite secretion by the cells: (1) cytotoxicity increased depending on ozone concentrations and exposure times; (2) during the first 5 min and for all ozone concentrations, an average decrease of 50% of activated cell mobility was observed; (3) H2O2 was increased, even in combination with ozone; (4) NO was detected at 731 nM, a result that was not affected by ozone pre-exposure.

Precipitation of silver-thiosulfate complex and immobilization of silver by Cupriavidus metallidurans CH34.

Cupriavidus metallidurans CH34 is a facultative chemolithotrophic bacterium that possesses two megaplasmids (pMOL28 and pMOL30) that confer resistance to eleven metals. The ability of Cupriavidus metallidurans CH34 to resist silver is described here. Electronic microscopy, energy-dispersive X-ray (EDX) and X-ray diffractometry (DRX) observations revealed that C. metallidurans CH34 strongly associated silver with the outer membrane, under chloride chemical form. Using derivate strains of C. metallidurans CH34, which carried only one or no megaplasmid, we show that this resistance seems to be carried by pMOL30.