Early-life house dust mite aeroallergen exposure augments cigarette smoke-induced myeloid inflammation and emphysema in mice.

BACKGROUND: Longitudinal studies have identified childhood asthma as a risk factor for obstructive pulmonary disease (COPD) and asthma-COPD overlap (ACO) where persistent airflow limitation can develop more aggressively. However, a causal link between childhood asthma and COPD/ACO remains to be established. Our study aimed to model the natural history of childhood asthma and COPD and to investigate the cellular/molecular mechanisms that drive disease progression. METHODS: Allergic airways disease was established in three-week-old young C57BL/6 mice using house dust mite (HDM) extract. Mice were subsequently exposed to cigarette smoke (CS) and HDM for 8 weeks. Airspace enlargement (emphysema) was measured by the mean linear intercept method. Flow cytometry was utilised to phenotype lung immune cells. Bulk RNA-sequencing was performed on lung tissue. Volatile organic compounds (VOCs) in bronchoalveolar lavage-fluid were analysed to screen for disease-specific biomarkers. RESULTS: Chronic CS exposure induced emphysema that was significantly augmented by HDM challenge. Increased emphysematous changes were associated with more abundant immune cell lung infiltration consisting of neutrophils, interstitial macrophages, eosinophils and lymphocytes. Transcriptomic analyses identified a gene signature where disease-specific changes induced by HDM or CS alone were conserved in the HDM-CS group, and further revealed an enrichment of Mmp12, Il33 and Il13, and gene expression consistent with greater expansion of alternatively activated macrophages. VOC analysis also identified four compounds increased by CS exposure that were paradoxically reduced in the HDM-CS group. CONCLUSIONS: Early-life allergic airways disease worsened emphysematous lung pathology in CS-exposed mice and markedly alters the lung transcriptome.

Low dose aspirin prevents endothelial dysfunction in the aorta and foetal loss in pregnant mice infected with influenza A virus.

Influenza A virus (IAV) infection in pregnancy resembles a preeclamptic phenotype characterised by vascular dysfunction and foetal growth retardation. Given that low dose aspirin (ASA) is safe in pregnancy and is used to prevent preeclampsia, we investigated whether ASA or NO-conjugated aspirin, NCX4016, resolve vascular inflammation and function to improve offspring outcomes following IAV infection in pregnant mice. Pregnant mice were intranasally infected with a mouse adapted IAV strain (Hkx31; 104 plaque forming units) and received daily treatments with either 200µg/kg ASA or NCX4016 via oral gavage. Mice were then culled and the maternal lungs and aortas collected for qPCR analysis, and wire myography was performed on aortic rings to assess endothelial and vascular smooth muscle functionality. Pup and placentas were weighed and pup growth rates and survival assessed. IAV infected mice had an impaired endothelial dependent relaxation response to ACh in the aorta, which was prevented by ASA and NCX4016 treatment. ASA and NCX4016 treatment prevented IAV dissemination and inflammation of the aorta as well as improving the pup placental ratios in utero, survival and growth rates at post-natal day 5. Low dose ASA is safe to use during pregnancy for preeclampsia and this study demonstrates that ASA may prove a promising treatment for averting the significant vascular complications associated with influenza infection during pregnancy.

Musculoskeletal crosstalk in chronic obstructive pulmonary disease and comorbidities: Emerging roles and therapeutic potentials.

Chronic Obstructive Pulmonary Disease (COPD) is a multifaceted respiratory disorder characterized by progressive airflow limitation and systemic implications. It has become increasingly apparent that COPD exerts its influence far beyond the respiratory system, extending its impact to various organ systems. Among these, the musculoskeletal system emerges as a central player in both the pathogenesis and management of COPD and its associated comorbidities. Muscle dysfunction and osteoporosis are prevalent musculoskeletal disorders in COPD patients, leading to a substantial decline in exercise capacity and overall health. These manifestations are influenced by systemic inflammation, oxidative stress, and hormonal imbalances, all hallmarks of COPD. Recent research has uncovered an intricate interplay between COPD and musculoskeletal comorbidities, suggesting that muscle and bone tissues may cross-communicate through the release of signalling molecules, known as "myokines" and "osteokines". We explored this dynamic relationship, with a particular focus on the role of the immune system in mediating the cross-communication between muscle and bone in COPD. Moreover, we delved into existing and emerging therapeutic strategies for managing musculoskeletal disorders in COPD. It underscores the development of personalized treatment approaches that target both the respiratory and musculoskeletal aspects of COPD, offering the promise of improved well-being and quality of life for individuals grappling with this complex condition. This comprehensive review underscores the significance of recognizing the profound impact of COPD on the musculoskeletal system and its comorbidities. By unravelling the intricate connections between these systems and exploring innovative treatment avenues, we can aspire to enhance the overall care and outcomes for COPD patients, ultimately offering hope for improved health and well-being.

Targeting accelerated pulmonary ageing to treat chronic obstructive pulmonary disease-induced neuropathological comorbidities.

Chronic obstructive pulmonary disease (COPD) is a major incurable health burden, ranking as the third leading cause of death worldwide, mainly driven by cigarette smoking. COPD is characterised by persistent airway inflammation, lung function decline and premature ageing with the presence of pulmonary senescent cells. This review proposes that cellular senescence, a state of stable cell cycle arrest linked to ageing, induced by inflammation and oxidative stress in COPD, extends beyond the lungs and affects the systemic circulation. This pulmonary senescent profile will reach other organs via extracellular vesicles contributing to brain inflammation and damage, and increasing the risk of neurological comorbidities, such as stroke, cerebral small vessel disease and Alzheimer's disease. The review explores the role of cellular senescence in COPD-associated brain conditions and investigates the relationship between cellular senescence and circadian rhythm in COPD. Additionally, it discusses potential therapies, including senomorphic and senolytic treatments, as novel strategies to halt or improve the progression of COPD.

Dual inhibition of airway inflammation and fibrosis by common ß cytokine receptor blockade.

BACKGROUND: Patients with severe asthma can present with eosinophilic type 2 (T2), neutrophilic, or mixed inflammation that drives airway remodeling and exacerbations and represents a major treatment challenge. The common ß (ßc) receptor signals for 3 cytokines, GM-CSF, IL-5, and IL-3, which collectively mediate T2 and neutrophilic inflammation. OBJECTIVE: To determine the pathogenesis of ßc receptor-mediated inflammation and remodeling in severe asthma and to investigate ßc antagonism as a therapeutic strategy for mixed granulocytic airway disease. METHODS: ßc gene expression was analyzed in bronchial biopsy specimens from patients with mild-to-moderate and severe asthma. House dust mite extract and Aspergillus fumigatus extract (ASP) models were used to establish asthma-like pathology and airway remodeling in human ßc transgenic mice. Lung tissue gene expression was analyzed by RNA sequencing. The mAb CSL311 targeting the shared cytokine binding site of ßc was used to block ßc signaling. RESULTS: ßc gene expression was increased in patients with severe asthma. CSL311 potently reduced lung neutrophils, eosinophils, and interstitial macrophages and improved airway pathology and lung function in the acute steroid-resistant house dust mite extract model. Chronic intranasal ASP exposure induced airway inflammation and fibrosis and impaired lung function that was inhibited by CSL311. CSL311 normalized the ASP-induced fibrosis-associated extracellular matrix gene expression network and strongly reduced signatures of cellular inflammation in the lung. CONCLUSIONS: ßc cytokines drive steroid-resistant mixed myeloid cell airway inflammation and fibrosis. The anti-ßc antibody CSL311 effectively inhibits mixed T2/neutrophilic inflammation and severe asthma-like pathology and reverses fibrosis gene signatures induced by exposure to commonly encountered environmental allergens.

Assessing Lung Inflammation and Pathology in Preclinical Models of Chronic Obstructive Pulmonary Disease.

Chronic obstructive pulmonary disease (COPD) is an incurable disease that is a major cause of mortality and morbidity worldwide. Cigarette smoking is a major cause of COPD and triggers progressive airflow limitation, chronic lung inflammation, and irreversible lung damage and decline in lung function. COPD patients often experience various extrapulmonary comorbid diseases, including cardiovascular disease, skeletal muscle wasting, lung cancer, and cognitive decline which markedly impact on disease morbidity, progression, and mortality. People with COPD are also susceptible to respiratory infections which cause exacerbations of the underlying disease (AECOPD). The mechanisms and mediators underlying COPD and its comorbidities are poorly understood and current COPD therapy is relatively ineffective. We and others have used animal modelling systems to explore the mechanisms underlying COPD, AECOPD, and comorbidities of COPD with the goal of identifying novel therapeutic targets. Here we provide a preclinical model and protocols to assess the cellular, molecular, and pathological consequences of cigarette smoke exposure and the development of comorbidities of COPD.

A promising new oral delivery mode for insulin using lipid-filled enteric-coated capsules.

The treatment of diabetes requires daily administration of the peptide insulin via subcutaneous (SC) injection due to poor stability following oral administration. Enteric capsules, designed to protect against low pH conditions in the stomach by providing a polymeric coating which only breaks down in the small intestine, have failed to significantly increase oral bioavailability for insulin. In parallel, amphiphilic lipid mesophases are versatile carrier materials which can protect encapsulated proteins and peptides from undesirable enzymatic degradation. Here we show the combined delivery capacity of a hydrated bicontinuous cubic lipid mesophase embedded within an enteric capsule. Animal studies demonstrated that the lipid filled enteric capsules could deliver insulin with bioavailabilities (relative to SC injection) as high as 99 % and 150 % for fast and slow acting insulin, respectively. These results provide a promising starting point towards further trials to develop an alternative, non-invasive mode for the delivery of insulin.

Inhibition of oxidative stress by apocynin attenuated chronic obstructive pulmonary disease progression and vascular injury by cigarette smoke exposure.

BACKGROUND AND PURPOSE: Cardiovascular disease affects up to half of the patients with chronic obstructive pulmonary disease (COPD), exerting deleterious impact on health outcomes and survivability. Vascular endothelial dysfunction marks the onset of cardiovascular disease. The present study examined the effect of a potent NADPH Oxidase (NOX) inhibitor and free-radical scavenger, apocynin, on COPD-related cardiovascular disease. EXPERIMENTAL APPROACH: Male BALB/c mice were exposed to either room air (Sham) or cigarette smoke (CS) generated from 9 cigarettes·day-1 , 5 days a week for up to 24 weeks with or without apocynin treatment (5 mg·kg-1 ·day-1 , intraperitoneal injection). KEY RESULTS: Eight-weeks of apocynin treatment reduced airway neutrophil infiltration (by 42%) and completely preserved endothelial function and endothelial nitric oxide synthase (eNOS) availability against the oxidative insults of cigarette smoke exposure. These preservative effects were maintained up until the 24-week time point. 24-week of apocynin treatment markedly reduced airway inflammation (reduced infiltration of macrophage, neutrophil and lymphocyte), lung function decline (hyperinflation) and prevented airway collagen deposition by cigarette smoke exposure. CONCLUSION AND IMPLICATIONS: Limiting NOX activity may slow COPD progression and lower cardiovascular disease risk, particularly when signs of oxidative stress become evident.

Cigarette smoke-induced pulmonary impairment is associated with social recognition memory impairments and alterations in microglial profiles within the suprachiasmatic nucleus of the hypothalamus.

Chronic obstructive pulmonary disease (COPD) is a major, incurable respiratory condition that is primarily caused by cigarette smoking (CS). Neurocognitive disorders including cognitive dysfunction, anxiety and depression are highly prevalent in people with COPD. It is understood that increased lung inflammation and oxidative stress from CS exposure may 'spill over' into the systemic circulation to promote the onset of these extra-pulmonary comorbidities, and thus impacts the quality of life of people with COPD. The precise role of the 'spill-over' of inflammation and oxidative stress in the onset of COPD-related neurocognitive disorders are unclear. The present study investigated the impact of chronic CS exposure on anxiety-like behaviors and social recognition memory, with a particular focus on the role of the 'spill-over' of inflammation and oxidative stress from the lungs. Adult male BALB/c mice were exposed to either room air (sham) or CS (9 cigarettes per day, 5 days a week) for 24 weeks and were either daily co-administered with the NOX2 inhibitor, apocynin (5 mg/kg, in 0.01 % DMSO diluted in saline, i.p.) or vehicle (0.01 % DMSO in saline) one hour before the initial CS exposure of the day. After 23 weeks, mice underwent behavioral testing and physiological diurnal rhythms were assessed by monitoring diurnal regulation profiles. Lungs were collected and assessed for hallmark features of COPD. Consistent with its anti-inflammatory and oxidative stress properties, apocynin treatment partially lessened lung inflammation and lung function decline in CS mice. CS-exposed mice displayed marked anxiety-like behavior and impairments in social recognition memory compared to sham mice, which was prevented by apocynin treatment. Apocynin was unable to restore the decreased Bmal1-positive cells, key in cells in diurnal regulation, in the suprachiasmatic nucleus of the hypothalamus to that of sham levels. CS-exposed mice treated with apocynin was associated with a restoration of microglial area per cell and basal serum corticosterone. This data suggests that we were able to model the CS-induced social recognition memory impairments seen in humans with COPD. The preventative effects of apocynin on memory impairments may be via a microglial dependent mechanism.

Influenza Virus Infection during Pregnancy as a Trigger of Acute and Chronic Complications.

Influenza A virus (IAV) infection during pregnancy disrupts maternal and fetal health through biological mechanisms, which are to date poorly characterised. During pregnancy, the viral clearance mechanisms from the lung are sub-optimal and involve hyperactive innate and adaptive immune responses that generate wide-spread inflammation. Pregnancy-related adaptations of the immune and the cardiovascular systems appear to result in delayed recovery post-viral infection, which in turn promotes a prolonged inflammatory phenotype, increasing disease severity, and causing maternal and fetal health problems. This has immediate and long-term consequences for the mother and fetus, with complications including acute cardiopulmonary distress syndrome in the mother that lead to perinatal complications such as intrauterine growth restriction (IUGR), and birth defects; cleft lip, cleft palate, neural tube defects and congenital heart defects. In addition, an increased risk of long-term neurological disorders including schizophrenia in the offspring is reported. In this review we discuss the pathophysiology of IAV infection during pregnancy and its striking similarity to other well-established complications of pregnancy such as preeclampsia. We discuss general features of vascular disease with a focus on vascular inflammation and define the "Vascular Storm" that is triggered by influenza infection during pregnancy, as a pivotal disease mechanism for short and long term cardiovascular complications.

Influenza A virus elicits peri-vascular adipose tissue inflammation and vascular dysfunction of the aorta in pregnant mice.

Influenza A virus (IAV) infection during pregnancy initiates significant aortic endothelial and vascular smooth muscle dysfunction, with inflammation and T cell activation, but the details of the mechanism are yet to be clearly defined. Here we demonstrate that IAV disseminates preferentially into the perivascular adipose tissue (PVAT) of the aorta in mice. IAV mRNA levels in the PVAT increased at 1-3 days post infection (d.p.i) with the levels being ~4-8 fold higher compared with the vessel wall. IAV infection also increased Ly6Clow patrolling monocytes and Ly6Chigh pro-inflammatory monocytes in the vessel wall at 3 d.p.i., which was then followed by a greater homing of these monocytes into the PVAT at 6 d.p.i. The vascular immune phenotype was characteristic of a "vascular storm"- like response, with increases in neutrophils, pro-inflammatory cytokines and oxidative stress markers in the PVAT and arterial wall, which was associated with an impairment in endothelium-dependent relaxation to acetylcholine. IAV also triggered a PVAT compartmentalised elevation in CD4+ and CD8+ activated T cells. In conclusion, the PVAT of the aorta is a niche that supports IAV dissemination and a site for perpetuating a profound innate inflammatory and adaptive T cell response. The manifestation of this inflammatory response in the PVAT following IAV infection may be central to the genesis of cardiovascular complications arising during pregnancy.

Glycolysis and the Pentose Phosphate Pathway Promote LPS-Induced NOX2 Oxidase- and IFN-ß-Dependent Inflammation in Macrophages.

Macrophages undergo a metabolic switch from oxidative phosphorylation to glycolysis when exposed to gram-negative bacterial lipopolysaccharide (LPS), which modulates antibacterial host defence mechanisms. Here, we show that LPS treatment of macrophages increased the classical oxidative burst response via the NADPH oxidase (NOX) 2 enzyme, which was blocked by 2-deoxyglucose (2-DG) inhibition of glycolysis. The inhibition of the pentose phosphate pathway with 6-aminonicotinamide (6-AN) also suppressed the LPS-induced increase in NOX2 activity and was associated with a significant reduction in the mRNA expression of NOX2 and its organizer protein p47phox. Notably, the LPS-dependent enhancement in NOX2 oxidase activity was independent of both succinate and mitochondrial reactive oxygen species (ROS) production. LPS also increased type I IFN-ß expression, which was suppressed by 2-DG and 6-AN and, therefore, is dependent on glycolysis and the pentose phosphate pathway. The type I IFN-ß response to LPS was also inhibited by apocynin pre-treatment, suggesting that NOX2-derived ROS promotes the TLR4-induced response to LPS. Moreover, recombinant IFN-ß increased NOX2 oxidase-dependent ROS production, as well as NOX2 and p47phox expression. Our findings identify a previously undescribed molecular mechanism where both glycolysis and the pentose phosphate pathway are required to promote LPS-induced inflammation in macrophages.

Cross-talk between IL-6 trans-signaling and AIM2 inflammasome/IL-1ß axes bridge innate immunity and epithelial apoptosis to promote emphysema.

Pulmonary emphysema is associated with dysregulated innate immune responses that promote chronic pulmonary inflammation and alveolar apoptosis, culminating in lung destruction. However, the molecular regulators of innate immunity that promote emphysema are ill-defined. Here, we investigated whether innate immune inflammasome complexes, comprising the adaptor ASC, Caspase-1 and specific pattern recognition receptors (PRRs), promote the pathogenesis of emphysema. In the lungs of emphysematous patients, as well as spontaneous gp130F/F and cigarette smoke (CS)-induced mouse models of emphysema, the expression (messenger RNA and protein) and activation of ASC, Caspase-1, and the inflammasome-associated PRR and DNA sensor AIM2 were up-regulated. AIM2 up-regulation in emphysema coincided with the biased production of the mature downstream inflammasome effector cytokine IL-1ß but not IL-18. These observations were supported by the genetic blockade of ASC, AIM2, and the IL-1 receptor and therapy with AIM2 antagonistic suppressor oligonucleotides, which ameliorated emphysema in gp130F/F mice by preventing elevated alveolar cell apoptosis. The functional requirement for AIM2 in driving apoptosis in the lung epithelium was independent of its expression in hematopoietic-derived immune cells and the recruitment of infiltrating immune cells in the lung. Genetic and inhibitor-based blockade of AIM2 also protected CS-exposed mice from pulmonary alveolar cell apoptosis. Intriguingly, IL-6 trans-signaling via the soluble IL-6 receptor, facilitated by elevated levels of IL-6, acted upstream of the AIM2 inflammasome to augment AIM2 expression in emphysema. Collectively, we reveal cross-talk between the AIM2 inflammasome/IL-1ß and IL-6 trans-signaling axes for potential exploitation as a therapeutic strategy for emphysema.

Cigarette Smoke Exposure Induces Neurocognitive Impairments and Neuropathological Changes in the Hippocampus.

Background and Objective: Neurocognitive dysfunction is present in up to ∼61% of people with chronic obstructive pulmonary disease (COPD), with symptoms including learning and memory deficiencies, negatively impacting the quality of life of these individuals. As the mechanisms responsible for neurocognitive deficits in COPD remain unknown, we explored whether chronic cigarette smoke (CS) exposure causes neurocognitive dysfunction in mice and whether this is associated with neuroinflammation and an altered neuropathology. Methods: Male BALB/c mice were exposed to room air (sham) or CS (9 cigarettes/day, 5 days/week) for 24 weeks. After 23 weeks, mice underwent neurocognitive tests to assess working and spatial memory retention. At 24 weeks, mice were culled and lungs were collected and assessed for hallmark features of COPD. Serum was assessed for systemic inflammation and the hippocampus was collected for neuroinflammatory and structural analysis. Results: Chronic CS exposure impaired lung function as well as driving pulmonary inflammation, emphysema, and systemic inflammation. CS exposure impaired working memory retention, which was associated with a suppression in hippocampal microglial number, however, these microglia displayed a more activated morphology. CS-exposed mice showed changes in astrocyte density as well as a reduction in synaptophysin and dendritic spines in the hippocampus. Conclusion: We have developed an experimental model of COPD in mice that recapitulates the hallmark features of the human disease. The altered microglial/astrocytic profiles and alterations in the neuropathology within the hippocampus may explain the neurocognitive dysfunction observed during COPD.

Targeting the human ßc receptor inhibits inflammatory myeloid cells and lung injury caused by acute cigarette smoke exposure.

BACKGROUND AND OBJECTIVE: Chronic obstructive pulmonary disease (COPD) is a devastating disease commonly caused by cigarette smoke (CS) exposure that drives tissue injury by persistently recruiting myeloid cells into the lungs. A significant portion of COPD patients also present with overlapping asthma pathology including eosinophilic inflammation. The ßc cytokine family includes granulocyte monocyte-colony-stimulating factor, IL-5 and IL-3 that signal through their common receptor subunit ßc to promote the expansion and survival of multiple myeloid cells including monocytes/macrophages, neutrophils and eosinophils. METHODS: We have used our unique human ßc receptor transgenic (hßc Tg) mouse strain that expresses human ßc instead of mouse ßc and ßIL3 in an acute CS exposure model. Lung tissue injury was assessed by histology and measurement of albumin and lactate dehydrogenase levels in the bronchoalveolar lavage (BAL) fluid. Transgenic mice were treated with an antibody (CSL311) that inhibits human ßc signalling. RESULTS: hßc Tg mice responded to acute CS exposure by expanding blood myeloid cell numbers and recruiting monocyte-derived macrophages (cluster of differentiation 11b+ [CD11b+ ] interstitial and exudative macrophages [IM and ExM]), neutrophils and eosinophils into the lungs. This inflammatory response was associated with lung tissue injury and oedema. Importantly, CSL311 treatment in CS-exposed mice markedly reduced myeloid cell numbers in the blood and BAL compartment. Furthermore, CSL311 significantly reduced lung CD11b+ IM and ExM, neutrophils and eosinophils, and this decline was associated with a significant reduction in matrix metalloproteinase-12 (MMP-12) and IL-17A expression, tissue injury and oedema. CONCLUSION: This study identifies CSL311 as a therapeutic antibody that potently inhibits immunopathology and lung injury caused by acute CS exposure.

Endothelial NOX4 Oxidase Negatively Regulates Inflammation and Improves Morbidity During Influenza A Virus Lung Infection in Mice.

Endosomal NOX2 oxidase-dependent ROS production promotes influenza pathogenicity, but the role of NOX4 oxidase, which is highly expressed in the lung endothelium, is largely unknown. The aim of this study was to determine if endothelial NOX4 expression can influence viral pathology in vivo, using a mouse model of influenza infection. WT and transgenic endothelial NOX4 overexpressing mice (NOX4 TG) were infected intranasally with the Hong Kong H3N2 X-31 influenza A virus (104 PFU; HK x-31) or PBS control. Mice were culled at either 3 or 7 days post-infection to analyse: airway inflammation by bronchoalveolar lavage fluid (BALF) cell counts; NOX4, as well as inflammatory cytokine and chemokine gene expression by QPCR; and ROS production by an L-012-enhanced chemiluminescence assay. Influenza A virus infection of WT mice resulted in a significant reduction in lung NOX4 mRNA at day 3, which persisted until day 7, when compared to uninfected mice. Influenza A virus infection of NOX4 TG mice resulted in significantly less weight loss than that of WT mice at 3-days post infection. Viral titres were decreased in infected NOX4 TG mice compared to the infected WT mice, at both 3- and 7-days post infection and there was significantly less lung alveolitis, peri-bronchial inflammation and neutrophil infiltration. The oxidative burst from BALF inflammatory cells extracted from infected NOX4 TG mice was significantly less than that in the WT mice. Expression of macrophage and neutrophil chemoattractants CXCL10, CCL3, CXCL1 and CXCL2 in the lung tissue were significantly lower in NOX4 TG mice compared to the WT mice at 3-days post infection. We conclude that endothelial NOX4 oxidase is protective against influenza morbidity and is a potential target for limiting influenza A virus-induced lung inflammation.

Therapeutic Targeting of Endosome and Mitochondrial Reactive Oxygen Species Protects Mice From Influenza Virus Morbidity.

There is an urgent need to develop effective therapeutic strategies including immunomodulators to combat influenza A virus (IAV) infection. Influenza A viruses increase ROS production, which suppress anti-viral responses and contribute to pathological inflammation and morbidity. Two major cellular sites of ROS production are endosomes via the NOX2-oxidase enzyme and the electron transport chain in mitochondria. Here we examined the effect of administration of Cgp91ds-TAT, an endosome-targeted NOX2 oxidase inhibitor, in combination with mitoTEMPO, a mitochondrial ROS scavenger and compared it to monotherapy treatment during an established IAV infection. Mice were infected with IAV (Hkx31 strain; 104PFU/mouse) and 24 h post infection were treated with Cgp91ds-TAT (0.2 mg/kg), mitoTEMPO (100 µg) or with a combination of these inhibitors [Cgp91ds-TAT (0.2 mg/kg)/mitoTEMPO (100 µg)] intranasally every day for up to 2 days post infection (pi). Mice were euthanized on Days 3 or 6 post infection for analyses of disease severity. A combination of Cgp91ds-TAT and mitoTEMPO treatment was more effective than the ROS inhibitors alone at reducing airway and neutrophilic inflammation, bodyweight loss, lung oedema and improved the lung pathology with a reduction in alveolitis following IAV infection. Dual ROS inhibition also caused a significant elevation in Type I IFN expression at the early phase of infection (day 3 pi), however, this response was suppressed at the later phase of infection (day 6 pi). Furthermore, combined treatment with Cgp91ds-TAT and mitoTEMPO resulted in an increase in IAV-specific CD8+ T cells in the lungs. In conclusion, this study demonstrates that the reduction of ROS production in two major subcellular sites, i.e. endosomes and mitochondria, by intranasal delivery of a combination of Cgp91ds-TAT and mitoTEMPO, suppresses the severity of influenza infection and highlights a novel immunomodulatory approach for IAV disease management.

Influenza A Virus-Driven Airway Inflammation may be Dissociated From Limb Muscle Atrophy in Cigarette Smoke-Exposed Mice.

Limb muscle dysfunction is a hallmark of Chronic Obstructive Pulmonary Disease (COPD) which is further worsened following a viral-induced acute exacerbation of COPD (AECOPD). An amplified airway inflammation underlies the aggravated respiratory symptoms seen during AECOPD, however, its contributory role to limb muscle dysfunction is unclear. The present study examined the impact of influenza A virus (IAV)-induced exacerbation on hind limb muscle parameters. Airway inflammation was established in male BALB/c mice by exposure to cigarette smoke (CS) for 8 weeks. Exacerbation was then induced via inoculation with IAV, and various lung and muscle parameters were assessed on day 3 (peak of airway inflammation) and day 10 (resolution phase) post-infection. IAV infection exacerbated CS-induced airway inflammation as evidenced by further increases in immune cell counts within bronchoalveolar lavage fluid. Despite no significant impact on muscle mass, IAV exacerbation worsened the force-generating capacity of the tibialis anterior (TA) muscle. Protein oxidation and myogenic disruption was observed in the TA following CS exposure, however, IAV exacerbation did not augment these detrimental processes. To further explore the contributory role of airway inflammation on myogenic signaling, cultured myotubes were exposed to conditioned medium (CM) derived from bronchial epithelial cells stimulated with polyinosinic:polycytidylic acid and cigarette smoke extract (CSE). Despite an amplified inflammatory response in the lung epithelial cells, the CM derived from these cells did not potentiate myogenic disruption in the C2C12 myotubes. In conclusion, our data suggest that certain parameters of limb muscle dysfunction seen during viral-induced AECOPD may be independent of airway inflammation.

Chronic obstructive pulmonary disease and atherosclerosis: common mechanisms and novel therapeutics.

Chronic obstructive pulmonary disease (COPD) and atherosclerosis are chronic irreversible diseases, that share a number of common causative factors including cigarette smoking. Atherosclerosis drastically impairs blood flow and oxygen availability to tissues, leading to life-threatening outcomes including myocardial infarction (MI) and stroke. Patients with COPD are most likely to die as a result of a cardiovascular event, with 30% of all COPD-related deaths being attributed to cardiovascular disease (CVD). Both atherosclerosis and COPD involve significant local (i.e. lung, vasculature) and systemic inflammation and oxidative stress, of which current pharmacological treatments have limited efficacy, hence the urgency for the development of novel life-saving therapeutics. Currently these diseases must be treated individually, with no therapies available that can effectively reduce the likelihood of comorbid CVD other than cessation of cigarette smoking. In this review, the important mechanisms that drive atherosclerosis and CVD in people with COPD are explained and we propose that modulation of both the oxidative stress and the inflammatory burden will provide a novel therapeutic strategy to treat both the pulmonary and systemic manifestations related to these diseases.

Ebselen prevents cigarette smoke-induced cognitive dysfunction in mice by preserving hippocampal synaptophysin expression.

BACKGROUND: Cigarette smoking (CS) is the leading cause of chronic obstructive pulmonary disease (COPD). The "spill-over" of pulmonary inflammation into the systemic circulation may damage the brain, leading to cognitive dysfunction. Cessation of CS can improve pulmonary and neurocognitive outcomes, however, its benefit on the neuroinflammatory profile remains uncertain. Here, we investigate how CS exposure impairs neurocognition and whether this can be reversed with CS cessation or an antioxidant treatment. METHODS: Male BALB/c mice were exposed to CS (9 cigarettes/day for 8 weeks) followed by 4 weeks of CS cessation. Another cohort of CS-exposed mice were co-administrated with a glutathione peroxidase mimetic, ebselen (10 mg/kg) or vehicle (5% CM-cellulose). We assessed pulmonary inflammation, spatial and working memory, and the hippocampal microglial, oxidative and synaptic profiles. RESULTS: CS exposure increased lung inflammation which was reduced following CS cessation. CS caused spatial and working memory impairments which were attributed to hippocampal microglial activation and suppression of synaptophysin. CS cessation did not improve memory deficits or alter microglial activation. Ebselen completely prevented the CS-induced working and spatial memory impairments, which was associated with restored synaptophysin expression without altering microglial activation. CONCLUSION: We were able to model the CS-induced memory impairment and microglial activation seen in human COPD. The preventative effects of ebselen on memory impairment is likely to be dependent on a preserved synaptogenic profile. Cessation alone also appears to be insufficient in correcting the memory impairment, suggesting the importance of incorporating antioxidant therapy to help maximising the benefit of cessation.