Ozone-induced COPD mouse model reflects the detrimental effects of aging as well as ozone on the lung

( So Ri Kim ),( Yong Chul Lee ),( Kyeong Hwa Park ),( Hae Jin Park ),( Navin Ray ),( Yeong Hun Choe ),( Seung Yong Park ),( Jae Seok Jeong ) 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

To understand the pathogenesis of chronic obstructive pulmonary diseases (COPD) and to screen new drug targets, the valid animal model is required. A cigarette smoke (CS)-induced model remains the most popular one. However, it usually requires 3-12 months to induce the COPD animal model by CS which is such a time-consuming and labor-intensive process. In this study, we aimed to establish the protocol for the ozone-induced COPD murine model and to differentiate the aging effects from the real ozone-induced detrimental effects on the lung in the mice. Ozone-induced COPD murine model was established by the exposure to 3 ppm of ozone twice a week for 7 weeks. Time-course analysis showed that the number of BAL cells was increased in air-exposed mice and ozone-exposed mice at the same age (i.e.,14 weeks) compared to young air-exposed mice (i.e.,6-7 weeks). The expression of TGF-β, IL-17, and IL-1β is gradually increased as ozone-exposed time goes by up to 7 weeks (TGF-β, IL-17) and 6 weeks (IL-1β), respectively. In addition, ozone-exposed mice for 7 weeks revealed that decreased lung function (FEV0.1) and increased alveolar destruction in lung tissues compared to air-exposed mice. Interestingly, old air-exposed mice (14 weeks) showed more severe lung destruction than young aged air-exposed mice (6-7 weeks). These findings suggest that ozone-exposed mice can be used for preclinical experiments regarding COPD as a better choice than CS-induced models and that this mouse model reflects the detrimental effects of aging as well as ozone on the lung more physiologically.

MCC 950 ameliorates house dust mite-induced asthma through the regulation of ASC oligomerization and NEK7-NLRP3 binding in NLRP3 inflammasome assembly

( Yong Chul Lee ),( Jae Seok Jeong ),( So Ri Kim ),( Hae Jin Park ),( Kyeong Hwa Park ),( Navin Ray ),( Yeong Hun Choe ),( Seung Yong Park ) 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

NLRP3 inflammasome, consisting of NLRP3, the adaptor protein ASC, and the protease caspase-1, is responsible for the production of active forms of IL-1β and IL-18. In this process, oligomerization of ASC is a key step in assembly/activation of inflammasome and, recently, NEK7, a serine and threonine kinase, has been also reported as an essential activator of the NLRP3 inflammasome. MCC 950 is a small-molecule inhibitor of NLRP3 inflammasome, however, molecular action mechanism of MCC 950 is not fully understood. We investigated the therapeutic effects of the MCC 950 and its action mechanism in house dust mite (HDM)-induced allergic lung inflammation, particularly focusing on the NLRP3 inflammasome assembly process involving ASC and NEK7. Respiratory HDM exposure into mice led to the significant increases of pulmonary NLRP3, caspase-1, and IL-1β. Furthermore, levels of ASC oligomers were elevated in lung tissues of HDM-exposed mice and we observed the cytoplasmic co-localization of immunofluorescence intensities of NLRP3 and NEK7 in bronchoalveolar lavage (BAL) cells. Notably, treatment with MCC 950 significantly reduced the HDM-induced increases of ASC oligomerization and NLRP3-NEK7 colocalization in the lung of mice, and that ameliorated the HDM-induced increases of airway inflammatory cells infiltration, airway hyper-reactivity, and pulmonary TH2 cytokines. These results suggest that MCC 950 may have potential for treating HDM-induced allergic asthma partly through the regulation of HDM-induced ASC oligomerization and NEK7-NLRP3 binding in NLRP3 inflammasome assembly.

Comparison of the therapeutic effects of dexamethasone on two different murine models of fungus-induced allergic lung inflammation

( Jae Seok Jeong ),( Yong Chul Lee ),( So Ri Kim ),( Hae Jin Park ),( Kyeong Hwa Park ),( Navin Ray ),( Yeong Hun Choe ),( Seung Yong Park ) 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

Fungal asthma endotype presents severe disease with frequent exacerbations and eosinophilic inflammation. The most severe form of allergic fungal disease is known to be associated with Aspergillus fumigatus (Af). Oxidative stress is involved in the development of corticosteroid (CS) resistance in chronic airway disorders and our previous study showed that mitochondrial reactive oxygen species (mtROS) generation is implicated in CS-resistant lung inflammation. Herein, we compared effects of dexamethasone on various features of fungal allergic lung inflammation between Af- and Alternaria alternata (Aa)-induced forms, and that investigated molecular basis of the fungus-induced CS resistance. Results showed that dexamethasone improved the Aa-induced eosinophilic lung inflammation and airway hyper-responsiveness, while it failed to attenuate Af-induced allergic lung inflammation. In addition, dexamethasone reduced Aa-induced increases in the fluorescence intensity of mtROS in bronchoalveolar lavage (BAL) cells, whereas it failed to reduce Af-induced increase in mtROS generation. Furthermore, while dexamethasone reduced Aa-induced increases in the nuclear translocation of nuclear factor (NF)-κB p65, it failed to lower Af-induced increases in NF-κB p65 in nuclear protein extracts of lung tissues. These findings suggest that difference in ability of dexamethasone to control inflammation in two murine models of fungal allergic lung inflammation is partly dependent on whether CS can regulate fungi-induced mtROS generation and related activation of NF-κB.

Pharmacologic effects of a novel PI3K-δ inhibitor, YH25487 on steroid-resistant eosinophilic asthma in mice

( So Ri Kim ),( Yong Chul Lee ),( Wonee Chong ),( Kyoung Kyu Ahn ),( Soongyu Choi ),( Hae Jin Park ),( Kyeong Hwa Park ),( Navin Ray ),( Yeong Hun Choe ),( Seung Yong Park ),( Jae Seok Jeong ) 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

Phosphoinositide 3-kinase (PI3K)-δ-dependent Akt activation play critical roles in various immune responses and PI3K-δ isoform is principally expressed in leukocytes. The restricted expression profile of PI3K-δ makes it an attractive drug target for various inflammatory conditions such as chronic airway disorders. Nowadays, it is believed that PI3K-δ contributes to the steroid resistance of severe respiratory diseases and more specifically that oxidative stress directly induces PI3K-δ-dependent Akt activation. With understanding the role of PI3K-δ in airway inflammatory disorders, recent developed PI3K-δ inhibitors are applying to control bronchial asthma or COPD in preclinical and clinical trials. In this study, we aimed to define the pharmacologic effects of YH25487, a newly developed PI3K-δ selective inhibitor on the steroid-resistant eosinophilic asthma using Aspergillus fumigatus (Af)-sensitized and -challenged mice. Our results showed that the intratracheal administration of YH25487 attenuated the steroid-resistant asthmatic manifestations including inflammatory signs and airway hyper-responsiveness in Af-exposed mice. Moreover, when the pharmacologic effects are compared to the positive control GSK2269557, we found that the anti-asthmatic effects of YH25487 are superior to those of GSK2269557 in the severe eosinophilic asthma animal model. These findings suggest that a novel compound targeting PI3K-δ-isoform, YH25487 can be a promising therapeutic agent for the severe eosinophilic asthma, specifically as an inhaled formulation.

Effects of phosphoinositide 3-kinase (PI3K)-δ blockade on various features of viable Aspergillus fumigatus (Af) conidium-induced allergic lung inflammation

( Yong Chul Lee ),( Jae Seok Jeong ),( So Ri Kim ),( Hae Jin Park ),( Kyeong Hwa Park ),( Navin Ray ),( Yeong Hun Choe ),( Seung Yong Park ) 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

Delta (δ) isoform of phosphoinositide 3-kinases (PI3Ks) may be one of important therapeutic targets for treating inflammatory diseases. However, there are also concerns on PI3K-δ blockade due to its potential to disturb protective immune responses. We evaluated whether PI3K-δ inhibition can affect on the progression of fungus-associated inflammation/infection in an experimental fungal allergic lung inflammation established through respiratory Aspergillus fumigatus (Af) conidium exposure. Respiratory exposure of Af led to the increases in the numbers of total cells, neutrophils, lymphocytes, and particularly eosinophils, in bronchoalveolar lavage (BAL) fluids. Levels of TH2 cytokines in lung tissues of Af-exposed mice were increased. IC87114, a potent inhibitor of PI3K-δ, significantly lowered the Af-induced increases of BAL cells, particularly eosinophils, and IL-4, IL-5, and IL-13 in the lung, while treatment with itraconazole failed to ameliorate. Concurrent treatment of IC87114 with itraconazole led to a tendency toward a better improvement of Af-induced allergic lung inflammation than IC87114 alone, although it was not statistically significant. We could not observe any significant increase in fungal colonization/infection associated with IC87114 in the lung and the numbers of neutrophils were not significantly changed in BAL fluids from Af-exposed mice. These data suggest that PI3K-δ blockade is effective for controlling Af-induced allergic lung inflammation without significant progression of inflammation and the infection in the current experimental system.

Phosphoinositide 3-kinase-δ influences fungi-induced NLRP3 inflammasome assembly/activation in human bronchial epithelial cells

( Yong Chul Lee ),( Jae Seok Jeong ),( So Ri Kim ),( Hae Jin Park ),( Kyeong Hwa Park ),( Navin Ray ),( Yeong Hun Choe ),( Seung Yong Park ) 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

Respiratory fungal exposure is associated with severe allergic lung inflammation. NLRP3 inflammasome and phosphoinositide 3-kinase (PI3K)-δ in airway epithelium are involved in various inflammatory processes. We previously reported that fungus- induced activation of PI3K-δ in airway epithelium leads to generation of mitochondrial reactive oxygen species (mtROS), thereby being involved in the corticosteroid resistance in fungal allergy model. In this study, we performed in vitro experiments using normal human bronchial epithelial (NHBE) cells to investigate the role of PI3K-δ in modulating NLRP3 inflammasome activation against fungal exposure, especially focusing on mtROS. We also checked NLRP3 expression in lung tissues from allergic bronchopulmonary aspergillosis (ABPA) patients. NLRP3, caspase-1, and ASC and their cytoplasmic co-localization were increased in Af-stimulated NHBE cells. Furthermore, mature IL-1β were increased in Af-stimulated NHBE cells. Notably, Af-induced increases of NLRP3 inflammasome components and their co-localizations were lowered by a potent PI3K-δ inhibitor or PI3K-δ specific siRNA. This modulatory role of PI3K-δ was mediated through the regulation of mtROS generation. In human lung tissue specimens, NLRP3 in ABPA patients was increased compared to that in disease control (idiopathic pulmonary fibrosis) or healthy controls. These findings suggest that fungi-induced activation of NLRP3 inflammasome in airway epithelium may be modulated by PI3K-δ, which is mediated partly through the regulation of mtROS.

Comparison of the severity of lung inflammation and the role of phosphoinositide 3-kinase-δ in Aspergillus fumigatus (Af)-induced allergic lung inflammation between male and female mice

( Yong Chul Lee ),( Jae Seok Jeong ),( So Ri Kim ),( Hae Jin Park ),( Kyeong Hwa Park ),( Navin Ray ),( Yeong Hun Choe ),( Seung Yong Park ) 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

Females are known to be more susceptible to many immunological disorders in humans. Several immunologic features in female gender have been suggested as the cause of this phenomenon. As for allergic lung inflammation, it has been reported that female mice are also more susceptible to the development of allergic inflammation against a typical experimental aeroallergen, ovalbumin, than male mice. We investigated whether there is also a gender gap regarding the degree of immune response against a potent fungal allergen, Aspergillus fumigatus (Af) in mice. Respiratory Af exposure led to the increases in the numbers of total cells, eosinophils, and lymphocytes in bronchoalveolar lavage (BAL) fluids from both genders compared to those of control mice. Furthermore, inhibition of phosphoinositide 3-kinase (PI3K)-δ significantly reduced the Af-induced increases in the numbers of total cells and eosinophils in BAL fluids from both male and female mice. Interestingly, female mice showed a tendency toward the more severe allergic lung inflammation than male mice. Intratracheal administration of IC87114, a potent inhibitor of PI3K-δ, into mice remarkably ameliorated the Af-induced increases of airway inflammatory cells infiltration, airway hyper-responsiveness, and pulmonary TH2 cytokines (IL-4, IL-5, and IL-13) in the lung. These findings suggest the presence of a gender gap in the severity of allergic lung inflammation against Af and PI3K-δ may play a key role in Af-induced allergic lung inflammation regardless of gender in mice.

Therapeutic potential of an indole derivative on murine models of environmental compound-associated lung injury

( Yong Chul Lee ),( Jae Seok Jeong ),( So Ri Kim ),( Hae Jin Park ),( Kyeong Hwa Park ),( Navin Ray ),( Yeong Hun Choe ),( Seung Yong Park ) 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

Various environmental compounds are implicated in fatal lung injury. Mitochondria are crucial organelles for normal lung function that can be impacted by many lung diseases. Recently, we developed an antioxidant indole-derived NecroX compound (NecroX), which preserves mitochondrial functionalities. In this study, we investigated therapeutic effects of NecroX on lung injury associated with polyhexamethylene guanidine (PHMG), a well-known chemical compound implicated in humidifier disinfectant- associated fatal lung injury in human, and bleomycin, a widely used chemical for inducing experimental lung fibrosis in mice, focusing on functionalities of mitochondria. Respiratory exposure to PHMG and bleomycin led to lung injury manifesting inflammation (increases of inflammatory cell infiltrations, TNF-α, IL-1β, IL-17, and KC) followed by fibrosis (elevated total collagen amount and TGF-β1) in the lung parenchyma, which was further verified by histopathologic and radiologic measurements. Exposure to these compounds impacted on mitochondria in regard to biogenesis, mitochondrial DNA (mtDNA) integrity, and generation of mitochondrial reactive oxygen species (mtROS) in various cells of the lung. Notably, NecroX significantly improved these pathobiologic features of the PHMG- and bleomycin-induced lung injury and ameliorated mitochondrial functionalities. These findings imply that NecroX has therapeutic potential in the treatment of environmental compound- induced lung injury.

Role of PI3K-δ-HIF-VEGF axis in the pathogenesis of sinonasal inverted papilloma

( Jae Seok Jeong ),( Yong Chul Lee ),( So Ri Kim ),( Jong Seung Kim ),( Hae Jin Park ),( Navin Ray ),( Kyeong Hwa Park ),( Yeong Hun Choe ),( Seung Yong Park ) 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

Sinonasal inverted papilloma (IP) is associated with the recurrent disease and invasion to adjacent tissues. In addition, IP has been regarded as a premalignant lesion, however, not enough information exists on the pathogenesis of the disease. Vascular endothelial growth factor (VEGF) is one of crucial mediators in airway inflammation in part through the regulation of vascular permeability. We previously demonstrated for the first time that vascular leakage and subsequent inflammation of airways can be modulated by PI3K-hypoxia-inducible factor (HIF)-1α-VEGF axis. Herein, we investigated the role of this axis in the pathogenesis of sinonasal IP. IP tissues were obtained from 10 patients during the endoscopic sinus surgery and inferior turbinate tissues as healthy control were obtained from 10 control subjects during septoplasty or skull base surgery. Western blot analyses showed that protein expressions of catalytic subunit of PI3K-δ (p110δ) and phosphorylated AKT, a key downstream mediator of PI3K-δ, were notably elevated in IP tissues compared to those in controls. Furthermore, confocal microscopic analysis demonstrated that expression of p110δ was prominent in the epithelial and submucosal layers, which substantially co-localized to VEGF and HIF-1α in sinonasal IP tissues. Expression levels of VEGF and HIF-1α protein were strongly correlated with p110δ protein levels in these tissues. These data suggest that PI3K-δ-HIF-VEGF axis may play a role in the pathogenesis of sinonasal inverted papilloma, in part through modulating vascular leakage and plasma exudation.

Involvement of phosphoinositide 3-kinase-delta in the development of eosinophilic nasal polyps

( Yong Chul Lee ),( Jae Seok Jeong ),( So Ri Kim ),( Jong Seung Kim ),( Hae Jin Park ),( Kyeong Hwa Park ),( Navin Ray ),( Yeong Hun Choe ),( Seung Yong Park ) 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

Nasal polyps (NP) cause diverse clinical symptoms of chronic rhinosinusitis (CRS). Chronic inflammation of sinonasal mucosa is known to be crucial in NP formation. We aimed to define the implications of phosphoinositide 3-kinase (PI3K)-δ in nasal inflammation associated with NP by analyzing NP tissue obtained from CRS patients. Results showed that expression of p110δ, a regulatory subunit of PI3K-δ, in NP tissue was increased compared to control tissue. Increased p110δ expression was closely correlated with more severe CRS features. Interestingly, p110δ expression was increased in eosinophilic NP, which are closely related to more complicated clinical courses of the disease. Furthermore, CRS patients possessing NP with higher p110δ expression displayed more eosinophils in NP tissue and blood, higher levels of IL-5 in NP tissue, and more severe features of the disease. Therefore, PI3K-δ may contribute to the formation of NP, especially eosinophilic NP associated with more severe clinical presentations and radiological features.