Effect of roflumilast on airway remodeling in asthmatic mice exposed to or not exposed to cigarette smoke: Comparison with the effect of dexamethasone.

Cigarette smoking constitutes a risk factor for severe asthma, which is frequently linked to remodeling of the airways. Appropriate drug treatment for smokers with asthma is uncertain because many smokers with asthma are less sensitive to glucocorticoid treatment than non-smokers with asthma. The purpose of this study was to compare the anti-airway remodeling effects of dexamethasone (Dex) and roflumilast (Rof), a selective phosphodiesterases-4 inhibitor, in smoking and non-smoking mice with asthma. BALB/c mice were sensitized with ovalbumin (OVA) and then challenged with OVA for two weeks, either with or without concurrent exposure to cigarette smoke (CS). Dex (1 mg/kg body weight), Rof (5 mg/kg body weight), or vehicle alone was given orally to the mice once daily. To assess the histopathological effects of airway remodeling, lung tissue sections were obtained. Repeated OVA challenges resulted in fibrosis, goblet cell hyperplasia, and thickening of the airway but not the smooth muscle layer. The presence of CS did not have an impact on the degree of airway remodeling brought on by repeated OVA challenges. In mice repeatedly exposed to OVA either with or without CS, Dex treatment reduced the remodeling alterations. In these mice group, the Rof Treatment had a less significant impact than the Dex treatment. Dex was still more effective than Rof at reducing airway remodeling in asthmatic smoking mice. According to the current study's findings, Dex effectively prevented airway remodeling in a two-week asthma model in mice exposed to CS or not. In contrast, we found that Rof had little to no inhibitory effect of Rof on the airway in our mouse model of asthma, whether or not it had been exposed to CS. We were unable to find solid proof to support CS-induced steroid resistance to treat airway remodeling.

Differential effects of dexamethasone and roflumilast on asthma in mice with or without short cigarette smoke exposure.

Appropriate drug treatment for smoking asthmatics is uncertain because most smokers with asthma are less sensitive to treatment with glucocorticoids compared with non-smokers with asthma. We hypothesized that roflumilast (Rof), a selective phosphodiesterases-4 inhibitor regarded as an add-on therapy for chronic obstructive pulmonary disease, might be more effective than glucocorticoids for improving asthma in smokers. To investigate this hypothesis, we compared the therapeutic effects of dexamethasone (Dex) and Rof in a mouse model of ovalbumin-induced asthma with or without concurrent cigarette smoke (CS) exposure for 2 weeks. We found that recurrent asthma attacks increased lung tissue resistance. CS exposure in asthmatic mice decreased the central airway resistance, increased lung compliance, and attenuated airway hyper-responsiveness (AHR). CS exposure in asthmatic mice also increased the number of neutrophils and macrophages in the bronchoalveolar fluid. Treatment with Dex in asthmatic mice without CS exposure reduced airway resistance, AHR and airway eosinophilia. In asthmatic mice with CS exposure, however, Dex treatment unexpectedly increased lung tissue resistance and restored AHR that had been otherwise suppressed. Dex treatment in asthmatic mice with CS exposure inhibited eosinophilic inflammation but conversely exacerbated neutrophilic inflammation. On the other hand, treatment with Rof in asthmatic mice without CS exposure reduced airway resistance and airway eosinophilia, although the inhibitory effect of Rof on AHR was unremarkable. In asthmatic mice with CS exposure, Rof treatment did not exacerbate lung tissue resistance but modestly restored AHR, without any significant effects on airway inflammation. These results suggest that CS exposure mitigates sensitivity to both Dex and Rof. In asthmatic mice with CS exposure, Dex is still effective in reducing eosinophilic inflammation but increases lung tissue resistance, AHR and neutrophilic inflammation. Rof is ineffective in improving lung function and inflammation in asthmatic mice with CS exposure. This study did not support our initial hypothesis that Rof might be more effective than glucocorticoids for improving asthma in smokers. However, glucocorticoids may have a detrimental effect on smoking asthmatics.

Systemic effects of acute cigarette smoke exposure in mice.

CONTEXT: Cigarette smoke (CS) causes both pulmonary and extrapulmonary disorders. OBJECTIVE: To determine the pulmonary and extrapulmonary effects of acute CS exposure in regard to inflammation, oxidative stress and DNA damage. MATERIALS AND METHODS: Mice were exposed to CS for 10 days and then their lungs, heart, liver, pancreas, kidneys, gastrocnemius muscle and subcutaneous (inguinal and flank) and visceral (retroperitoneum and periuterus) adipose tissues were excised. Bronchoalveolar lavage fluid samples were obtained for differential cell analysis. Inflammatory cell infiltration of the tissues was assessed by immunohistochemistry for Mac-3(+) cells, F4/80(+) cells and CD45(+) cells. Oxidative stress was determined by immunohistochemistry for thymidine glycol (a marker of DNA peroxidation) and 4-hydroxy hexenal (a marker of lipid peroxidation), by enzyme-linked immunosorbent assay for protein carbonyls (a marker of protein peroxidation) and by measurements of enzyme activities of glutathione peroxidase, superoxide dismutase and catalase. DNA double-strand breaks were assessed by immunohistochemistry for γH2AX. RESULTS: CS exposure-induced inflammatory cell infiltration, oxidative stress and DNA damage in the lung. Neither inflammatory cell infiltration nor DNA damage was observed in any extrapulmonary organs. However, oxidative stress was increased in the heart and inguinal adipose tissue. DISCUSSIONS: Induction of inflammatory cell infiltration and DNA damage by acute CS exposure was confined to the lung. However, an increased oxidative burden occurred in the heart and some adipose tissue, as well as in the lung. CONCLUSIONS: Although extrapulmonary effects of CS are relatively modest compared with the pulmonary effects, some extrapulmonary organs are vulnerable to CS-induced oxidative stress.

Pulmonary emphysema induced by cigarette smoke solution and lipopolysaccharide in guinea pigs.

Exposure of animals to cigarette smoke for longer than 3 months leads to the development of chronic obstructive pulmonary disease (COPD) showing pulmonary emphysema. We attempted to create a COPD model with emphysema that could be established in a shorter period of time. Guinea pigs were intratracheally treated once a day on days 0-3, 5-8, 10-13 and 15-18 with a cigarette smoke solution (CSS), which was prepared by bubbling a stream of smoke into saline. Additionally, lipopolysaccharide (LPS) was administered intratracheally as an exacerbation factor on days 4, 9 and 14. By day 19, there was a gradual elevation of specific airway resistance (sRaw). In addition, both residual volume and functional residual capacity were found to be significantly higher on day 19. In the lungs, there was a marked increase in leukocytes, especially neutrophils. Histologically, we observed epithelial hyperplasia and emphysema. On the other hand, daily oral administration of theophylline during the administration of CSS and LPS suppressed the sRaw increase and the epithelial hyperplasia, but not other functional structural changes. In conclusion, we established an experimental COPD model in guinea pigs by using intratracheal instillations of CSS and LPS over a considerably shorter term than has been reported for other models.

Aquaporin-3 potentiates allergic airway inflammation in ovalbumin-induced murine asthma.

Oxidative stress plays a pivotal role in the pathogenesis of asthma. Aquaporin-3 (AQP3) is a small transmembrane water/glycerol channel that may facilitate the membrane uptake of hydrogen peroxide (H2O2). Here we report that AQP3 potentiates ovalbumin (OVA)-induced murine asthma by mediating both chemokine production from alveolar macrophages and T cell trafficking. AQP3 deficient (AQP3(-/-)) mice exhibited significantly reduced airway inflammation compared to wild-type mice. Adoptive transfer experiments showed reduced airway eosinophilic inflammation in mice receiving OVA-sensitized splenocytes from AQP3(-/-) mice compared with wild-type mice after OVA challenge, consistently with fewer CD4(+) T cells from AQP3(-/-) mice migrating to the lung than from wild-type mice. Additionally, in vivo and vitro experiments indicated that AQP3 induced the production of some chemokines such as CCL24 and CCL22 through regulating the amount of cellular H2O2 in M2 polarized alveolar macrophages. These results imply a critical role of AQP3 in asthma, and AQP3 may be a novel therapeutic target.

A cardiac rhabdomyoma in a Guinea pig.

A guinea pig (9-week-old) that had been placed in a control group for a pharmacological test was found to have a single nodule on the surface of the right ventricular wall. In a transverse section of the heart after fixation, a whitish mass was found that extended from the subendocardium to the subepicardium of the right ventricular wall. Histopathological examination revealed a spongy network consisting of vacuolated spaces in the myocardium of the right ventricle extending to the myocardium and subepicardium of the right atrium. The vacuolated space was PAS-positive. Immunohistochemical examinations revealed that the lesions contained striated fibers that were positive for anti-desmin and anti-myoglobin. Electron micrographs revealed the lesions resulting in affected striated muscle fibers and accumulations of many glycogen granules. Based on the findings, the lesions were diagnosed as a cardiac rhabdomyoma. This is the first report of application of immunohistochemical examinations to diagnosis of cardiac rhabdomyoma in the guinea pig.

Vulnerability of gastric mucosa to prednisolone in rats chronically exposed to cigarette smoke.

We examined gastric mucosal vulnerability in a rat model of chronic obstructive pulmonary disease (COPD). Male Wistar rats were exposed to cigarette smoke for 12 weeks (CSE rats), and on the last 4 days of exposure, prednisolone was given to induce gastric mucosal injury. Histopathology, pulmonary function, arterial blood gases, and levels of lipid peroxides (LPO), prostaglandin E(2) (PGE(2)), hypoxia-inducible factor 1 alpha subunit (HIF-1alpha), and vascular endothelial growth factor (VEGF) in gastric mucosa were examined. We also tested the effect of rebamipide on prednisolone-induced gastric lesions. In CSE rats, although no gastric lesions were detected, LPO, PGE(2), HIF-1alpha, and VEGF levels were higher than in control rats. Prednisolone induced gastric hemorrhagic lesions more readily in CSE rats than controls, with concomitant decrease in PaO(2) and increased levels of LPO, HIF-1alpha, and VEGF. Rebamipide reversed gastric lesions without affecting any parameters examined. CSE rats were found to be a useful animal model of COPD, and COPD appeared to render the gastric mucosa vulnerable to prednisolone.