James Pinkerton

Macrolide therapy suppresses key features of experimental steroid-sensitive and steroid-insensitive asthma

Background Steroid-insensitive endotypes of asthma are an important clinical problem and effective therapies are required. They are associated with bacterial infection and non-eosinophilic inflammatory responses in the asthmatic lung. Macrolide therapy is effective in steroid-insensitive endotypes, such as non-eosinophilic asthma. However, whether the effects of macrolides are due to antimicrobial or anti-inflammatory mechanisms is not known. Objective To determine and assess the efficacy of macrolide (ie, clarithromycin) and non-macrolide (ie, amoxicillin) antibiotic treatments in experimental models of infection-induced, severe, steroid-insensitive neutrophilic allergic airways disease (SSIAAD), compared with steroid-sensitive AAD and to delineate the antimicrobial and anti-inflammatory effects of macrolide therapy. Methods We developed and used novel mouse models of Chlamydia and Haemophilus lung infection-induced SSIAAD. We used these models to investigate the effects of clarithromycin and amoxicillin treatment on immune responses and airways hyper-responsiveness (AHR) in Ova-induced, T helper lymphocyte (Th) 2 -associated steroid-sensitive AAD and infection-induced Th1/Th17-associated SSIAAD compared with dexamethasone treatment. Results Clarithromycin and amoxicillin had similar antimicrobial effects on infection. Amoxicillin did attenuate some features, but did not broadly suppress either form of AAD. It did restore steroid sensitivity in SSIAAD by reducing infection. In contrast, clarithromycin alone widely suppressed inflammation and AHR in both steroid-sensitive AAD and SSIAAD. This occurred through reductions in Th2 responses that drive steroid-sensitive eosinophilic AAD and tumour necrosis factor α and interleukin 17 responses that induce SSIAAD. Conclusions Macrolides have broad anti-inflammatory effects in AAD that are likely independent of their antimicrobial effects. The specific responses that are suppressed are dependent upon the responses that dominate during AAD.

Role for NLRP3 Inflammasome-mediated, IL-1β-dependent Responses in Severe, Steroid-resistant Asthma.

&NA; Rationale: Severe, steroid‐resistant asthma is the major unmet need in asthma therapy. Disease heterogeneity and poor understanding of pathogenic mechanisms hampers the identification of therapeutic targets. Excessive nucleotide‐binding oligomerization domain‐like receptor family, pyrin domain‐containing 3 (NLRP3) inflammasome and concomitant IL‐1&bgr; responses occur in chronic obstructive pulmonary disease, respiratory infections, and neutrophilic asthma. However, the direct contributions to pathogenesis, mechanisms involved, and potential for therapeutic targeting remain poorly understood, and are unknown in severe, steroid‐resistant asthma. Objectives: To investigate the roles and therapeutic targeting of the NLRP3 inflammasome and IL‐1&bgr; in severe, steroid‐resistant asthma. Methods: We developed mouse models of Chlamydia and Haemophilus respiratory infection‐mediated, ovalbumin‐induced severe, steroid‐resistant allergic airway disease. These models share the hallmark features of human disease, including elevated airway neutrophils, and NLRP3 inflammasome and IL‐1&bgr; responses. The roles and potential for targeting of NLRP3 inflammasome, caspase‐1, and IL‐1&bgr; responses in experimental severe, steroid‐resistant asthma were examined using a highly selective NLRP3 inhibitor, MCC950; the specific caspase‐1 inhibitor Ac‐YVAD‐cho; and neutralizing anti‐IL‐1&bgr; antibody. Roles for IL‐1&bgr;‐induced neutrophilic inflammation were examined using IL‐1&bgr; and anti‐Ly6G. Measurements and Main Results: Chlamydia and Haemophilus infections increase NLRP3, caspase‐1, IL‐1&bgr; responses that drive steroid‐resistant neutrophilic inflammation and airway hyperresponsiveness. Neutrophilic airway inflammation, disease severity, and steroid resistance in human asthma correlate with NLRP3 and IL‐1&bgr; expression. Treatment with anti‐IL‐1&bgr;, Ac‐YVAD‐cho, and MCC950 suppressed IL‐1&bgr; responses and the important steroid‐resistant features of disease in mice, whereas IL‐1&bgr; administration recapitulated these features. Neutrophil depletion suppressed IL‐1&bgr;‐induced steroid‐resistant airway hyperresponsiveness. Conclusions: NLRP3 inflammasome responses drive experimental severe, steroid‐resistant asthma and are potential therapeutic targets in this disease.

MicroRNA-21 drives severe, steroid-insensitive experimental asthma by amplifying phosphoinositide 3-kinase-mediated suppression of histone deacetylase 2.

Background: Severe steroid‐insensitive asthma is a substantial clinical problem. Effective treatments are urgently required, however, their development is hampered by a lack of understanding of the mechanisms of disease pathogenesis. Steroid‐insensitive asthma is associated with respiratory tract infections and noneosinophilic endotypes, including neutrophilic forms of disease. However, steroid‐insensitive patients with eosinophil‐enriched inflammation have also been described. The mechanisms that underpin infection‐induced, severe steroid‐insensitive asthma can be elucidated by using mouse models of disease. Objective: We sought to develop representative mouse models of severe, steroid‐insensitive asthma and to use them to identify pathogenic mechanisms and investigate new treatment approaches. Methods: Novel mouse models of Chlamydia, Haemophilus influenzae, influenza, and respiratory syncytial virus respiratory tract infections and ovalbumin‐induced, severe, steroid‐insensitive allergic airway disease (SSIAAD) in BALB/c mice were developed and interrogated. Results: Infection induced increases in the levels of microRNA (miRNA)‐21 (miR‐21) expression in the lung during SSIAAD, whereas expression of the miR‐21 target phosphatase and tensin homolog was reduced. This was associated with an increase in levels of phosphorylated Akt, an indicator of phosphoinositide 3‐kinase (PI3K) activity, and decreased nuclear histone deacetylase (HDAC)2 levels. Treatment with an miR‐21–specific antagomir (Ant‐21) increased phosphatase and tensin homolog levels. Treatment with Ant‐21, or the pan‐PI3K inhibitor LY294002, reduced PI3K activity and restored HDAC2 levels. This led to suppression of airway hyperresponsiveness and restored steroid sensitivity to allergic airway disease. These observations were replicated with SSIAAD associated with 4 different pathogens. Conclusion: We identify a previously unrecognized role for an miR‐21/PI3K/HDAC2 axis in SSIAAD. Our data highlight miR‐21 as a novel therapeutic target for the treatment of this form of asthma. GRAPHICAL ABSTRACT Figure. No caption available.

A short-term mouse model that reproduces the immunopathological features of rhinovirus-induced exacerbation of COPD

Viral exacerbations of chronic obstructive pulmonary disease (COPD), commonly caused by rhinovirus (RV) infections, are poorly controlled by current therapies. This is due to a lack of understanding of the underlying immunopathological mechanisms. Human studies have identified a number of key immune responses that are associated with RV-induced exacerbations including neutrophilic inflammation, expression of inflammatory cytokines and deficiencies in innate anti-viral interferon. Animal models of COPD exacerbation are required to determine the contribution of these responses to disease pathogenesis. We aimed to develop a short-term mouse model that reproduced the hallmark features of RV-induced exacerbation of COPD. Evaluation of complex protocols involving multiple dose elastase and lipopolysaccharide (LPS) administration combined with RV1B infection showed suppression rather than enhancement of inflammatory parameters compared with control mice infected with RV1B alone. Therefore, these approaches did not accurately model the enhanced inflammation associated with RV infection in patients with COPD compared with healthy subjects. In contrast, a single elastase treatment followed by RV infection led to heightened airway neutrophilic and lymphocytic inflammation, increased expression of tumour necrosis factor (TNF)-α, C-X-C motif chemokine 10 (CXCL10)/IP-10 (interferon γ-induced protein 10) and CCL5 [chemokine (C-C motif) ligand 5]/RANTES (regulated on activation, normal T-cell expressed and secreted), mucus hypersecretion and preliminary evidence for increased airway hyper-responsiveness compared with mice treated with elastase or RV infection alone. In summary, we have developed a new mouse model of RV-induced COPD exacerbation that mimics many of the inflammatory features of human disease. This model, in conjunction with human models of disease, will provide an essential tool for studying disease mechanisms and allow testing of novel therapies with potential to be translated into clinical practice.

Inflammasomes in the lung

HighlightsActivation of the inflammasome plays a critical role in the innate immune response against noxious stimuli in the lung.Excessive activation of the inflammasome may be associated with the development of chronic lung diseases.Inflammasome‐targeted, site‐specific therapeutics may be beneficial in suppressing inflammasome‐associated disease. Abstract Innate immune responses act as first line defences upon exposure to potentially noxious stimuli. The innate immune system has evolved numerous intracellular and extracellular receptors that undertake surveillance for potentially damaging particulates. Inflammasomes are intracellular innate immune multiprotein complexes that form and are activated following interaction with these stimuli. Inflammasome activation leads to the cleavage of pro‐IL‐1&bgr; and release of the pro‐inflammatory cytokine, IL‐1&bgr;, which initiates acute phase pro‐inflammatory responses, and other responses are also involved (IL‐18, pyroptosis). However, excessive activation of inflammasomes can result in chronic inflammation, which has been implicated in a range of chronic inflammatory diseases. The airways are constantly exposed to a wide variety of stimuli. Inflammasome activation and downstream responses clears these stimuli. However, excessive activation may drive the pathogenesis of chronic respiratory diseases such as severe asthma and chronic obstructive pulmonary disease. Thus, there is currently intense interest in the role of inflammasomes in chronic inflammatory lung diseases and in their potential for therapeutic targeting. Here we review the known associations between inflammasome‐mediated responses and the development and exacerbation of chronic lung diseases.

NLRP3 inflammasome-mediated, IL-1 beta-dependent inflammatory responses drive severe, steroid-insensitive asthma

Introduction: Studies suggest that 10% of children with an ARI have persistent cough at day 21. There are no studies in Indigenous children who have a high risk of chronic lung disease. We aimed to identify the incidence and outcomes of ARI with cough as a symptom in urban Indigenous children. Methods:This is a prospective study of Indigenous children aged<5 years registered with a primary health service. Children are followed for a period of 12 months via monthly contacts. Children who develop cough as a symptom at any time are followed weekly for 4 weeks to ascertain cough outcomes. Results: To date, 162 children are enrolled, totalling 1065 child-months of observation. Two-hundred ARI episodes with cough have been reported (29.6 episodes/100 child-months at risk). Thirty-four ARIs (17%) have progressed to persistent cough at day 28 in 24 children. Of these, 15 children had 1 episode, 5 had 2, 4 had 3 and 1 had 4 during the follow-up period. The majority of children with persistent cough were diagnosed (by a respiratory physician) with protracted bacterial bronchitis and/or bronchiectasis. Conclusions:The proportion of children developing persistent cough postARI is higher than that currently reported (10%) with the majority suggesting protracted bacterial infection. GrantSupport:AQUTAPAaward, aQCMRIProgramGrant, UQFoundation Research Excellence Award, a QUT Indigenous Health Research StartUpGrant and the NHMRCCRE for LungHealth in Aboriginal and TorresStrait Islander Children. Declaration of Interest: None to Declare

A Sheep Model to Examine the Effects of Maternal Asthma On Fetal Outcomes

Introduction: Studies suggest that 10% of children with an ARI have persistent cough at day 21. There are no studies in Indigenous children who have a high risk of chronic lung disease. We aimed to identify the incidence and outcomes of ARI with cough as a symptom in urban Indigenous children. Methods:This is a prospective study of Indigenous children aged<5 years registered with a primary health service. Children are followed for a period of 12 months via monthly contacts. Children who develop cough as a symptom at any time are followed weekly for 4 weeks to ascertain cough outcomes. Results: To date, 162 children are enrolled, totalling 1065 child-months of observation. Two-hundred ARI episodes with cough have been reported (29.6 episodes/100 child-months at risk). Thirty-four ARIs (17%) have progressed to persistent cough at day 28 in 24 children. Of these, 15 children had 1 episode, 5 had 2, 4 had 3 and 1 had 4 during the follow-up period. The majority of children with persistent cough were diagnosed (by a respiratory physician) with protracted bacterial bronchitis and/or bronchiectasis. Conclusions:The proportion of children developing persistent cough postARI is higher than that currently reported (10%) with the majority suggesting protracted bacterial infection. GrantSupport:AQUTAPAaward, aQCMRIProgramGrant, UQFoundation Research Excellence Award, a QUT Indigenous Health Research StartUpGrant and the NHMRCCRE for LungHealth in Aboriginal and TorresStrait Islander Children. Declaration of Interest: None to Declare