Biomedical subjects

P Chitano

Publications and source records attributed to P Chitano.

At least 19 recordsLinked to original sources

PubMed1998

Immunization and challenge with toluene diisocyanate decrease tachykinin and calcitonin gene-related peptide immunoreactivity in guinea pig central airways.

Toluene diisocyanate (TDI) is a potent sensitizer that causes occupational asthma in a significant proportion of subjects exposed. We used an animal model to investigate whether neuropeptide changes occur in the airways of immunized and TDI-challenged guinea pigs. Animals were immunized by weekly intradermal injections, challenged with TDI (5 to 20 ppb) after the third injection, and killed 6 h after exposure. Control guinea pigs received injections of saline. Lung tissue was processed immediately and analyzed for nerves using the streptavidin-biotin complex peroxidase method with antisera to the neural marker protein gene product 9.5 (PGP 9.5), substance P (SP), and calcitonin gene- related peptide (CGRP). We also quantified the inflammatory infiltrate in the submucosa of central airways, and we measured the serum level of specific IgG and IgG1. Specific antibodies against TDI were present only in immunized animals. Immunized as compared with nonimmunized animals had a significant increase in eosinophils in the submucosa of central airways, and a further increase was observed 6 h after TDI challenge. Immunization and TDI challenge did not modify the number of mononuclear cells in the submucosa of central airways in both nonimmunized and immunized animals. TDI exposure did not change the overall innervation in both nonimmunized and immunized animals, but the density of PGP 9.5-positive nerves was significantly different between nonimmunized and immunized TDI-challenged animals. The density of SP-, and CGRP-immunostained nerves was significantly lower in immunized TDI-challenged than in nonimmunized animals. TDI exposure significantly decreased the density of SP-positive nerves in nonimmunized animals. A negative relationship was found between the presence of airway inflammation, as indexed by eosinophil cell infiltration, and the density of PGP 9.5-, SP-, and CGRP-immunostained nerves. In conclusion, TDI produces airway inflammation and neuropeptides changes in the central airways of immunized guinea pigs 6 h after TDI challenge. These findings support an interaction between tachykinins, inflammatory (i.e., eosinophils) and possibly immune cells.

Animals↗

PubMed1998

Dehydroepiandrosterone and analogs inhibit DNA binding of AP-1 and airway smooth muscle proliferation.

The adrenal steroid dehydroepiandrosterone (DHEA) and its analogs reduce growth of immortalized and malignant cell lines. We therefore explored their effects on the growth of airway smooth muscle, whose hyperplasia may lead to fixed airways obstruction and enhanced airways hyperresponsiveness in severe chronic asthma. DHEA and its potent analog 16 alpha-bromoepiandrosterone dramatically reduced proliferation in primary cultures of rat tracheal smooth muscle stimulated with fetal bovine serum or platelet-derived growth factor. Growth inhibition was dose-dependent and could not be attributed to interference with glucose-6-phosphate dehydrogenase activity or cholesterol metabolism, as reported for immortalized or malignant cell lines, respectively. Expression of the early response gene c-fos remained intact, but DHEA and 16 alpha-bromoepiandrosterone decreased DNA binding of the transcription factor activator protein-1, a later response important for expression of genes that mediate DNA synthesis and cell cycle progression. These results suggest that the nonglucocorticoid steroid DHEA and its analogs may impair activation of secondary growth response genes in a fashion analogous to that reported for glucocorticoids and that they may prove useful for treatment of asthmatic airway remodeling in the human.

Androsterone↗

PubMed1996

Increased response to antigen and histamine release in smaller sensitized canine bronchi.

We studied the Schultz-Dale response in vitro in large and small size branches from 3rd to 6th generation bronchi from ragweed-sensitized dogs. The response to electric field stimulation (EFS) increased after antigen from 65.56 +/- 8.11 to 78.6 +/- 9.0 mN/mm2 of smooth muscle (P < 0.01), but no topographical difference was observed. The response to ragweed (% of the response to EFS) was 158.3 +/- 12 and 67.1 +/- 11.7 in strips from small and large branches respectively (P < 0.01), while no difference was observed between generations; when clustering bronchi according to dimension, it was 129.9 +/- 13.4 in small and 71.9 +/- 19.8 in large bronchi (P < 0.01). Histamine released from small and large branches was 2.90 +/- 1.01 and 0.76 +/- 0.20 (ng/mg of tissue) respectively (P < 0.05); no difference was found between generations. In conclusion, in sensitized dogs a greater response to antigen, which involves a higher histamine release, occurs in small compared to large bronchi. We suggest that control of distribution of ventilation occurs mainly at small bronchi level, which becomes the elective tissue to study the Schultz-Dale response. Finally, the classification of bronchi into generations is inadequate to study allergic bronchospasm.

Allergens↗

PubMed1996

Effect of subchronic in vivo exposure to nitrogen dioxide on lung tissue inflammation, airway microvascular leakage, and in vitro bronchial muscle responsiveness in rats.

OBJECTIVES: In a previous study on bronchoalveolar lavage fluid from rats exposed in vivo for seven days to 10 ppm nitrogen dioxide (NO2), it has been shown that there is an influx of macrophages into the airways. The present study investigated the effect of seven day exposure to 10 ppm NO2, on: (a) lung tissue inflammation and morphology; (b) airway microvascular leakage; (c) in vitro contractile response of main bronchi. METHODS: Lung tissue was studied by light microscopy, after fixing the lungs by inflation with 4% formalin at a pressure of 20 cm H2O. Microvascular leakage was measured by extravasation of Evans blue dye in the larynx, trachea, main bronchi, and intrapulmonary airways. Smooth muscle responsiveness was evaluated by concentration-responses curves to acetylcholine (10(-9)-10(-3) M), serotonin (10(-9)-10(-4) M), and voltage-response curves (12-28 V) to electrical field stimulation. RESULTS: Histology showed an increased total inflammation at the level of respiratory bronchioles and alveoli. No influx of inflammatory cells was found in the main bronchi. A loss of cilia in the epithelium of small airways and ectasia of alveolar capillaries was also found. By contrast, no alterations to microvascular permeability or modification of bronchial smooth muscle responsiveness was found. CONCLUSIONS: Subchronic exposure to 10 ppm NO2 causes airway inflammation and structural damage, but does not cause any persistent alteration to microvascular permeability or bronchial smooth muscle responsiveness in rats.

Acetylcholine↗

PubMed1996

Inflammatory events in the blood and airways of guinea pigs immunized to toluene diisocyanate.

Toluene diisocyanate (TDI)-induced asthma is a common cause of occupational lung disease. We used a model to investigate the course of bronchopulmonary inflammation following immunization with TDI. Guinea pigs were immunized by weekly intradermal injections and challenged with TDI 7 d after the third injection. The animals were killed at different times after challenge and prepared for histologic examination of central and peripheral airways, for immunohistochemical studies of T lymphocyte and eosinophil distribution, and for hematologic and serologic investigations. Specific IgG1 against TDI were present only in immunized animals. In immunized TDI-challenged animals there was a significant increase in the number of metachromatic cells (at 24 h) and a late increase of eosinophils (at 48 h) in the peripheral blood. Mast cells and eosinophils were also increased in the submucosa of central airways of immunized TDI-challenged animals. A similar pattern was observed in the animals' peripheral airways. Additionally, a significant increase of T-lymphocytes and eosinophils was found in the lamina propria at 6 h after exposure in immunized TDI-challenged animals as compared with control animals. In these immunized animals, TDI challenge caused a significant increase of eosinophils, T-lymphocytes, and CD4+ T cells. These findings indicate that intradermal injections of TDI induced a specific antibody response as well as an inflammatory process in both central and peripheral airways. T cells, particularly CD4+ T cells and eosinophils, are the key cells in the immunopathologic alterations induced by TDI in the guinea pig lung.

Animals↗

PubMed1996

In vivo exposure to nitrogen dioxide (NO2) induces a decrease in calcitonin gene-related peptide (CGRP) and tachykinin immunoreactivity in guinea-pig peripheral airways.

The mammalian respiratory tract is densely innervated by sensory and autonomic fibres. Subsets of the nerves contain bioactive regulatory peptides, such as substance P, calcitonin gene-related peptide (CGRP), and neurokinins. The sensory nervous system responds to inhaled irritants, resulting in a release of neuropeptides and, thus, a decrease in the peptide immunoreactivity of the fibres. We examined the effects of inhaled nitrogen dioxide (NO2), a well-known indoor and outdoor air pollutant, on pulmonary sensory neuropeptides. Guinea-pigs were exposed for 4 h to 18 parts per million (ppm) NO2 or to air (n = 5 each). At the end of the exposure, they were killed with urethane and their lungs were fixed in 1% paraformaldehyde in phosphate-buffered saline. Cryostat sections were stained with antisera to an anatomical nerve marker, protein gene product (PGP) 9.5, and to CGRP and tachykinins, utilizing the avidin-biotinylated peroxidase method. In the noncartilaginous airways (diameter < 250 microns) of NO2-exposed animals, less tachykinin- and CGRP-immunoreactive nerve fibres were found compared with controls. No change was seen in the total nerve fibre distribution (PGP 9.5). It is concluded that the peptidergic nerves of guinea-pig peripheral airways are a sensitive indicator of exposure to nitrogen dioxide.

Administration, Inhalation↗

PubMed1996

Isotonic smooth muscle response in human bronchi exposed in vitro to nitrogen dioxide.

Exposure to nitrogen dioxide (NO2), a common oxidant airborne pollutant, has been shown to cause reversible effects on lung function and airway responsiveness, in addition to airways inflammation. However, there have been conflicting reports concerning NO2-induced airway hyperresponsiveness. In the present study, we investigated the isotonic smooth muscle response in isolated human bronchi previously exposed in vitro to NO2. Bronchial segments were obtained from 12 patients who had undergone thoracotomy for lung cancer. Bronchial segments from each patient were exposed to air and to 2.5 parts per million (ppm) NO2 for 4 h. The contractile response of bronchial rings to acetylcholine, neurokinin A (NKA), and substance P was then studied under isotonic conditions. The response to NKA was also studied in rings, with or without epithelium, exposed either to air or 7 ppm NO2. No NO2-induced alteration of the bronchial smooth muscle isotonic response was found under any of the experimental conditions. We conclude that in vitro exposure to up to 7 ppm nitrogen dioxide does not cause alterations of the human bronchial smooth muscle shortening capacity.

Acetylcholine↗

PubMed1995

The Schultz-Dale response of sensitized canine bronchial smooth muscle.

We wished to determine why in vitro agonist dose-response curves show reduced leftward shift (1/2-1 log dose units) in sensitized canine airway smooth muscle compared to curves elicited in vivo (2-3 log dose unit). The Schultz-Dale response was studied in sensitized dog tracheal (TSM) and bronchial (BSM) smooth muscle. Sensitized TSM challenged with specific antigen showed greater mechanical response, but only on exposure to 300 micrograms/ml; BSM responded to concentrations of ragweed as low as 0.001 microgram/ml. This result resolved the problem cited at the outset. Control TSM and BSM showed no response. The response in BSM is mediated through histamine release, and to a smaller extent by acetylcholine. With challenge release of histamine and acetylcholine increased significantly in sensitized airway smooth muscle. Integrated contractile responses obtained with high and low concentrations of antigen showed a dose-response relationship. Increased sensitivity of BSM to antigen compared to TSM indicates the former is the preparation of choice for study of allergic bronchoconstriction.

Acetylcholine↗

PubMed1995

In-vitro exposure of guinea pig main bronchi to 2.5 ppm of nitrogen dioxide does not alter airway smooth muscle response.

In order to investigate whether the oxidant airborne pollutant nitrogen dioxide (NO2) affects airway smooth muscle responsiveness, the contractile response of guinea pig main bronchi after in vitro exposure to 2.5 ppm of nitrogen dioxide was studied. Main bronchi were cannulated and exposed for 2 or 4 h to a constant flow of either NO2 or air. After exposure, bronchial rings were obtained and placed in a 37 degrees C jacketed organ bath filled with Krebs-Henseleit solution. Concentration-response curves were performed for acetylcholine (10(-9)-10(-3) M), substance P (10(-9)-10(-4) M), and neurokinin A (10(-10)-10(-5) M), and voltage-response curves (12-28 V) were performed for electrical field stimulation. There was no significant difference in either the smooth muscle maximal contractile response, or sensitivity between the bronchi exposed to NO2 and those exposed to air. We conclude that in vitro exposure to 2.5 ppm of NO2 does not alter airway smooth muscle responsiveness in guinea pigs.

Acetylcholine↗

PubMed1995

Effect of oxidant air pollutants on the respiratory system: insights from experimental animal research.

In the present paper, we have reviewed experimental animal studies on the effects of the two most important oxidant airborne pollutants, nitrogen dioxide and ozone, on the respiratory system. The toxic effects depend on concentration and length of exposure, and are generally similar for both oxidants, with ozone operative at lower concentrations. High doses of both oxidants cause death due to lung oedema. Exposure to sublethal levels causes functional alterations such as airflow limitation and airway hyperresponsiveness to bronchoconstrictor stimuli. These effects, which are generally reversible, are associated with epithelial injury, oedema and airway and parenchymal infiltration by inflammatory cells. Loss of cilia of airway epithelium and necrosis of type I alveolar epithelial cells are the most prominent consequences at the epithelial level. Inflammation is characterized by early neutrophilic infiltration, followed by an increased number of mononuclear cells, predominantly alveolar macrophages. After long-term exposure, whilst nitrogen dioxide causes predominantly emphysema, ozone produces mainly pulmonary fibrosis. Biochemical effects include lipid peroxidation, increased antioxidant metabolism, and alteration of enzyme activity. Nitrogen dioxide and ozone may also alter the immunological response and reduce the defence against infections, increasing the susceptibility of exposed animals to infections.

Animals↗

PubMed1994

The effects of toluene diisocyanate and of capsaicin on human bronchial smooth muscle in vitro.

Toluene diisocyanate contracts guinea-pig bronchial smooth muscle through a mechanism involving capsaicin-sensitive sensory nerves. In the present study, we investigated the effects of toluene diisocyanate, capsaicin and tachykinins on isolated human bronchi. In 44 rings, toluene diisocyanate (0.3 mM) produced a relaxation which averaged 16.9 +/- 1.1%, in ten rings it produced a shortening that was 15.1 +/- 3.3% and in ten preparations it gave no response. A second administration of toluene diisocyanate (0.3 mM) always produced a relaxation (n = 13, 18.1 +/- 3.9%). Capsaicin (0.03 mM) produced shortening in 15 (35 +/- 6.6%) and relaxation in 11 preparations (41 +/- 6.8%), whereas a second administration caused shortening in nine (25.1 +/- 6.1%) and relaxation in 16 rings (36.4 +/- 4.9%). When toluene diisocyanate was given after two consecutive capsaicin administrations, we observed shortening in two rings (10.0 +/- 3.6%), relaxation in ten rings (15.9 +/- 3.6%), and no response in four preparations. To test the role of NK1 and NK2 receptors in these conflicting responses, we performed concentration-response curves to different tachykinins. Substance P, neurokinin A and neurokinin A-(4-10), a specific NK2 receptor agonist, gave a concentration-dependent shortening, with neurokinin A being the most effective and neurokinin A-(4-10) the least. The specific NK1 receptor agonist, [Sar9, Met(O2)11]substance P, produced both shortening and relaxation. We conclude that toluene diisocyanate and capsaicin may produce both shortening and relaxation in isolated human bronchi through NK1 receptors.

Acetylcholine↗

PubMed1994

In vitro exposure to nitrogen dioxide (NO2) does not alter bronchial smooth muscle responsiveness in ovalbumin-sensitized guinea-pigs.

The aim of this study was to investigate whether in vitro exposure to NO2 affects responsiveness in ovalbumin-sensitized guinea-pig bronchi. Twenty-three animals were sensitized by three weekly intraperitoneal injections of 1 mg ovalbumin in saline with Freund's adjuvant; twenty-one control guinea-pigs received the diluent alone. From each animal, the two main bronchi were obtained and cannulated, then exposed in vitro to a constant intraluminal flow of: (i) either air or 2.5 ppm NO2 with four spikes of 10 ppm NO2 for 2 h; (ii) either air or 10 ppm NO2 for 4 h. A bronchial ring obtained from each animal before exposure was kept in aerated Krebs-Henseleit solution. Rings from bronchi exposed to air, NO2, or kept in Krebs solution were studied isometrically. We performed overall and non-adrenergic non-cholinergic voltage-response curves to electrical field stimulation, concentration-response curves to acetylcholine and to neurokinin A, followed by administration of 10 mg/ml ovalbumin. We did not find any significant difference in bronchial smooth muscle responsiveness between nonexposed, air-exposed and NO2-exposed bronchi, as well as between bronchi from control and sensitized animals. We conclude that in vitro exposure to NO2 does not alter bronchial smooth muscle responsiveness to either specific or non-specific stimuli.

Acetylcholine↗

PubMed1994

Mechanisms and pathology of occupational asthma.

Since the pathogenesis and the pathological features of occupational asthma are similar to those of nonoccupational asthma, the former represents a very useful model for the investigation of the pathogenesis of asthma in general. More than one mechanism may be operative in occupational asthma. Among the mechanisms proposed, immunological mechanisms and airway inflammation play an important role. There is evidence to confirm that T-lymphocyte activation and local accumulation in the bronchial wall of activated eosinophils occurs in asthma of diverse aetiology, i.e. immunoglobulin E (IgE)-mediated, occupational and intrinsic. Neurogenic pathways should be further investigated as a potential mechanism of modulation and amplification of airway inflammation in occupational asthma.

Adult↗

PubMed1993

The effect of compound 48/80 on contractions induced by toluene diisocyanate in isolated guinea-pig bronchus.

We have investigated the ability of compound 48/80 and of histamine H1 and H2 receptor antagonists to inhibit toluene diisocyanate-induced contractions in isolated guinea-pig bronchi. Compound 48/80 (100 micrograms/ml) significantly inhibited toluene diisocyanate-induced contractions. By contrast, the two histamine H1 and H2 receptor antagonists, chlorpheniramine (10 microM) and cimetidine, (10 microM) did not affect toluene diisocyanate-induced contractions, but significantly inhibited contractions induced by exogenously applied histamine (100 microM) and by 48/80. We investigated which mechanisms 48/80 used to inhibit toluene diisocyanate-induced contractions, paying particular attention to the possible involvement of capsaicin-sensitive primary afferents. In vitro capsaicin desensitization (10 microM for 30 min followed by washing) significantly reduced compound 48/80-induced contractions. A capsaicin-resistant component of contraction was also evident. Ruthenium red (3 microM), an inorganic dye which acts as a selective functional antagonist of capsaicin, did not affect 48/80-induced contraction. MEN 10,207 (Tyr5,D-Trp6,8,9,Arg10)-neurokinin A (4-10) (3 microM) a selective antagonist of NK2-tachykinin receptors significantly reduced 48/80-induced contractions. These results show that compound 48/80 inhibits toluene diisocyanate-induced contractions in isolated guinea-pig bronchi. It is likely that two mechanisms are involved in the inhibition: (1) the release of mediators other than histamine by mast cells, (2) an effect of 48/80 on sensory nerves.

Animals↗

PubMed1993

Relevance of classification by size to topographical differences in bronchial smooth muscle response.

To investigate heterogeneity of airway smooth muscle response, we studied strips of large and small branches from third- to sixth-generation bronchi obtained from ragweed antigen-sensitized and control dogs. The response to electrical field stimulation and carbamylcholine chloride was greater in strips from larger branches of the same generation when expressed as "tissue stress" (force per unit cross-sectional area of the whole tissue), whereas no difference emerged with use of the more appropriate "smooth muscle stress" (force per unit cross-sectional area of the muscle tissue). The response to histamine was significantly higher in small branches than in large ones, and histamine sensitivity [mean effective concentration (EC50)] was 7.79 x 10(-6) [geometric standard error of the mean (GSEM) 1.20] and 1.49 x 10(-5) M (GSEM 1.14), respectively (P < 0.01). Strips from control and sensitized animals at each site and strips from different generations did not show any significant difference. When we clustered our preparations according to dimensions, the response to histamine was significantly higher in small bronchi than in large ones and histamine EC50 was 8.95 x 10(-6) (GSEM 1.17) and 1.57 x 10(-5) M (GSEM 1.18), respectively (P < 0.05). We conclude that evaluation of muscle response in different tissues requires appropriate normalization. Furthermore, classification into generations is inadequate to study bronchial responsiveness, inasmuch as major differences originate from airway size.

Animals↗

PubMed1992

The products of the reaction between toluene diisocyanate and water contract isolated guinea pig bronchi.

We have investigated the ability of the products of the reaction between toluene diisocyanate (TDI) and water to contract bronchial smooth muscle. The experiments were performed in isolated guinea pig bronchi. TDI, both 2,4- and 2,6-toluenediamine (TDA) and mixtures of 2,4- and 2,6-TDA (ratio 80:20 and 20:80) caused concentration-dependent contraction in the isolated bronchi. The mixture of disubstituted urea and biuret also contracted the bronchi, but not in a concentration-dependent fashion. Our results provide evidence that all products of the reaction between toluene diisocyanate and water have the ability to contract isolated bronchial smooth muscle in guinea pigs. Whatever the role of toluenediamine in the adverse respiratory effects induced by exposure to isocyanates, our findings reveal the necessity of in vivo studies on the metabolism of inhaled toluene diisocyanate in humans to improve our understanding of the mechanism of action of isocyanates.

Animals↗

PubMed1992

The effect of phosphoramidon and epithelium removal on toluene diisocyanate-induced contractions in guinea-pig bronchi.

To evaluate the role of airway neutral endopeptidase 24.11 (NEP) and epithelium removal in the contraction of airway smooth muscle in response to toluene diisocyanate (TDI), we studied the effects of the NEP inhibitor, phosphoramidon, on TDI-induced contractions of guinea-pig bronchial rings with intact epithelium and without epithelium. In preparations with intact epithelium, phosphoramidon (10 microM) potentiated the contractile response to TDI (0.3 mM) (mean +/- SEM, 23.7 +/- 2.5% versus 67.9 +/- 10.3%, p less than 0.01). Phosphoramidon also increased TDI-induced contractions in tissues without epithelium (36.9 +/- 4.9% versus 52.5 +/- 7.1%, p less than 0.05). Removal of the epithelium increased the contractile response to TDI (23.7 +/- 2.5% versus 36.9 +/- 4.9%, p less than 0.05). These results demonstrate the response to TDI is increased in epithelium-free compared to intact bronchi and that NEP 24.11 modulates the effects of endogenously released tachykinins by TDI at all of the sites where NEP is found in the airways.

Animals↗

PubMed1991

Bronchial smooth muscle responses evoked by toluene diisocyanate are inhibited by ruthenium red and by indomethacin.

We have investigated the ability of ruthenium red, an inorganic dye with Ca2+ entry-blocking properties and a selective antagonist of capsaicin, and of indomethacin, a cyclooxygenase inhibitor, to inhibit bronchial smooth muscle responses evoked by toluene diisocyanate in guinea pigs. Previous exposure of isolated guinea pig bronchi to ruthenium red significantly decreased the response produced by toluene diisocyanate. Further, the response to toluene diisocyanate was significantly decreased by pretreatment with indomethacin. These findings provide evidence that toluene diisocyanate-induced contractions of guinea pig bronchi are produced indirectly by generation of a prostanoid that activates capsaicin-sensitive afferents via a ruthenium red-sensitive mechanism.

Animals↗