276 results on '"Darryl A. Knight"'
Search Results
2. Substrate stiffness engineered to replicate disease conditions influence senescence and fibrotic responses in primary lung fibroblasts
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Kaj E. C. Blokland, Mehmet Nizamoglu, Habibie Habibie, Theo Borghuis, Michael Schuliga, Barbro N. Melgert, Darryl A. Knight, Corry-Anke Brandsma, Simon D. Pouwels, and Janette K. Burgess
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cellular senescence ,stiffness ,GelMA hydrogels ,fibrosis ,SASP ,Therapeutics. Pharmacology ,RM1-950 - Abstract
In fibrosis remodelling of ECM leads to changes in composition and stiffness. Such changes can have a major impact on cell functions including proliferation, secretory profile and differentiation. Several studies have reported that fibrosis is characterised by increased senescence and accumulating evidence suggests that changes to the ECM including altered composition and increased stiffness may contribute to premature cellular senescence. This study investigated if increased stiffness could modulate markers of senescence and/or fibrosis in primary human lung fibroblasts. Using hydrogels representing stiffnesses that fall within healthy and fibrotic ranges, we cultured primary fibroblasts from non-diseased lung tissue on top of these hydrogels for up to 7 days before assessing senescence and fibrosis markers. Fibroblasts cultured on stiffer (±15 kPa) hydrogels showed higher Yes-associated protein-1 (YAP) nuclear translocation compared to soft hydrogels. When looking at senescence-associated proteins we also found higher secretion of receptor activator of nuclear factor kappa-B ligand (RANKL) but no change in transforming growth factor-β1 (TGF-β1) or connective tissue growth factor (CTGF) expression and higher decorin protein deposition on stiffer matrices. With respect to genes associated with fibrosis, fibroblasts on stiffer hydrogels compared to soft had higher expression of smooth muscle alpha (α)-2 actin (ACTA2), collagen (COL) 1A1 and fibulin-1 (Fbln1) and higher Fbln1 protein deposition after 7 days. Our results show that exposure of lung fibroblasts to fibrotic stiffness activates genes and secreted factors that are part of fibrotic responses and part of the Senescence-associated secretory phenotype (SASP). This overlap may contribute to the creation of a feedback loop whereby fibroblasts create a perpetuating cycle reinforcing progression of a fibrotic response.
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- 2022
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3. IL-4Rα blockade reduces influenza-associated morbidity in a murine model of allergic asthma
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Kimia Shahangian, David A. Ngan, H. H. Rachel Chen, Yeni Oh, Anthony Tam, Jing Wen, Chung Cheung, Darryl A. Knight, Delbert R. Dorscheid, Tillie L. Hackett, Michael R. Hughes, Kelly M. McNagny, Jeremy A. Hirota, Masahiro Niikura, S. F. Paul Man, and Don D. Sin
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Diseases of the respiratory system ,RC705-779 - Abstract
Abstract Background Asthma was identified as the most common comorbidity in hospitalized patients during the 2009 H1N1 influenza pandemic. We determined using a murine model of allergic asthma whether these mice experienced increased morbidity from pandemic H1N1 (pH1N1) viral infection and whether blockade of interleukin-4 receptor α (IL-4Rα), a critical mediator of Th2 signalling, improved their outcomes. Methods Male BALB/c mice were intranasally sensitized with house dust mite antigen (Der p 1) for 2 weeks; the mice were then inoculated intranasally with a single dose of pandemic H1N1 (pH1N1). The mice were administered intraperitoneally anti-IL-4Rα through either a prophylactic or a therapeutic treatment strategy. Results Infection with pH1N1 of mice sensitized to house dust mite (HDM) led to a 24% loss in weight by day 7 of infection (versus 14% in non-sensitized mice; p
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- 2021
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4. Inhibition of β-Catenin/CREB Binding Protein Signaling Attenuates House Dust Mite-Induced Goblet Cell Metaplasia in Mice
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Virinchi N. S. Kuchibhotla, Malcolm R. Starkey, Andrew T. Reid, Irene H. Heijink, Martijn C. Nawijn, Philip M. Hansbro, and Darryl A. Knight
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asthma ,airway inflammation ,β-catenin ,ICG-001 ,goblet cell metaplasia ,Physiology ,QP1-981 - Abstract
Excessive mucus production is a major feature of allergic asthma. Disruption of epithelial junctions by allergens such as house dust mite (HDM) results in the activation of β-catenin signaling, which has been reported to stimulate goblet cell differentiation. β-catenin interacts with various co-activators including CREB binding protein (CBP) and p300, thereby regulating the expression of genes involved in cell proliferation and differentiation, respectively. We specifically investigated the role of the β-catenin/CBP signaling pathway in goblet cell metaplasia in a HDM-induced allergic airway disease model in mice using ICG-001, a small molecule inhibitor that blocks the binding of CBP to β-catenin. Female 6- 8-week-old BALB/c mice were sensitized to HDM/saline on days 0, 1, and 2, followed by intranasal challenge with HDM/saline with or without subcutaneous ICG-001/vehicle treatment from days 14 to 17, and samples harvested 24 h after the last challenge/treatment. Differential inflammatory cells in bronchoalveolar lavage (BAL) fluid were enumerated. Alcian blue (AB)/Periodic acid–Schiff (PAS) staining was used to identify goblet cells/mucus production, and airway hyperresponsiveness (AHR) was assessed using invasive plethysmography. Exposure to HDM induced airway inflammation, goblet cell metaplasia and increased AHR, with increased airway resistance in response to the non-specific spasmogen methacholine. Inhibition of the β-catenin/CBP pathway using treatment with ICG-001 significantly attenuated the HDM-induced goblet cell metaplasia and infiltration of macrophages, but had no effect on eosinophils, neutrophils, lymphocytes or AHR. Increased β-catenin/CBP signaling may promote HDM-induced goblet cell metaplasia in mice.
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- 2021
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5. Airway mechanical compression: its role in asthma pathogenesis and progression
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Punnam Chander Veerati, Jennifer A. Mitchel, Andrew T. Reid, Darryl A. Knight, Nathan W. Bartlett, Jin-Ah Park, and Chris L. Grainge
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Diseases of the respiratory system ,RC705-779 - Abstract
The lung is a mechanically active organ, but uncontrolled or excessive mechanical forces disrupt normal lung function and can contribute to the development of disease. In asthma, bronchoconstriction leads to airway narrowing and airway wall buckling. A growing body of evidence suggests that pathological mechanical forces induced by airway buckling alone can perpetuate disease processes in asthma. Here, we review the data obtained from a variety of experimental models, including in vitro, ex vivo and in vivo approaches, which have been used to study the impact of mechanical forces in asthma pathogenesis. We review the evidence showing that mechanical compression alters the biological and biophysical properties of the airway epithelium, including activation of the epidermal growth factor receptor pathway, overproduction of asthma-associated mediators, goblet cell hyperplasia, and a phase transition of epithelium from a static jammed phase to a mobile unjammed phase. We also define questions regarding the impact of mechanical forces on the pathology of asthma, with a focus on known triggers of asthma exacerbations such as viral infection.
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- 2020
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6. Airway Epithelial Cell Immunity Is Delayed During Rhinovirus Infection in Asthma and COPD
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Punnam Chander Veerati, Niamh M. Troy, Andrew T. Reid, Ngan Fung Li, Kristy S. Nichol, Parwinder Kaur, Steven Maltby, Peter A. B. Wark, Darryl A. Knight, Anthony Bosco, Chris L. Grainge, and Nathan W. Bartlett
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rhinovirus ,interferon response ,innate immunity ,asthma ,chronic obstructive pulmonary disease (COPD) ,air-liquid interface (ALI) culture ,Immunologic diseases. Allergy ,RC581-607 - Abstract
Respiratory viral infections, particularly those caused by rhinovirus, exacerbate chronic respiratory inflammatory diseases, such as asthma and chronic obstructive pulmonary disease (COPD). Airway epithelial cells are the primary site of rhinovirus replication and responsible of initiating the host immune response to infection. Numerous studies have reported that the anti-viral innate immune response (including type I and type III interferon) in asthma is less effective or deficient leading to the conclusion that epithelial innate immunity is a key determinant of disease severity during a rhinovirus induced exacerbation. However, deficient rhinovirus-induced epithelial interferon production in asthma has not always been observed. We hypothesized that disparate in vitro airway epithelial infection models using high multiplicity of infection (MOI) and lacking genome-wide, time course analyses have obscured the role of epithelial innate anti-viral immunity in asthma and COPD. To address this, we developed a low MOI rhinovirus model of differentiated primary epithelial cells obtained from healthy, asthma and COPD donors. Using genome-wide gene expression following infection, we demonstrated that gene expression patterns are similar across patient groups, but that the kinetics of induction are delayed in cells obtained from asthma and COPD donors. Rhinovirus-induced innate immune responses were defined by interferons (type-I, II, and III), interferon response factors (IRF1, IRF3, and IRF7), TLR signaling and NF-κB and STAT1 activation. Induced gene expression was evident at 24 h and peaked at 48 h post-infection in cells from healthy subjects. In contrast, in cells from donors with asthma or COPD induction was maximal at or beyond 72–96 h post-infection. Thus, we propose that propensity for viral exacerbations of asthma and COPD relate to delayed (rather than deficient) expression of epithelial cell innate anti-viral immune genes which in turns leads to a delayed and ultimately more inflammatory host immune response.
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- 2020
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7. Influenza A virus infection dysregulates the expression of microRNA-22 and its targets; CD147 and HDAC4, in epithelium of asthmatics
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Fatemeh Moheimani, Jorinke Koops, Teresa Williams, Andrew T. Reid, Philip M. Hansbro, Peter A. Wark, and Darryl A. Knight
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Severe asthma ,Epithelial cells ,microRNA ,Influenza A virus ,Airway remodeling ,Diseases of the respiratory system ,RC705-779 - Abstract
Abstract Background Specific microRNAs (miRNAs) play essential roles in airway remodeling in asthma. Infection with influenza A virus (IAV) may also magnify pre-existing airway remodeling leading to asthma exacerbation. However, these events remain to be fully defined. We investigated the expression of miRNAs with diverse functions including proliferation (miR-20a), differentiation (miR-22) or innate/adaptive immune responses (miR-132) in primary bronchial epithelial cells (pBECs) of asthmatics following infection with the H1N1 strain of IAV. Methods pBECs from subjects (n = 5) with severe asthma and non-asthmatics were cultured as submerged monolayers or at the air-liquid-interface (ALI) conditions and incubated with IAV H1N1 (MOI 5) for up to 24 h. Isolated miRNAs were subjected to Taqman miRNAs assays. We confirmed miRNA targets using a specific mimic and antagomir. Taqman mRNAs assays and immunoblotting were used to assess expression of target genes and proteins, respectively. Results At baseline, these miRNAs were expressed at the same level in pBECs of asthmatics and non-asthmatics. After 24 h of infection, miR-22 expression increased significantly which was associated with the suppression of CD147 mRNA and HDAC4 mRNA and protein expression in pBECs from non-asthmatics, cultured in ALI. In contrast, miR-22 remained unchanged while CD147 expression increased and HDAC4 remained unaffected in cells from asthmatics. IAV H1N1 mediated increases in SP1 and c-Myc transcription factors may underpin the induction of CD147 in asthmatics. Conclusion The different profile of miR-22 expression in differentiated epithelial cells from non-asthmatics may indicate a self-defense mechanism against aberrant epithelial responses through suppressing CD147 and HDAC4, which is compromised in epithelial cells of asthmatics.
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- 2018
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8. Corticosteroid suppression of antiviral immunity increases bacterial loads and mucus production in COPD exacerbations
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Aran Singanayagam, Nicholas Glanville, Jason L. Girkin, Yee Man Ching, Andrea Marcellini, James D. Porter, Marie Toussaint, Ross P. Walton, Lydia J. Finney, Julia Aniscenko, Jie Zhu, Maria-Belen Trujillo-Torralbo, Maria Adelaide Calderazzo, Chris Grainge, Su-Ling Loo, Punnam Chander Veerati, Prabuddha S. Pathinayake, Kristy S. Nichol, Andrew T. Reid, Phillip L. James, Roberto Solari, Peter A. B. Wark, Darryl A. Knight, Miriam F. Moffatt, William O. Cookson, Michael R. Edwards, Patrick Mallia, Nathan W. Bartlett, and Sebastian L. Johnston
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Science - Abstract
Corticosteroid therapy is frequently used for chronic obstructive pulmonary disease (COPD) but its use is associated with increased risk of pneumonia. Here the authors show that corticosteroid use impairs innate and adaptive immunity to rhinovirus infection, which is restored by exogenous IFNβ.
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- 2018
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9. Accumulation mode particles and LPS exposure induce TLR-4 dependent and independent inflammatory responses in the lung
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Angela M. Fonceca, Graeme R. Zosky, Elizabeth M. Bozanich, Erika N. Sutanto, Anthony Kicic, Paul S. McNamara, Darryl A. Knight, Peter D. Sly, Debra J. Turner, and Stephen M. Stick
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Asthma ,TLR-4 ,PM ,LPS ,AMP ,COPD ,Diseases of the respiratory system ,RC705-779 - Abstract
Abstract Background Accumulation mode particles (AMP) are formed from engine combustion and make up the inhalable vapour cloud of ambient particulate matter pollution. Their small size facilitates dispersal and subsequent exposure far from their original source, as well as the ability to penetrate alveolar spaces and capillary walls of the lung when inhaled. A significant immuno-stimulatory component of AMP is lipopolysaccharide (LPS), a product of Gram negative bacteria breakdown. As LPS is implicated in the onset and exacerbation of asthma, the presence or absence of LPS in ambient particulate matter (PM) may explain the onset of asthmatic exacerbations to PM exposure. This study aimed to delineate the effects of LPS and AMP on airway inflammation, and potential contribution to airways disease by measuring airway inflammatory responses induced via activation of the LPS cellular receptor, Toll-like receptor 4 (TLR-4). Methods The effects of nebulized AMP, LPS and AMP administered with LPS on lung function, cellular inflammatory infiltrate and cytokine responses were compared between wildtype mice and mice not expressing TLR-4. Results The presence of LPS administered with AMP appeared to drive elevated airway resistance and sensitivity via TLR-4. Augmented TLR4 driven eosinophilia and greater TNF-α responses observed in AMP-LPS treated mice independent of TLR-4 expression, suggests activation of allergic responses by TLR4 and non-TLR4 pathways larger than those induced by LPS administered alone. Treatment with AMP induced macrophage recruitment independent of TLR-4 expression. Conclusions These findings suggest AMP-LPS as a stronger stimulus for allergic inflammation in the airways then LPS alone.
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- 2018
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10. Dysregulated Notch Signaling in the Airway Epithelium of Children with Wheeze
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Thomas Iosifidis, Erika N. Sutanto, Samuel T. Montgomery, Patricia Agudelo-Romero, Kevin Looi, Kak-Ming Ling, Nicole C. Shaw, Luke W. Garratt, Jessica Hillas, Kelly M. Martinovich, Elizabeth Kicic-Starcevich, Shyan Vijayasekaran, Francis J. Lannigan, Paul J. Rigby, Darryl A. Knight, Stephen M. Stick, and Anthony Kicic
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pediatrics ,wheeze ,airway epithelium ,wound repair ,Notch ,Medicine - Abstract
The airway epithelium of children with wheeze is characterized by defective repair that contributes to disease pathobiology. Dysregulation of developmental processes controlled by Notch has been identified in chronic asthma. However, its role in airway epithelial cells of young children with wheeze, particularly during repair, is yet to be determined. We hypothesized that Notch is dysregulated in primary airway epithelial cells (pAEC) of children with wheeze contributing to defective repair. This study investigated transcriptional and protein expression and function of Notch in pAEC isolated from children with and without wheeze. Primary AEC of children with and without wheeze were found to express all known Notch receptors and ligands, although pAEC from children with wheeze expressed significantly lower NOTCH2 (10-fold, p = 0.004) and higher JAG1 (3.5-fold, p = 0.002) mRNA levels. These dysregulations were maintained in vitro and cultures from children with wheeze displayed altered kinetics of both NOTCH2 and JAG1 expression during repair. Following Notch signaling inhibition, pAEC from children without wheeze failed to repair (wound closure rate of 76.9 ± 3.2%). Overexpression of NOTCH2 in pAEC from children with wheeze failed to rescue epithelial repair following wounding. This study illustrates the involvement of the Notch pathway in airway epithelial wound repair in health and disease, where its dysregulation may contribute to asthma development.
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- 2021
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11. A Senescence Bystander Effect in Human Lung Fibroblasts
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David W. Waters, Michael Schuliga, Prabuddha S. Pathinayake, Lan Wei, Hui-Ying Tan, Kaj E. C. Blokland, Jade Jaffar, Glen P. Westall, Janette K. Burgess, Cecilia M. Prêle, Steven E. Mutsaers, Christopher L. Grainge, and Darryl A. Knight
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collagen ,idiopathic pulmonary fibrosis (IPF) ,lung fibroblasts ,senescence ,Biology (General) ,QH301-705.5 - Abstract
Idiopathic pulmonary fibrosis (IPF) is a chronic disease characterised by a dense fibrosing of the lung parenchyma. An association between IPF and cellular senescence is well established and several studies now describe a higher abundance of senescent fibroblasts and epithelial cells in the lungs of IPF patients compared with age-matched controls. The cause of this abnormal accumulation of senescent cells is unknown but evidence suggests that, once established, senescence can be transferred from senescent to non-senescent cells. In this study, we investigated whether senescent human lung fibroblasts (LFs) and alveolar epithelial cells (AECs) could induce a senescent-like phenotype in “naïve” non-senescent LFs in vitro. Primary cultures of LFs from adult control donors (Ctrl-LFs) with a low baseline of senescence were exposed to conditioned medium (CM) from: (i) Ctrl-LFs induced to become senescent using H2O2 or etoposide; (ii) LFs derived from IPF patients (IPF-LFs) with a high baseline of senescence; or (iii) senescence-induced A549 cells, an AEC line. Additionally, ratios of non-senescent Ctrl-LFs and senescence-induced Ctrl-LFs (100:0, 0:100, 50:50, 90:10, 99:1) were co-cultured and their effect on induction of senescence measured. We demonstrated that exposure of naïve non-senescent Ctrl-LFs to CM from senescence-induced Ctrl-LFs and AECs and IPF-LFs increased the markers of senescence including nuclear localisation of phosphorylated-H2A histone family member X (H2AXγ) and expression of p21, IL-6 and IL-8 in Ctrl-LFs. Additionally, co-cultures of non-senescent and senescence-induced Ctrl-LFs induced a senescent-like phenotype in the non-senescent cells. These data suggest that the phenomenon of “senescence-induced senescence” can occur in vitro in primary cultures of human LFs, and provides a possible explanation for the abnormal abundance of senescent cells in the lungs of IPF patients.
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- 2021
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12. Previous Influenza Infection Exacerbates Allergen Specific Response and Impairs Airway Barrier Integrity in Pre-Sensitized Mice
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Kevin Looi, Alexander N. Larcombe, Kara L. Perks, Luke J. Berry, Graeme R. Zosky, Paul Rigby, Darryl A. Knight, Anthony Kicic, and Stephen M. Stick
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house dust mite ,lung function ,BALB/c mice ,influenza ,tight junctions ,epithelial barrier integrity ,Biology (General) ,QH301-705.5 ,Chemistry ,QD1-999 - Abstract
In this study we assessed the effects of antigen exposure in mice pre-sensitized with allergen following viral infection on changes in lung function, cellular responses and tight junction expression. Female BALB/c mice were sensitized to ovalbumin and infected with influenza A before receiving a second ovalbumin sensitization and challenge with saline, ovalbumin (OVA) or house dust mite (HDM). Fifteen days post-infection, bronchoalveolar inflammation, serum antibodies, responsiveness to methacholine and barrier integrity were assessed. There was no effect of infection alone on bronchoalveolar lavage cellular inflammation 15 days post-infection; however, OVA or HDM challenge resulted in increased bronchoalveolar inflammation dominated by eosinophils/neutrophils or neutrophils, respectively. Previously infected mice had higher serum OVA-specific IgE compared with uninfected mice. Mice previously infected, sensitized and challenged with OVA were most responsive to methacholine with respect to airway resistance, while HDM challenge caused significant increases in both tissue damping and tissue elastance regardless of previous infection status. Previous influenza infection was associated with decreased claudin-1 expression in all groups and decreased occludin expression in OVA or HDM-challenged mice. This study demonstrates the importance of the respiratory epithelium in pre-sensitized individuals, where influenza-infection-induced barrier disruption resulted in increased systemic OVA sensitization and downstream effects on lung function.
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- 2021
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13. Regulation of Cellular Senescence Is Independent from Profibrotic Fibroblast-Deposited ECM
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Kaj E. C. Blokland, Habibie Habibie, Theo Borghuis, Greta J. Teitsma, Michael Schuliga, Barbro N. Melgert, Darryl A. Knight, Corry-Anke Brandsma, Simon D. Pouwels, and Janette K. Burgess
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extracellular matrix ,senescence ,idiopathic pulmonary fibrosis ,proinflammatory ,profibrotic ,Cytology ,QH573-671 - Abstract
Idiopathic pulmonary fibrosis (IPF) is a devastating lung disease with poor survival. Age is a major risk factor, and both alveolar epithelial cells and lung fibroblasts in this disease exhibit features of cellular senescence, a hallmark of ageing. Accumulation of fibrotic extracellular matrix (ECM) is a core feature of IPF and is likely to affect cell function. We hypothesize that aberrant ECM deposition augments fibroblast senescence, creating a perpetuating cycle favouring disease progression. In this study, primary lung fibroblasts were cultured on control and IPF-derived ECM from fibroblasts pretreated with or without profibrotic and prosenescent stimuli, and markers of senescence, fibrosis-associated gene expression and secretion of cytokines were measured. Untreated ECM derived from control or IPF fibroblasts had no effect on the main marker of senescence p16Ink4a and p21Waf1/Cip1. However, the expression of alpha smooth muscle actin (ACTA2) and proteoglycan decorin (DCN) increased in response to IPF-derived ECM. Production of the proinflammatory cytokines C-X-C Motif Chemokine Ligand 8 (CXCL8) by lung fibroblasts was upregulated in response to senescent and profibrotic-derived ECM. Finally, the profibrotic cytokines transforming growth factor β1 (TGF-β1) and connective tissue growth factor (CTGF) were upregulated in response to both senescent- and profibrotic-derived ECM. In summary, ECM deposited by IPF fibroblasts does not induce cellular senescence, while there is upregulation of proinflammatory and profibrotic cytokines and differentiation into a myofibroblast phenotype in response to senescent- and profibrotic-derived ECM, which may contribute to progression of fibrosis in IPF.
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- 2021
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14. Reduced SOCS1 Expression in Lung Fibroblasts from Patients with IPF Is Not Mediated by Promoter Methylation or Mir155
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Cecilia M. Prêle, Thomas Iosifidis, Robin J. McAnulty, David R. Pearce, Bahareh Badrian, Tylah Miles, Sarra E. Jamieson, Matthias Ernst, Philip J. Thompson, Geoffrey J. Laurent, Darryl A. Knight, and Steven E. Mutsaers
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L-6 ,Jak/STAT pathway ,SOCS1 ,miR155 ,fibroblast ,fibrosis ,Biology (General) ,QH301-705.5 - Abstract
The interleukin (IL)-6 family of cytokines and exaggerated signal transducer and activator of transcription (STAT)3 signaling is implicated in idiopathic pulmonary fibrosis (IPF) pathogenesis, but the mechanisms regulating STAT3 expression and function are unknown. Suppressor of cytokine signaling (SOCS)1 and SOCS3 block STAT3, and low SOCS1 levels have been reported in IPF fibroblasts and shown to facilitate collagen production. Fibroblasts and lung tissue from IPF patients and controls were used to examine the mechanisms underlying SOCS1 down-regulation in IPF. A significant reduction in basal SOCS1 mRNA in IPF fibroblasts was confirmed. However, there was no difference in the kinetics of activation, and methylation of SOCS1 in control and IPF lung fibroblasts was low and unaffected by 5′-aza-2′-deoxycytidine’ treatment. SOCS1 is a target of microRNA-155 and although microRNA-155 levels were increased in IPF tissue, they were reduced in IPF fibroblasts. Therefore, SOCS1 is not regulated by SOCS1 gene methylation or microRNA155 in these cells. In conclusion, we confirmed that IPF fibroblasts had lower levels of SOCS1 mRNA compared with control fibroblasts, but we were unable to determine the mechanism. Furthermore, although SOCS1 may be important in the fibrotic process, we were unable to find a significant role for SOCS1 in regulating fibroblast function.
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- 2021
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15. Acute cigarette smoke exposure activates apoptotic and inflammatory programs but a second stimulus is required to induce epithelial to mesenchymal transition in COPD epithelium
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Lynne A. Murray, Rebecca Dunmore, Ana Camelo, Carla A. Da Silva, Malin J. Gustavsson, David M. Habiel, Tillie L Hackett, Cory M. Hogaboam, Matthew A. Sleeman, and Darryl A. Knight
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TGFβ1 ,Poly I:C ,Remodelling ,Apoptosis ,Diseases of the respiratory system ,RC705-779 - Abstract
Abstract Background Smoking and aberrant epithelial responses are risk factors for lung cancer as well as chronic obstructive pulmonary disease and idiopathic pulmonary fibrosis. In these conditions, disease progression is associated with epithelial damage and fragility, airway remodelling and sub-epithelial fibrosis. The aim of this study was to assess the acute effects of cigarette smoke on epithelial cell phenotype and pro-fibrotic responses in vitro and in vivo. Results Apoptosis was significantly greater in unstimulated cells from COPD patients compared to control, but proliferation and CXCL8 release were not different. Cigarette smoke dose-dependently induced apoptosis, proliferation and CXCL8 release with normal epithelial cells being more responsive than COPD patient derived cells. Cigarette smoke did not induce epithelial-mesenchymal transition. In vivo, cigarette smoke exposure promoted epithelial apoptosis and proliferation. Moreover, mimicking a virus-induced exacerbation by exposing to mice to poly I:C, exaggerated the inflammatory responses, whereas expression of remodelling genes was similar in both. Conclusions Collectively, these data indicate that cigarette smoke promotes epithelial cell activation and hyperplasia, but a secondary stimulus is required for the remodelling phenotype associated with COPD.
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- 2017
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16. Senescence of IPF Lung Fibroblasts Disrupt Alveolar Epithelial Cell Proliferation and Promote Migration in Wound Healing
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Kaj E. C. Blokland, David W. Waters, Michael Schuliga, Jane Read, Simon D. Pouwels, Christopher L. Grainge, Jade Jaffar, Glen Westall, Steven E. Mutsaers, Cecilia M. Prêle, Janette K. Burgess, and Darryl A. Knight
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senescence ,fibroblasts ,alveolar epithelial cell ,fibrosis ,aberrant repair ,cell-cycle inhibition ,Pharmacy and materia medica ,RS1-441 - Abstract
Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease marked by excessive accumulation of lung fibroblasts (LFs) and collagen in the lung parenchyma. The mechanisms that underlie IPF pathophysiology are thought to reflect repeated alveolar epithelial injury leading to an aberrant wound repair response. Recent work has shown that IPF-LFs display increased characteristics of senescence including growth arrest and a senescence-associated secretory phenotype (SASP) suggesting that senescent LFs contribute to dysfunctional wound repair process. Here, we investigated the influence of senescent LFs on alveolar epithelial cell repair responses in a co-culture system. Alveolar epithelial cell proliferation was attenuated when in co-culture with cells or conditioned media from, senescence-induced control LFs or IPF-LFs. Cell-cycle analyses showed that a larger number of epithelial cells were arrested in G2/M phase when co-cultured with IPF-LFs, than in monoculture. Paradoxically, the presence of LFs resulted in increased A549 migration after mechanical injury. Our data suggest that senescent LFs may contribute to aberrant re-epithelialization by inhibiting proliferation in IPF.
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- 2020
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17. The Processes and Mechanisms of Cardiac and Pulmonary Fibrosis
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Lucy A. Murtha, Michael J. Schuliga, Nishani S. Mabotuwana, Sean A. Hardy, David W. Waters, Janette K. Burgess, Darryl A. Knight, and Andrew J. Boyle
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cardiac fibrosis ,pulmonary fibrosis ,heart failure ,myocardial infarction ,idiopathic pulmonary hypertension ,acute respiratory distress syndrome ,Physiology ,QP1-981 - Abstract
Fibrosis is the formation of fibrous connective tissue in response to injury. It is characterized by the accumulation of extracellular matrix components, particularly collagen, at the site of injury. Fibrosis is an adaptive response that is a vital component of wound healing and tissue repair. However, its continued activation is highly detrimental and a common final pathway of numerous disease states including cardiovascular and respiratory disease. Worldwide, fibrotic diseases cause over 800,000 deaths per year, accounting for ~45% of total deaths. With an aging population, the incidence of fibrotic disease and subsequently the number of fibrosis-related deaths will rise further. Although, fibrosis is a well-recognized cause of morbidity and mortality in a range of disease states, there are currently no viable therapies to reverse the effects of chronic fibrosis. Numerous predisposing factors contribute to the development of fibrosis. Biological aging in particular, interferes with repair of damaged tissue, accelerating the transition to pathological remodeling, rather than a process of resolution and regeneration. When fibrosis progresses in an uncontrolled manner, it results in the irreversible stiffening of the affected tissue, which can lead to organ malfunction and death. Further investigation into the mechanisms of fibrosis is necessary to elucidate novel, much needed, therapeutic targets. Fibrosis of the heart and lung make up a significant proportion of fibrosis-related deaths. It has long been established that the heart and lung are functionally and geographically linked when it comes to health and disease, and thus exploring the processes and mechanisms that contribute to fibrosis of each organ, the focus of this review, may help to highlight potential avenues of therapeutic investigation.
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- 2017
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18. Correction: Granzyme B Cleaves Decorin, Biglycan and Soluble Betaglycan, Releasing Active Transforming Growth Factor-β1.
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Wendy A. Boivin, Marlo Shackleford, Amanda Vanden Hoek, Hongyan Zhao, Tillie L. Hackett, Darryl A. Knight, and David J. Granville
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Medicine ,Science - Published
- 2012
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19. The role of pathological aging in cardiac and pulmonary fibrosis
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Aaron L. Sverdlov, Michael Schuliga, Doan T.M. Ngo, Lucy A. Murtha, Andrew J. Boyle, Matthew Morten, David W Waters, N. Mabotuwana, Janette K. Burgess, Darryl A. Knight, and Sean A. Hardy
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0301 basic medicine ,Senescence ,autophagy ,senescence ,MITOCHONDRIAL DYSFUNCTION ,Cardiac fibrosis ,Disease ,heart ,Bioinformatics ,Pathology and Forensic Medicine ,lung ,MECHANISMS ,03 medical and health sciences ,Idiopathic pulmonary fibrosis ,0302 clinical medicine ,INFLAMMATION ,Fibrosis ,Pulmonary fibrosis ,medicine ,PLASMINOGEN-ACTIVATOR ,Lung ,pulmonary fibrosis ,business.industry ,aging ,CELLULAR SENESCENCE ,Cell Biology ,DIASTOLIC DYSFUNCTION ,MOUSE MODEL ,DNA ,medicine.disease ,030104 developmental biology ,medicine.anatomical_structure ,Heart failure ,HEART-FAILURE ,Original Article ,inflammaging ,Neurology (clinical) ,Geriatrics and Gerontology ,business ,030217 neurology & neurosurgery - Abstract
Aging promotes a range of degenerative pathologies characterized by progressive losses of tissue and/or cellular function. Fibrosis is the hardening, overgrowth and scarring of various tissues characterized by the accumulation of extracellular matrix components. Aging is an important predisposing factor common for fibrotic heart and respiratory disease. Age-related processes such as senescence, inflammaging, autophagy and mitochondrial dysfunction are interconnected biological processes that diminish the regenerative capacity of the aged heart and lung and have been shown to play a crucial role in cardiac fibrosis and idiopathic pulmonary fibrosis. This review focuses on these four processes of aging in relation to their role in fibrosis. It has long been established that the heart and lung are linked both functionally and anatomically when it comes to health and disease, with an ever-expanding aging population, the incidence of fibrotic disease and therefore the number of fibrosis-related deaths will continue to rise. There are currently no feasible therapies to treat the effects of chronic fibrosis therefore highlighting the importance of exploring the processes of aging and its role in inducing and exacerbating fibrosis of each organ. The focus of this review may help to highlight potential avenues of therapeutic exploration.
- Published
- 2023
20. Conditionally reprogrammed asthmatic bronchial epithelial cells express lower FOXJ1 at terminal differentiation and lower IFNs following RV-A1 infection
- Author
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Punnam Chander Veerati, Kristy S. Nichol, Jane M. Read, Nathan W. Bartlett, Peter A. B. Wark, Darryl A. Knight, Christopher L. Grainge, and Andrew T. Reid
- Subjects
Pulmonary and Respiratory Medicine ,Physiology ,Physiology (medical) ,Cell Biology - Abstract
Primary bronchial epithelial cells (pBECs) obtained from donors have limited proliferation capacity. Recently, conditional reprogramming (CR) technique has overcome this and has provided the potential for extended passaging and subsequent differentiation of cells at air-liquid interface (ALI). However, there has been no donor-specific comparison of cell morphology, baseline gene expression, barrier function, and antiviral responses compared with their “parent” pBECs, especially cells obtained from donors with asthma. We, therefore, collected and differentiated pBECs at ALI from mild donors with asthma ( n = 6) for the parent group. The same cells were conditionally reprogrammed and later differentiated at ALI. Barrier function was measured during the differentiation phase. Morphology and baseline gene expression were compared at terminal differentiation. Viral replication kinetics and antiviral responses were assessed following rhinovirus (RV) infection over 96 h. Barrier function during the differentiation phase and cell structural morphology at terminal differentiation appear similar in both parent and CR groups, however, there were elongated cell structures superficial to basal cells and significantly lower FOXJ1 expression in CR group. IFN gene expression was also significantly lower in CR group compared with parent asthma group following RV infection. The CR technique is a beneficial tool to proliferate pBECs over extended passages. Considering lower FOXJ1 expression, viral replication kinetics and antiviral responses, a cautious approach should be taken while choosing CR cells for experiments. In addition, as lab-to-lab cell culture techniques vary, the most appropriate technique must be utilized to best match individual cell functions and morphologies to address specific research questions and experimental reproducibility across the labs.
- Published
- 2022
21. Airway and parenchymal transcriptomics in a novel model of asthma and COPD overlap
- Author
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Xiaofan Tu, Richard Y. Kim, Alexandra C. Brown, Emma de Jong, Bernadette Jones-Freeman, Md Khadem Ali, Henry M. Gomez, Kurtis F. Budden, Malcolm R. Starkey, Guy J.M. Cameron, Svenja Loering, Duc H. Nguyen, Prema Mono Nair, Tatt Jhong Haw, Charlotte A. Alemao, Alen Faiz, Hock L. Tay, Peter A.B. Wark, Darryl A. Knight, Paul S. Foster, Anthony Bosco, Jay C. Horvat, Philip M. Hansbro, and Chantal Donovan
- Subjects
Allergy ,Immunology ,Asthma ,Mice ,Pulmonary Disease, Chronic Obstructive ,1107 Immunology ,Eosinophilia ,Respiratory Hypersensitivity ,Immunology and Allergy ,Animals ,RNA ,Female ,Transcriptome ,Transcription Factors - Abstract
BACKGROUND: Asthma and chronic obstructive pulmonary disease (COPD) are common chronic respiratory diseases, and some patients have overlapping disease features, termed asthma-COPD overlap (ACO). Patients characterized with ACO have increased disease severity; however, the mechanisms driving this have not been widely studied. OBJECTIVES: This study sought to characterize the phenotypic and transcriptomic features of experimental ACO in mice induced by chronic house dust mite antigen and cigarette smoke exposure. METHODS: Female BALB/c mice were chronically exposed to house dust mite antigen for 11 weeks to induce experimental asthma, cigarette smoke for 8 weeks to induce experimental COPD, or both concurrently to induce experimental ACO. Lung inflammation, structural changes, and lung function were assessed. RNA-sequencing was performed on separated airway and parenchyma lung tissues to assess transcriptional changes. Validation of a novel upstream driver SPI1 in experimental ACO was assessed using the pharmacological SPI1 inhibitor, DB2313. RESULTS: Experimental ACO recapitulated features of both asthma and COPD, with mixed pulmonary eosinophilic/neutrophilic inflammation, small airway collagen deposition, and increased airway hyperresponsiveness. Transcriptomic analysis identified common and distinct dysregulated gene clusters in airway and parenchyma samples in experimental asthma, COPD, and ACO. Upstream driver analysis revealed increased expression of the transcription factor Spi1. Pharmacological inhibition of SPI1 using DB2313, reduced airway remodeling and airway hyperresponsiveness in experimental ACO. CONCLUSIONS: A new experimental model of ACO featuring chronic dual exposures to house dust mite and cigarette smoke mimics key disease features observed in patients with ACO and revealed novel disease mechanisms, including upregulation of SPI1, that are amenable to therapy.
- Published
- 2022
22. Epithelial Mesenchymal Transition in Respiratory Disease
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Stephen M. Stick, Anthony Kicic, Michael Schuliga, Darryl A. Knight, and Christopher Grainge
- Subjects
Pulmonary and Respiratory Medicine ,Pathology ,medicine.medical_specialty ,COPD ,biology ,business.industry ,Respiratory disease ,Critical Care and Intensive Care Medicine ,medicine.disease ,Cystic fibrosis ,Cystic fibrosis transmembrane conductance regulator ,Extracellular matrix ,Idiopathic pulmonary fibrosis ,Fibrosis ,medicine ,biology.protein ,Epithelial–mesenchymal transition ,Cardiology and Cardiovascular Medicine ,business - Published
- 2020
23. Airway epithelial-targeted nanoparticles for asthma therapy
- Author
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Darryl A. Knight, Christopher Grainge, Dewi Melani Hariyadi, Stanislav Kan, Mingtao Liang, and Nathan W. Bartlett
- Subjects
0301 basic medicine ,Pulmonary and Respiratory Medicine ,Physiology ,Disease ,Epithelium ,03 medical and health sciences ,Drug Delivery Systems ,0302 clinical medicine ,Immune system ,Physiology (medical) ,medicine ,Animals ,Humans ,Anti-Asthmatic Agents ,Asthma ,Drug Carriers ,business.industry ,Cell Biology ,respiratory system ,medicine.disease ,Mucus ,respiratory tract diseases ,030104 developmental biology ,030228 respiratory system ,Bronchial hyperresponsiveness ,Immunology ,Drug delivery ,Airway Remodeling ,Nanoparticles ,Respiratory epithelium ,business ,Airway - Abstract
Asthma is a common chronic inflammatory disease associated with intermittent airflow obstruction caused by airway inflammation, mucus overproduction, and bronchial hyperresponsiveness. Despite current treatment and management options, a large number of patients with asthma still have poorly controlled disease and are susceptible to acute exacerbations, usually caused by a respiratory virus infection. As a result, there remains a need for novel therapies to achieve better control and prevent/treat exacerbations. Nanoparticles (NPs), including extracellular vesicles (EV) and their synthetic counterparts, have been developed for drug delivery in respiratory diseases. In the case of asthma, where airway epithelium dysfunction, including dysregulated differentiation of epithelial cells, impaired barrier, and immune response, is a driver of disease, targeting airway epithelial cells with NPs may offer opportunities to repair or reverse these dysfunctions with therapeutic interventions. EVs possess multiple advantages for airway epithelial targeting, such as their natural intrinsic cell-targeting properties and low immunogenicity. Synthetic NPs can be coated with muco-inert polymers to overcome biological barriers such as mucus and the phagocytic response of immune cells. Targeting ligands could be also added to enhance targeting specificity to epithelial cells. The review presents current understanding and advances in NP-mediated drug delivery to airway epithelium for asthma therapy. Future perspectives in this therapeutic strategy will also be discussed, including the development of novel formulations and physiologically relevant preclinical models.
- Published
- 2020
24. Inhibition of β‐catenin/CBP signalling improves airway epithelial barrier function and suppresses CCL20 release
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Darryl A. Knight, Jacobien A. Noordhoek, Marnix R. Jonker, Harold G. de Bruin, Virinchi N. S. Kuchibhotla, Irene H. Heijink, Martijn C. Nawijn, and Groningen Research Institute for Asthma and COPD (GRIAC)
- Subjects
Chemokine CCL20 ,Chemistry ,Immunology ,Inflammation ,Epithelium ,Cell biology ,CCL20 ,medicine.anatomical_structure ,Signalling ,Catenin ,E-CADHERIN ,medicine ,Humans ,Immunology and Allergy ,Epithelial barrier function ,medicine.symptom ,Letters to the Editor ,Airway ,Letter to the Editor ,beta Catenin ,CELL-ADHESION ,Signal Transduction - Published
- 2020
25. A cGAS-dependent response links DNA damage and senescence in alveolar epithelial cells: A potential drug target in IPF
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Cecilia M. Prêle, Claire Thomson, Darryl A. Knight, Steven E. Mutsaers, Amama Kanwal, Kaj E C Blokland, Jane Read, Michael Schuliga, Janette K. Burgess, Christopher Grainge, Nathan W. Bartlett, Allen James, Restoring Organ Function by Means of Regenerative Medicine (REGENERATE), and Groningen Research Institute for Asthma and COPD (GRIAC)
- Subjects
Pulmonary and Respiratory Medicine ,Senescence ,Cyclin-Dependent Kinase Inhibitor p21 ,Mitochondrial DNA ,Physiology ,DNA damage ,Alveolar Epithelium ,Drug target ,education ,Biology ,DNA, Mitochondrial ,Pathogenesis ,Idiopathic pulmonary fibrosis ,Physiology (medical) ,Cell Line, Tumor ,medicine ,Deoxyribonuclease I ,Humans ,RNA, Small Interfering ,Cellular Senescence ,Benzofurans ,Etoposide ,Alveolar epithelial cell ,Cell Biology ,respiratory system ,medicine.disease ,Epithelial Cell Adhesion Molecule ,Nucleotidyltransferases ,Idiopathic Pulmonary Fibrosis ,Mitochondria ,respiratory tract diseases ,A549 Cells ,Alveolar Epithelial Cells ,Cancer research ,Cytokines ,RNA Interference ,DNA Damage ,Signal Transduction - Abstract
Alveolar epithelial cell (AEC) senescence is implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF). Mitochondrial dysfunction including release of mitochondrial DNA (mtDNA) is a feature of senescence, which led us to investigate the role of the DNA-sensing guanine monophosphate-adenine monophosphate (GMP-AMP) synthase (cGAS) in IPF, with a focus on AEC senescence. cGAS expression in fibrotic tissue from lungs of patients with IPF was detected within cells immunoreactive for epithelial cell adhesion molecule (EpCAM) and p21, epithelial and senescence markers, respectively. Submerged primary cultures of AECs isolated from lung tissue of patients with IPF (IPF-AECs, n = 5) exhibited higher baseline senescence than AECs from control donors (Ctrl-AECs, n = 5–7), as assessed by increased nuclear histone 2AXγ phosphorylation, p21 mRNA, and expression of senescence-associated secretory phenotype (SASP) cytokines. Pharmacological cGAS inhibition using RU.521 diminished IPF-AEC senescence in culture and attenuated induction of Ctrl-AEC senescence following etoposide-induced DNA damage. Short interfering RNA (siRNA) knockdown of cGAS also attenuated etoposide-induced senescence of the AEC line, A549. Higher levels of mtDNA were detected in the cytosol and culture supernatants of primary IPF- and etoposide-treated Ctrl-AECs when compared with Ctrl-AECs at baseline. Furthermore, ectopic mtDNA augmented cGAS-dependent senescence of Ctrl-AECs, whereas DNAse I treatment diminished IPF-AEC senescence. This study provides evidence that a self-DNA-driven, cGAS-dependent response augments AEC senescence, identifying cGAS as a potential therapeutic target for IPF.
- Published
- 2021
26. Plasma cell but not CD20-mediated B-cell depletion protects from bleomycin-induced lung fibrosis
- Author
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Cecilia M. Prêle, Tylah Miles, David R. Pearce, Robert J. O'Donoghue, Chris Grainge, Lucy Barrett, Kimberly Birnie, Andrew D. Lucas, Svetlana Baltic, Matthias Ernst, Catherine Rinaldi, Geoffrey J. Laurent, Darryl A. Knight, Mark Fear, Gerard Hoyne, Robin J. McAnulty, and Steven E. Mutsaers
- Subjects
Pulmonary and Respiratory Medicine ,Mice ,Bleomycin ,Plasma Cells ,Humans ,Animals ,Lung Diseases, Interstitial ,Lung ,Idiopathic Pulmonary Fibrosis - Abstract
Idiopathic pulmonary fibrosis (IPF) is an interstitial lung disease associated with chronic inflammation and tissue remodelling leading to fibrosis, reduced pulmonary function, respiratory failure and death. Bleomycin (Blm)-induced lung fibrosis in mice replicates several clinical features of human IPF, including prominent lymphoid aggregates of predominantly B-cells that accumulate in the lung adjacent to areas of active fibrosis. We have shown previously a requirement for B-cells in the development of Blm-induced lung fibrosis in mice. To determine the therapeutic potential of inhibiting B-cell function in pulmonary fibrosis, we examined the effects of anti-CD20 B-cell ablation therapy to selectively remove mature B-cells from the immune system and inhibit Blm-induced lung fibrosis. Anti-CD20 B-cell ablation did not reduce fibrosis in this model; however, immune phenotyping of peripheral blood and lung resident cells revealed that anti-CD20-treated mice retained a high frequency of CD19+CD138+plasma cells. Interestingly, high levels of CD138+cells were also identified in the lung tissue of patients with IPF, consistent with the mouse model. Treatment of mice with bortezomib, which depletes plasma cells, reduced the level of Blm-induced lung fibrosis, implicating plasma cells as important effector cells in the development and progression of pulmonary fibrosis.
- Published
- 2022
27. Dysregulated actin cytoskeleton associated with barrier dysfunction in asthma
- Author
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Christopher Grainge, Andrew T. Reid, Peter A. B. Wark, Nathan W. Bartlett, Kristy Nichol, Ngan Fung Li, and Darryl A. Knight
- Subjects
business.industry ,Genetics ,medicine ,medicine.disease ,business ,Actin cytoskeleton ,Molecular Biology ,Biochemistry ,Biotechnology ,Cell biology ,Asthma - Published
- 2021
28. Reduced SOCS1 Expression in Lung Fibroblasts from Patients with IPF Is Not Mediated by Promoter Methylation or Mir155
- Author
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Geoffrey J. Laurent, Philip J. Thompson, David R. Pearce, Sarra E. Jamieson, Matthias Ernst, Tylah Miles, Cecilia M Prêle, Robin J. McAnulty, Darryl A. Knight, Bahareh Badrian, Steven E. Mutsaers, and Thomas Iosifidis
- Subjects
QH301-705.5 ,medicine.medical_treatment ,Medicine (miscellaneous) ,Article ,General Biochemistry, Genetics and Molecular Biology ,fibroblast ,Idiopathic pulmonary fibrosis ,Fibrosis ,Medicine ,SOCS1 ,SOCS3 ,Biology (General) ,STAT3 ,Fibroblast ,Uncategorized ,miR155 ,biology ,business.industry ,Suppressor of cytokine signaling 1 ,fibrosis ,L-6 ,respiratory system ,medicine.disease ,humanities ,respiratory tract diseases ,Cytokine ,medicine.anatomical_structure ,biology.protein ,STAT protein ,Cancer research ,Jak/STAT pathway ,business - Abstract
The interleukin (IL)-6 family of cytokines and exaggerated signal transducer and activator of transcription (STAT)3 signaling is implicated in idiopathic pulmonary fibrosis (IPF) pathogenesis, but the mechanisms regulating STAT3 expression and function are unknown. Suppressor of cytokine signaling (SOCS)1 and SOCS3 block STAT3, and low SOCS1 levels have been reported in IPF fibroblasts and shown to facilitate collagen production. Fibroblasts and lung tissue from IPF patients and controls were used to examine the mechanisms underlying SOCS1 down-regulation in IPF. A significant reduction in basal SOCS1 mRNA in IPF fibroblasts was confirmed. However, there was no difference in the kinetics of activation, and methylation of SOCS1 in control and IPF lung fibroblasts was low and unaffected by 5′-aza-2′-deoxycytidine’ treatment. SOCS1 is a target of microRNA-155 and although microRNA-155 levels were increased in IPF tissue, they were reduced in IPF fibroblasts. Therefore, SOCS1 is not regulated by SOCS1 gene methylation or microRNA155 in these cells. In conclusion, we confirmed that IPF fibroblasts had lower levels of SOCS1 mRNA compared with control fibroblasts, but we were unable to determine the mechanism. Furthermore, although SOCS1 may be important in the fibrotic process, we were unable to find a significant role for SOCS1 in regulating fibroblast function.
- Published
- 2021
29. Pharmacological HIF-1 stabilization promotes intestinal epithelial healing through regulation of α-integrin expression and function
- Author
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Gang Liu, Kyra Minahan, Jay C. Horvat, Bridie J. Goggins, Wai S. Soh, Simonne Sherwin, Simon Keely, Marjorie M. Walker, Jennifer Pryor, Andrea Mathe, Jessica Bruce, and Darryl A. Knight
- Subjects
Physiology ,Colon ,Integrin ,Integrin alpha2 ,Integrin alpha6 ,Cell Movement ,Physiology (medical) ,Cell Line, Tumor ,medicine ,Animals ,Humans ,Intestinal Mucosa ,Cell Proliferation ,Mice, Inbred BALB C ,Wound Healing ,Hepatology ,biology ,Gastroenterology & Hepatology ,Chemistry ,Protein Stability ,Gastroenterology ,Prolyl-Hydroxylase Inhibitors ,Colitis ,Hypoxia-Inducible Factor 1, alpha Subunit ,Epithelium ,Cell biology ,Disease Models, Animal ,medicine.anatomical_structure ,0606 Physiology, 1116 Medical Physiology ,Integrin expression ,Trinitrobenzenesulfonic Acid ,Epithelial restitution ,biology.protein ,Female ,Wound healing ,Integrin alpha Chains ,Function (biology) ,Signal Transduction - Abstract
Intestinal epithelia are critical for maintaining gastrointestinal homeostasis. Epithelial barrier injury, causing inflammation and vascular damage, results in inflammatory hypoxia, and thus, healing occurs in an oxygen-restricted environment. The transcription factor hypoxia-inducible factor (HIF)-1 regulates genes important for cell survival and repair, including the cell adhesion protein β1-integrin. Integrins function as αβ-dimers, and α-integrin-matrix binding is critical for cell migration. We hypothesized that HIF-1 stabilization accelerates epithelial migration through integrin-dependent pathways. We aimed to examine functional and posttranslational activity of α-integrins during HIF-1-mediated intestinal epithelial healing. Wound healing was assessed in T84 monolayers over 24 h with/without prolyl-hydroxylase inhibitor (PHDi) (GB-004), which stabilizes HIF-1. Gene and protein expression were measured by RT-PCR and immunoblot, and α-integrin localization was assessed by immunofluorescence. α-integrin function was assessed by antibody-mediated blockade, and integrin α6 regulation was determined by HIF-1α chromatin immunoprecipitation. Models of mucosal wounding and 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis were used to examine integrin expression and localization in vivo. PHDi treatment accelerated wound closure and migration within 12 h, associated with increased integrin α2 and α6 protein, but not α3. Functional blockade of integrins α2 and α6 inhibited PHDi-mediated accelerated wound closure. HIF-1 bound directly to the integrin α6 promoter. PHDi treatment accelerated mucosal healing, which was associated with increased α6 immunohistochemical staining in wound-associated epithelium and wound-adjacent tissue. PHDi treatment increased α6 protein levels in colonocytes of TNBS mice and induced α6 staining in regenerating crypts and reepithelialized inflammatory lesions. Together, these data demonstrate a role for HIF-1 in regulating both integrin α2 and α6 responses during intestinal epithelial healing.NEW & NOTEWORTHY HIF-1 plays an important role in epithelial restitution, selectively inducing integrins α6 and α2 to promote migration and proliferation, respectively. HIF-stabilizing prolyl-hydroxylase inhibitors accelerate intestinal mucosal healing by inducing epithelial integrin expression.
- Published
- 2021
30. STAT3 Regulates the Onset of Oxidant-induced Senescence in Lung Fibroblasts
- Author
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Lan Wei, Steven E. Mutsaers, Christopher Grainge, Jade Jaffar, Michael Schuliga, Prabuddha S. Pathinayake, Janette K. Burgess, Glen P. Westall, Philip M. Hansbro, Darryl A. Knight, Kaj E C Blokland, Nathan W. Bartlett, Cecilia M. Prêle, David W Waters, Restoring Organ Function by Means of Regenerative Medicine (REGENERATE), and Groningen Research Institute for Asthma and COPD (GRIAC)
- Subjects
0301 basic medicine ,senescence ,Respiratory System ,Clinical Biochemistry ,SECRETORY PHENOTYPE ,Mitochondrion ,fibroblast ,ACTIVATION ,chemistry.chemical_compound ,Idiopathic pulmonary fibrosis ,0302 clinical medicine ,Fibrosis ,Phosphorylation ,STAT3 ,PHOSPHORYLATION ,Lung ,Cellular Senescence ,biology ,Superoxide ,CELLULAR SENESCENCE ,Oxidants ,CANCER ,Mitochondria ,Cell biology ,Protein Transport ,Phenotype ,medicine.anatomical_structure ,signal transducer and activator of transcription 3 ,MITOCHONDRIAL STAT3 ,STAT3 Transcription Factor ,Pulmonary and Respiratory Medicine ,Senescence ,PROTEINS ,Cell Respiration ,03 medical and health sciences ,mitochondrial dysfunction ,medicine ,Humans ,Polycyclic Compounds ,Fibroblast ,Molecular Biology ,Cell Nucleus ,fibrosis ,Editorials ,Cell Biology ,Fibroblasts ,medicine.disease ,GENE ,030104 developmental biology ,030228 respiratory system ,chemistry ,DNA-DAMAGE ,CELLS ,biology.protein ,STAT protein - Abstract
Copyright © 2019 by the American Thoracic Society. Idiopathic pulmonary fibrosis (IPF) is a chronic lung disease of unknown cause with a median survival of only 3 years. Other investigators and we have shown that fibroblasts derived from IPF lungs display characteristics of senescent cells, and that dysregulated activation of the transcription factor signal transducer and activator of transcription 3 (STAT3) correlates with IPF progression. The question of whether STAT3 activation is involved in fibroblast senescence remains unanswered. We hypothesized that inhibiting STAT3 activation after oxidantinduced senescence would attenuate characteristics of the senescent phenotype. We aimed to characterize a model of oxidant-induced senescence in human lung fibroblasts and to determine the effect of inhibiting STAT3 activity on the development of senescence. Exposing human lung fibroblasts to 150 μM hydrogen peroxide (H2O2) resulted in increased senescence-associated β-galactosidase content and expression of p21 and IL-6, all of which are features of senescence. The shift into senescence was accompanied by an increase of STAT3 translocation to the nucleus and mitochondria. Additionally, Seahorse analysis provided evidence of increased mitochondrial respiration characterized by increased basal respiration, proton leak, and an associated increase in superoxide (O2-) production in senescent fibroblasts. Targeting STAT3 activity using the small-molecule inhibitor STA-21 attenuated IL-6 production, reduced p21 levels, decreased senescence-associated b-galactosidase accumulation, and restored normalmitochondrial function. The results of this study illustrate that stress-induced senescence in lung fibroblasts involves the activation of STAT3, which can be pharmacologically modulated.
- Published
- 2019
31. IL-4Rα blockade reduces influenza-associated morbidity in a murine model of allergic asthma
- Author
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Kelly M. McNagny, Darryl A. Knight, Tillie L. Hackett, Delbert R. Dorscheid, Michael R. Hughes, Don D. Sin, Yeni Oh, Kimia Shahangian, Masahiro Niikura, S. F. Paul Man, David A. Ngan, Chung Cheung, Jeremy A. Hirota, Jing Wen, H. H. Rachel Chen, and Anthony Tam
- Subjects
0301 basic medicine ,Male ,Receptors, Cell Surface ,Systemic inflammation ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Influenza A Virus, H1N1 Subtype ,Antigen ,Influenza, Human ,medicine ,Hypersensitivity ,Eosinophilia ,Animals ,Humans ,Asthma ,House dust mite ,lcsh:RC705-779 ,Mice, Inbred BALB C ,biology ,business.industry ,Research ,Pyroglyphidae ,Antibodies, Monoclonal ,lcsh:Diseases of the respiratory system ,medicine.disease ,biology.organism_classification ,Blockade ,respiratory tract diseases ,Disease Models, Animal ,030104 developmental biology ,030228 respiratory system ,Immunology ,Nasal administration ,medicine.symptom ,business ,Viral load - Abstract
Background Asthma was identified as the most common comorbidity in hospitalized patients during the 2009 H1N1 influenza pandemic. We determined using a murine model of allergic asthma whether these mice experienced increased morbidity from pandemic H1N1 (pH1N1) viral infection and whether blockade of interleukin-4 receptor α (IL-4Rα), a critical mediator of Th2 signalling, improved their outcomes. Methods Male BALB/c mice were intranasally sensitized with house dust mite antigen (Der p 1) for 2 weeks; the mice were then inoculated intranasally with a single dose of pandemic H1N1 (pH1N1). The mice were administered intraperitoneally anti-IL-4Rα through either a prophylactic or a therapeutic treatment strategy. Results Infection with pH1N1 of mice sensitized to house dust mite (HDM) led to a 24% loss in weight by day 7 of infection (versus 14% in non-sensitized mice; p Conclusion Together, these data implicate allergic asthma as a significant risk factor for H1N1-related morbidity and reveal a potential therapeutic role for IL-4Rα signalling blockade in reducing the severity of influenza infection in those with allergic airway disease.
- Published
- 2021
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32. Airway mechanical compression: its role in asthma pathogenesis and progression
- Author
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Christopher Grainge, Jin-Ah Park, Andrew T. Reid, Jennifer A. Mitchel, Punnam Chander Veerati, Nathan W. Bartlett, and Darryl A. Knight
- Subjects
0301 basic medicine ,Pulmonary and Respiratory Medicine ,Disease ,Respiratory Mucosa ,Models, Biological ,Article ,Pathogenesis ,03 medical and health sciences ,0302 clinical medicine ,In vivo ,medicine ,Humans ,Asthma ,lcsh:RC705-779 ,Lung ,business.industry ,lcsh:Diseases of the respiratory system ,respiratory system ,medicine.disease ,respiratory tract diseases ,ErbB Receptors ,030104 developmental biology ,medicine.anatomical_structure ,030228 respiratory system ,Immunology ,Respiratory epithelium ,Bronchoconstriction ,Goblet Cells ,Stress, Mechanical ,medicine.symptom ,Airway ,business - Abstract
The lung is a mechanically active organ, but uncontrolled or excessive mechanical forces disrupt normal lung function and can contribute to the development of disease. In asthma, bronchoconstriction leads to airway narrowing and airway wall buckling. A growing body of evidence suggests that pathological mechanical forces induced by airway buckling alone can perpetuate disease processes in asthma. Here, we review the data obtained from a variety of experimental models, includingin vitro,ex vivoandin vivoapproaches, which have been used to study the impact of mechanical forces in asthma pathogenesis. We review the evidence showing that mechanical compression alters the biological and biophysical properties of the airway epithelium, including activation of the epidermal growth factor receptor pathway, overproduction of asthma-associated mediators, goblet cell hyperplasia, and a phase transition of epithelium from a static jammed phase to a mobile unjammed phase. We also define questions regarding the impact of mechanical forces on the pathology of asthma, with a focus on known triggers of asthma exacerbations such as viral infection.
- Published
- 2020
33. Regulation of cellular senescence by extracellular matrix during chronic fibrotic diseases
- Author
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Simon D. Pouwels, Kaj E C Blokland, Janette K. Burgess, Michael Schuliga, Darryl A. Knight, Groningen Research Institute for Asthma and COPD (GRIAC), and Restoring Organ Function by Means of Regenerative Medicine (REGENERATE)
- Subjects
0301 basic medicine ,Senescence ,Aging ,Cell Homeostasis & Autophagy ,Antifibrotics ,extracellular matrix ,Disease ,Cell Communication ,Matrix (biology) ,Molecular Bases of Health & Disease ,Pathogenesis ,Extracellular matrix ,03 medical and health sciences ,0302 clinical medicine ,Fibrosis ,medicine ,Animals ,Homeostasis ,Humans ,Review Articles ,Cellular Senescence ,DAMPs ,business.industry ,Senolytics ,Mesenchymal stem cell ,fibrosis ,General Medicine ,medicine.disease ,030104 developmental biology ,030220 oncology & carcinogenesis ,Chronic Disease ,Cell Cycle, Growth & Proliferation ,Cancer research ,Cell Migration, Adhesion & Morphology ,business - Abstract
The extracellular matrix (ECM) is a complex network of macromolecules surrounding cells providing structural support and stability to tissues. The understanding of the ECM and the diverse roles it plays in development, homoeostasis and injury have greatly advanced in the last three decades. The ECM is crucial for maintaining tissue homoeostasis but also many pathological conditions arise from aberrant matrix remodelling during ageing. Ageing is characterised as functional decline of tissue over time ultimately leading to tissue dysfunction, and is a risk factor in many diseases including cardiovascular disease, diabetes, cancer, dementia, glaucoma, chronic obstructive pulmonary disease (COPD) and fibrosis. ECM changes are recognised as a major driver of aberrant cell responses. Mesenchymal cells in aged tissue show signs of growth arrest and resistance to apoptosis, which are indicative of cellular senescence. It was recently postulated that cellular senescence contributes to the pathogenesis of chronic fibrotic diseases in the heart, kidney, liver and lung. Senescent cells negatively impact tissue regeneration while creating a pro-inflammatory environment as part of the senescence-associated secretory phenotype (SASP) favouring disease progression. In this review, we explore and summarise the current knowledge around how aberrant ECM potentially influences the senescent phenotype in chronic fibrotic diseases. Lastly, we will explore the possibility for interventions in the ECM–senescence regulatory pathways for therapeutic potential in chronic fibrotic diseases.
- Published
- 2020
34. The contribution of animal models to understanding the role of the immune system in human idiopathic pulmonary fibrosis
- Author
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Gerard F. Hoyne, Steven E Mutsaers, Darryl A. Knight, Cecilia M Prêle, Mark W. Fear, and Tylah Miles
- Subjects
0301 basic medicine ,lcsh:Immunologic diseases. Allergy ,Immunology ,Inflammation ,Bleomycin ,Pathogenesis ,Lung Disorder ,03 medical and health sciences ,chemistry.chemical_compound ,Idiopathic pulmonary fibrosis ,0302 clinical medicine ,Immune system ,Special Feature Review ,Pulmonary fibrosis ,medicine ,Immunology and Allergy ,General Nursing ,Lung ,bleomycin ,business.industry ,medicine.disease ,fibrogenesis ,animal models ,innate and adaptive immune system ,030104 developmental biology ,medicine.anatomical_structure ,030228 respiratory system ,chemistry ,inflammation ,medicine.symptom ,business ,lcsh:RC581-607 - Abstract
Pulmonary fibrosis occurs in a heterogeneous group of lung disorders and is characterised by an excessive deposition of extracellular matrix proteins within the pulmonary interstitium, leading to impaired gas transfer and a loss of lung function. In the past 10 years, there has been a dramatic increase in our understanding of the immune system and how it contributes to fibrogenic processes within the lung. This review will compare some of the models used to investigate the pathogenesis and treatment of pulmonary fibrosis, in particular those used to study immune cell pathogenicity in idiopathic pulmonary fibrosis, highlighting their advantages and disadvantages in dissecting human disease., This review will compare some of the models used to investigate the pathogenesis and treatment of pulmonary fibrosis, in particular those used to study immune cell pathogenicity in idiopathic pulmonary fibrosis, highlighting their advantages, disadvantages and suitability in representing the human disease.
- Published
- 2020
35. Intrinsic Asthma and Type-2 Cytokines Mediated STAT1 Response to Rhinovirus in Bronchial Epithelial Cells
- Author
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Nathan W. Bartlett, Lan Wei, Christopher Grainge, Jane Read, Darryl A. Knight, and Andrew T. Reid
- Subjects
biology ,business.industry ,Intrinsic asthma ,Immunology ,biology.protein ,medicine ,STAT1 ,Rhinovirus ,medicine.disease_cause ,business - Published
- 2020
36. The Fibrogenic Actions of IL-25 and Its Potential Role in Idiopathic Pulmonary Fibrosis (IPF)
- Author
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A. Kanwal, Nathan W. Bartlett, Christopher Grainge, Michael Schuliga, and Darryl A. Knight
- Subjects
Pathology ,medicine.medical_specialty ,Idiopathic pulmonary fibrosis ,business.industry ,medicine ,medicine.disease ,business - Published
- 2020
37. Senescence of IPF Lung Fibroblasts Disrupt Alveolar Epithelial Cell Proliferation and Promote Migration in Wound Healing
- Author
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Michael Schuliga, Cecilia M Prêle, Steven E Mutsaers, Jane Read, Simon D. Pouwels, Janette K. Burgess, Darryl A. Knight, Kaj E C Blokland, Christopher Grainge, David W Waters, Jade Jaffar, Glen P. Westall, Groningen Research Institute for Asthma and COPD (GRIAC), and Restoring Organ Function by Means of Regenerative Medicine (REGENERATE)
- Subjects
Senescence ,STRESS ,senescence ,lcsh:RS1-441 ,Pharmaceutical Science ,Article ,lcsh:Pharmacy and materia medica ,03 medical and health sciences ,Idiopathic pulmonary fibrosis ,0302 clinical medicine ,Fibrosis ,fibroblasts ,Parenchyma ,INJURY ,aberrant repair ,Medicine ,alveolar epithelial cell ,030304 developmental biology ,0303 health sciences ,Lung ,IDIOPATHIC PULMONARY ,business.industry ,fibrosis ,respiratory system ,medicine.disease ,Phenotype ,Pathophysiology ,respiratory tract diseases ,cell-cycle inhibition ,medicine.anatomical_structure ,030220 oncology & carcinogenesis ,Cancer research ,business ,Wound healing - Abstract
Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease marked by excessive accumulation of lung fibroblasts (LFs) and collagen in the lung parenchyma. The mechanisms that underlie IPF pathophysiology are thought to reflect repeated alveolar epithelial injury leading to an aberrant wound repair response. Recent work has shown that IPF-LFs display increased characteristics of senescence including growth arrest and a senescence-associated secretory phenotype (SASP) suggesting that senescent LFs contribute to dysfunctional wound repair process. Here, we investigated the influence of senescent LFs on alveolar epithelial cell repair responses in a co-culture system. Alveolar epithelial cell proliferation was attenuated when in co-culture with cells or conditioned media from, senescence-induced control LFs or IPF-LFs. Cell-cycle analyses showed that a larger number of epithelial cells were arrested in G2/M phase when co-cultured with IPF-LFs, than in monoculture. Paradoxically, the presence of LFs resulted in increased A549 migration after mechanical injury. Our data suggest that senescent LFs may contribute to aberrant re-epithelialization by inhibiting proliferation in IPF.
- Published
- 2020
38. Epithelial cell dysfunction, a major driver of asthma development
- Author
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Ian Sayers, Darryl A. Knight, Tania Maes, Irene H. Heijink, Martijn C. Nawijn, Virinchi N. S. Kuchibhotla, and Mirjam P. Roffel
- Subjects
0301 basic medicine ,HAY-FEVER ,TYPE-2 INFLAMMATION ,Respiratory System ,RESPIRATORY SYNCYTIAL VIRUS ,Disease ,Review Article ,Epithelium ,type 2 responses ,0302 clinical medicine ,immune system diseases ,Medicine and Health Sciences ,Immunology and Allergy ,Medicine ,Review Articles ,Barrier function ,(epi)genetics ,respiratory system ,medicine.anatomical_structure ,type 2 ,THYMIC STROMAL LYMPHOPOIETIN ,VIRUS ,epithelial barrier ,TRANSITION ,GROWTH-FACTOR ,Thymic stromal lymphopoietin ,Immunology ,Respiratory Mucosa ,airway remodelling ,03 medical and health sciences ,E-CADHERIN ,HOUSE-DUST MITE ,MESENCHYMAL ,Humans ,Epigenetics ,RESPIRATORY SYNCYTIAL ,Asthma ,MESENCHYMAL TRANSITION ,business.industry ,Epithelial Cells ,AIRWAY RESPONSIVENESS ,asthma ,Allergens ,medicine.disease ,respiratory tract diseases ,030104 developmental biology ,030228 respiratory system ,BARRIER FUNCTION ,responses ,Respiratory epithelium ,business ,Airway - Abstract
Airway epithelial barrier dysfunction is frequently observed in asthma and may have important implications. The physical barrier function of the airway epithelium is tightly interwoven with its immunomodulatory actions, while abnormal epithelial repair responses may contribute to remodelling of the airway wall. We propose that abnormalities in the airway epithelial barrier play a crucial role in the sensitization to allergens and pathogenesis of asthma. Many of the identified susceptibility genes for asthma are expressed in the airway epithelium, supporting the notion that events at the airway epithelial surface are critical for the development of the disease. However, the exact mechanisms by which the expression of epithelial susceptibility genes translates into a functionally altered response to environmental risk factors of asthma are still unknown. Interactions between genetic factors and epigenetic regulatory mechanisms may be crucial for asthma susceptibility. Understanding these mechanisms may lead to identification of novel targets for asthma intervention by targeting the airway epithelium. Moreover, exciting new insights have come from recent studies using single‐cell RNA sequencing (scRNA‐Seq) to study the airway epithelium in asthma. This review focuses on the role of airway epithelial barrier function in the susceptibility to develop asthma and novel insights in the modulation of epithelial cell dysfunction in asthma.
- Published
- 2020
39. Blocking Notch3 Signaling Abolishes MUC5AC Production in Airway Epithelial Cells from Individuals with Asthma
- Author
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Nathan W. Bartlett, Kristy Nichol, Philip M. Hansbro, Fatemeh Moheimani, Stephen M. Stick, Christopher Grainge, Andrew T. Reid, Peter A. B. Wark, Punnam Chander Veerati, Darryl A. Knight, and Anthony Kicic
- Subjects
0301 basic medicine ,Pulmonary and Respiratory Medicine ,Male ,Clinical Biochemistry ,Notch signaling pathway ,Bronchi ,Respiratory Mucosa ,Biology ,Mucin 5AC ,03 medical and health sciences ,0302 clinical medicine ,medicine ,Humans ,RNA, Small Interfering ,Molecular Biology ,Lung ,Receptor, Notch3 ,Cells, Cultured ,Aged ,Gene knockdown ,Goblet cell ,Mucin ,Cell Differentiation ,Epithelial Cells ,Cell Biology ,respiratory system ,Middle Aged ,Mucus ,Epithelium ,Asthma ,respiratory tract diseases ,Cell biology ,030104 developmental biology ,medicine.anatomical_structure ,030228 respiratory system ,Respiratory epithelium ,Female ,Goblet Cells ,Airway ,Signal Transduction - Abstract
In asthma, goblet cell numbers are increased within the airway epithelium, perpetuating the production of mucus that is more difficult to clear and results in airway mucus plugging. Notch1, Notch2, or Notch3, or a combination of these has been shown to influence the differentiation of airway epithelial cells. How the expression of specific Notch isoforms differs in fully differentiated adult asthmatic epithelium and whether Notch influences mucin production after differentiation is currently unknown. We aimed to quantify different Notch isoforms in the airway epithelium of individuals with severe asthma and to examine the impact of Notch signaling on mucin MUC5AC. Human lung sections and primary bronchial epithelial cells from individuals with and without asthma were used in this study. Primary bronchial epithelial cells were differentiated at the air-liquid interface for 28 days. Notch isoform expression was analyzed by Taqman quantitative PCR. Immunohistochemistry was used to localize and quantify Notch isoforms in human airway sections. Notch signaling was inhibited in vitro using dibenzazepine or Notch3-specific siRNA, followed by analysis of MUC5AC. NOTCH3 was highly expressed in asthmatic airway epithelium compared with nonasthmatic epithelium. Dibenzazepine significantly reduced MUC5AC production in air-liquid interface cultures of primary bronchial epithelial cells concomitantly with suppression of NOTCH3 intracellular domain protein. Specific knockdown using NOTCH3 siRNA recapitulated the dibenzazepine-induced reduction in MUC5AC. We demonstrate that NOTCH3 is a regulator of MUC5AC production. Increased NOTCH3 signaling in the asthmatic airway epithelium may therefore be an underlying driver of excess MUC5AC production.
- Published
- 2020
40. IL-4Rα Blockade Reduces Influenza-Associated Morbidity in a Murine Model of Allergic Asthma
- Author
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Kimia Shahangian, David A. Ngan, H.H. Rachel Chen, Yeni Oh, Darryl A. Knight, Delbert Dorscheid, Tillie L. Hackett, Michael R. Hughes, Kelly M. McNagny, Jeremy A. Hirota, Masahiro Niikura, S.F. Paul Man, and Donald Sin
- Published
- 2020
41. Mitochondrial dysfunction contributes to the senescent phenotype of IPF lung fibroblasts
- Author
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Cecilia M. Prêle, David W Waters, Michael Schuliga, Darryl A. Knight, Jade Jaffar, Cory M. Hogaboam, Christopher Grainge, Kaj E C Blokland, Nasreen Khalil, Jane Read, Steven E. Mutsaers, Dmitri V. Pechkovsky, Janette K. Burgess, Glen P. Westall, Andrew T. Reid, Groningen Research Institute for Asthma and COPD (GRIAC), and Restoring Organ Function by Means of Regenerative Medicine (REGENERATE)
- Subjects
0301 basic medicine ,senescence ,antioxidant ,endogenous compound ,mTORC1 ,reactive oxygen metabolite ,Mitochondrion ,DNA damage response ,etoposide ,cyclin‐dependent kinase inhibitors ,rotenone ,cyclin-dependent kinase inhibitors ,stress ,chemistry.chemical_compound ,homeostasis ,peroxisome proliferator‐activated receptor gamma coactivator 1‐alpha ,mitochondrion ,Myofibroblasts ,Lung ,Cellular Senescence ,primary culture ,Superoxide ,adult ,article ,respiratory system ,idiopathic pulmonary fibrosis ,mechanistic target of rapamycin complex 1 ,enzyme activity ,Cell biology ,mitochondria ,medicine.anatomical_structure ,Molecular Medicine ,Original Article ,superoxide ,Signal transduction ,signal transduction ,Senescence ,phenotype ,DNA damage ,Down-Regulation ,cyclin dependent kinase inhibitor ,peroxisome proliferator-activated receptor gamma coactivator 1-alpha ,Cyclic N-Oxides ,03 medical and health sciences ,peroxisome proliferator activated receptor gamma coactivator 1alpha ,fibroblasts ,medicine ,Humans ,controlled study ,human ,Fibroblast ,Sirolimus ,reactive oxygen species and mitoTEMPO ,rapamycin ,human cell ,Original Articles ,mammalian target of rapamycin complex 1 ,Cell Biology ,Acetylcysteine ,respiratory tract diseases ,fibrosing alveolitis ,030104 developmental biology ,chemistry ,lung fibroblast ,Biomarkers ,Homeostasis - Abstract
Increasing evidence highlights that senescence plays an important role in idiopathic pulmonary fibrosis (IPF). This study delineates the specific contribution of mitochondria and the superoxide they form to the senescent phenotype of lung fibroblasts from IPF patients (IPF‐LFs). Primary cultures of IPF‐LFs exhibited an intensified DNA damage response (DDR) and were more senescent than age‐matched fibroblasts from control donors (Ctrl‐LFs). Furthermore, IPF‐LFs exhibited mitochondrial dysfunction, exemplified by increases in mitochondrial superoxide, DNA, stress and activation of mTORC1. The DNA damaging agent etoposide elicited a DDR and augmented senescence in Ctrl‐LFs, which were accompanied by disturbances in mitochondrial homoeostasis including heightened superoxide production. However, etoposide had no effect on IPF‐LFs. Mitochondrial perturbation by rotenone involving sharp increases in superoxide production also evoked a DDR and senescence in Ctrl‐LFs, but not IPF‐LFs. Inhibition of mTORC1, antioxidant treatment and a mitochondrial targeting antioxidant decelerated IPF‐LF senescence and/or attenuated pharmacologically induced Ctrl‐LF senescence. In conclusion, increased superoxide production by dysfunctional mitochondria reinforces lung fibroblast senescence via prolongation of the DDR. As part of an auto‐amplifying loop, mTORC1 is activated, altering mitochondrial homoeostasis and increasing superoxide production. Deeper understanding the mechanisms by which mitochondria contribute to fibroblast senescence in IPF has potentially important therapeutic implications.
- Published
- 2018
42. Influenza A virus infection dysregulates the expression of microRNA-22 and its targets; CD147 and HDAC4, in epithelium of asthmatics
- Author
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Darryl A. Knight, Fatemeh Moheimani, Jorinke Koops, Philip M. Hansbro, Teresa Williams, Peter A. B. Wark, Andrew T. Reid, and Faculteit Medische Wetenschappen/UMCG
- Subjects
Male ,0301 basic medicine ,Severe asthma ,Respiratory System ,Epithelial cells ,medicine.disease_cause ,chemistry.chemical_compound ,Influenza A Virus, H1N1 Subtype ,Influenza A virus ,Cells, Cultured ,microRNA ,PROLIFERATION ,Airway remodeling ,Middle Aged ,3. Good health ,medicine.anatomical_structure ,DIFFERENTIATION ,Female ,Adult ,RNA EXPRESSION ,Respiratory Mucosa ,Biology ,Histone Deacetylases ,03 medical and health sciences ,Immune system ,Influenza, Human ,medicine ,Humans ,Antagomir ,Aged ,lcsh:RC705-779 ,Messenger RNA ,MESENCHYMAL TRANSITION ,Innate immune system ,H1N1 INFLUENZA ,Research ,MIR-22 ,lcsh:Diseases of the respiratory system ,Asthma ,Epithelium ,Repressor Proteins ,AIRWAY EPITHELIUM ,MicroRNAs ,030104 developmental biology ,chemistry ,Immunology ,CELLS ,Basigin ,INNATE IMMUNITY ,Respiratory epithelium ,INFLAMMATORY RESPONSES - Abstract
Background Specific microRNAs (miRNAs) play essential roles in airway remodeling in asthma. Infection with influenza A virus (IAV) may also magnify pre-existing airway remodeling leading to asthma exacerbation. However, these events remain to be fully defined. We investigated the expression of miRNAs with diverse functions including proliferation (miR-20a), differentiation (miR-22) or innate/adaptive immune responses (miR-132) in primary bronchial epithelial cells (pBECs) of asthmatics following infection with the H1N1 strain of IAV. Methods pBECs from subjects (n = 5) with severe asthma and non-asthmatics were cultured as submerged monolayers or at the air-liquid-interface (ALI) conditions and incubated with IAV H1N1 (MOI 5) for up to 24 h. Isolated miRNAs were subjected to Taqman miRNAs assays. We confirmed miRNA targets using a specific mimic and antagomir. Taqman mRNAs assays and immunoblotting were used to assess expression of target genes and proteins, respectively. Results At baseline, these miRNAs were expressed at the same level in pBECs of asthmatics and non-asthmatics. After 24 h of infection, miR-22 expression increased significantly which was associated with the suppression of CD147 mRNA and HDAC4 mRNA and protein expression in pBECs from non-asthmatics, cultured in ALI. In contrast, miR-22 remained unchanged while CD147 expression increased and HDAC4 remained unaffected in cells from asthmatics. IAV H1N1 mediated increases in SP1 and c-Myc transcription factors may underpin the induction of CD147 in asthmatics. Conclusion The different profile of miR-22 expression in differentiated epithelial cells from non-asthmatics may indicate a self-defense mechanism against aberrant epithelial responses through suppressing CD147 and HDAC4, which is compromised in epithelial cells of asthmatics. Electronic supplementary material The online version of this article (10.1186/s12931-018-0851-7) contains supplementary material, which is available to authorized users.
- Published
- 2018
43. The fibrogenic actions of the coagulant and plasminogen activation systems in pulmonary fibrosis
- Author
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Michael Schuliga, Glen P. Westall, Christopher Grainge, and Darryl A. Knight
- Subjects
0301 basic medicine ,Plasmin ,Pulmonary Fibrosis ,Biochemistry ,Plasminogen Activators ,03 medical and health sciences ,Idiopathic pulmonary fibrosis ,Fibrosis ,Pulmonary fibrosis ,medicine ,Animals ,Humans ,Fibrin ,Factor XII ,biology ,Chemistry ,Fibrinolysis ,Plasminogen ,Cell Biology ,respiratory system ,medicine.disease ,Urokinase receptor ,030104 developmental biology ,Cancer research ,biology.protein ,Vitronectin ,Plasminogen activator ,medicine.drug - Abstract
Fibrosis causes irreversible damage to lung structure and function in restrictive lung diseases such as idiopathic pulmonary fibrosis (IPF). Extravascular coagulation involving fibrin formation in the intra-alveolar compartment is postulated to have a pivotal role in the development of pulmonary fibrosis, serving as a provisional matrix for migrating fibroblasts. Furthermore, proteases of the coagulation and plasminogen activation (plasminergic) systems that form and breakdown fibrin respectively directly contribute to pulmonary fibrosis. The coagulants, thrombin and factor Xa (FXa) evoke fibrogenic effects via cleavage of the N-terminus of protease-activated receptors (PARs). Whilst the formation and activity of plasmin, the principle plasminergic mediator is suppressed in the airspaces of patients with IPF, localized increases are likely to occur in the lung interstitium. Plasmin-evoked proteolytic activation of factor XII (FXII), matrix metalloproteases (MMPs) and latent, matrix-bound growth factors such as epidermal growth factor (EGF) indirectly implicate plasmin in pulmonary fibrosis. Another plasminergic protease, urokinase plasminogen activator (uPA) is associated with regions of fibrosis in the remodelled lung of IPF patients and elicits fibrogenic activity via binding its receptor (uPAR). Plasminogen activator inhibitor-1 (PAI-1) formed in the injured alveolar epithelium also contributes to pulmonary fibrosis in a manner that involves vitronectin binding. This review describes the mechanisms by which components of the two systems primarily involved in fibrin homeostasis contribute to interstitial fibrosis, with a particular focus on IPF. Selectively targeting the receptor-mediated mechanisms of coagulant and plasminergic proteases may limit pulmonary fibrosis, without the bleeding complications associated with conventional anti-coagulant and thrombolytic therapies.
- Published
- 2018
44. Autophagy and the unfolded protein response promote profibrotic effects of TGF-β1 in human lung fibroblasts
- Author
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Nicholas J. Kenyon, Ian M.C. Dixon, Afshin Samali, John B. Patterson, Andrew J. Halayko, Javad Alizadeh, Helmut Unruh, Adel Rezaei Moghadam, Amir A. Zeki, Darryl A. Knight, Shahla Shojaei, Martin Post, Saeid Ghavami, Thomas Klonisch, Sean Ott, and Behzad Yeganeh
- Subjects
0301 basic medicine ,Pulmonary and Respiratory Medicine ,Pathology ,medicine.medical_specialty ,XBP1 ,biology ,Physiology ,Autophagy ,Cell Biology ,respiratory system ,medicine.disease ,respiratory tract diseases ,3. Good health ,Fibronectin ,03 medical and health sciences ,Idiopathic pulmonary fibrosis ,030104 developmental biology ,Fibrosis ,Physiology (medical) ,Pulmonary fibrosis ,biology.protein ,medicine ,Cancer research ,Unfolded protein response ,Transforming growth factor - Abstract
Idiopathic pulmonary fibrosis (IPF) is a lethal fibrotic lung disease in adults with limited treatment options. Autophagy and the unfolded protein response (UPR), fundamental processes induced by cell stress, are dysregulated in lung fibroblasts and epithelial cells from humans with IPF. Human primary cultured lung parenchymal and airway fibroblasts from non-IPF and IPF donors were stimulated with transforming growth factor-β1 (TGF-β1) with or without inhibitors of autophagy or UPR (IRE1 inhibitor). Using immunoblotting, we monitored temporal changes in abundance of protein markers of autophagy (LC3βII and Atg5-12), UPR (BIP, IRE1α, and cleaved XBP1), and fibrosis (collagen 1α2 and fibronectin). Using fluorescent immunohistochemistry, we profiled autophagy (LC3βII) and UPR (BIP and XBP1) markers in human non-IPF and IPF lung tissue. TGF-β1-induced collagen 1α2 and fibronectin protein production was significantly higher in IPF lung fibroblasts compared with lung and airway fibroblasts from non-IPF donors. TGF-β1 induced the accumulation of LC3βII in parallel with collagen 1α2 and fibronectin, but autophagy marker content was significantly lower in lung fibroblasts from IPF subjects. TGF-β1-induced collagen and fibronectin biosynthesis was significantly reduced by inhibiting autophagy flux in fibroblasts from the lungs of non-IPF and IPF donors. Conversely, only in lung fibroblasts from IPF donors did TGF-β1 induce UPR markers. Treatment with an IRE1 inhibitor decreased TGF-β1-induced collagen 1α2 and fibronectin biosynthesis in IPF lung fibroblasts but not those from non-IPF donors. The IRE1 arm of the UPR response is uniquely induced by TGF-β1 in lung fibroblasts from human IPF donors and is required for excessive biosynthesis of collagen and fibronectin in these cells.
- Published
- 2018
45. Accumulation mode particles and LPS exposure induce TLR-4 dependent and independent inflammatory responses in the lung
- Author
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Graeme R. Zosky, Darryl A. Knight, Erika N. Sutanto, Debra J. Turner, Anthony Kicic, Paul S. McNamara, Angela Fonceca, Peter D. Sly, Elizabeth M. Bozanich, and Stephen M. Stick
- Subjects
0301 basic medicine ,Lipopolysaccharides ,LPS ,Lipopolysaccharide ,PM ,medicine.medical_treatment ,Allergic inflammation ,03 medical and health sciences ,chemistry.chemical_compound ,Mice ,0302 clinical medicine ,Airway resistance ,medicine ,Animals ,COPD ,Receptor ,Lung ,AMP ,Inflammation ,Mice, Knockout ,lcsh:RC705-779 ,Mice, Inbred C3H ,Chemistry ,Research ,Airway Resistance ,TLR-4 ,lcsh:Diseases of the respiratory system ,Asthma ,Toll-Like Receptor 4 ,030104 developmental biology ,Cytokine ,medicine.anatomical_structure ,030228 respiratory system ,Immunology ,TLR4 ,Tumor necrosis factor alpha ,Particulate Matter ,lipids (amino acids, peptides, and proteins) ,Inflammation Mediators - Abstract
Background: Accumulation mode particles (AMP) are formed from engine combustion and make up the inhalable vapour cloud of ambient particulate matter pollution. Their small size facilitates dispersal and subsequent exposure far from their original source, as well as the ability to penetrate alveolar spaces and capillary walls of the lung when inhaled. A significant immuno-stimulatory component of AMP is lipopolysaccharide (LPS), a product of Gram negative bacteria breakdown. As LPS is implicated in the onset and exacerbation of asthma, the presence or absence of LPS in ambient particulate matter (PM) may explain the onset of asthmatic exacerbations to PM exposure. This study aimed to delineate the effects of LPS and AMP on airway inflammation, and potential contribution to airways disease by measuring airway inflammatory responses induced via activation of the LPS cellular receptor, Toll-like receptor 4 (TLR-4). Methods: The effects of nebulized AMP, LPS and AMP administered with LPS on lung function, cellular inflammatory infiltrate and cytokine responses were compared between wildtype mice and mice not expressing TLR-4. Results: The presence of LPS administered with AMP appeared to drive elevated airway resistance and sensitivity via TLR-4. Augmented TLR4 driven eosinophilia and greater TNF-α responses observed in AMP-LPS treated mice independent of TLR-4 expression, suggests activation of allergic responses by TLR4 and non-TLR4 pathways larger than those induced by LPS administered alone. Treatment with AMP induced macrophage recruitment independent of TLR-4 expression. Conclusions: These findings suggest AMP-LPS as a stronger stimulus for allergic inflammation in the airways then LPS alone.
- Published
- 2018
46. Conditionally reprogrammed primary airway epithelial cells maintain morphology, lineage and disease specific functional characteristics
- Author
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Francis J. Lannigan, K. Martinovich, Erika N. Sutanto, Stephen M. Stick, Darryl A. Knight, Anthony Kicic, Samuel T. Montgomery, Nicole C. Shaw, Alysia G. Buckley, Luke W. Garratt, Kevin Looi, Thomas Iosifidis, E. Kicic-Starcevich, and Kak-Ming Ling
- Subjects
0301 basic medicine ,Male ,Lineage (genetic) ,Cystic Fibrosis ,Cellular differentiation ,Population ,lcsh:Medicine ,Respiratory Mucosa ,Biology ,Cystic fibrosis ,Article ,03 medical and health sciences ,Mice ,medicine ,Animals ,Humans ,Cell Lineage ,Cellular Reprogramming Techniques ,Fibroblast ,education ,lcsh:Science ,Cells, Cultured ,education.field_of_study ,Multidisciplinary ,lcsh:R ,Cell Differentiation ,Fibroblasts ,medicine.disease ,Phenotype ,Asthma ,Cell biology ,030104 developmental biology ,medicine.anatomical_structure ,Child, Preschool ,Female ,lcsh:Q ,Airway - Abstract
Current limitations to primary cell expansion led us to test whether airway epithelial cells derived from healthy children and those with asthma and cystic fibrosis (CF), co-cultured with an irradiated fibroblast feeder cell in F-medium containing 10 µM ROCK inhibitor could maintain their lineage during expansion and whether this is influenced by underlying disease status. Here, we show that conditionally reprogrammed airway epithelial cells (CRAECs) can be established from both healthy and diseased phenotypes. CRAECs can be expanded, cryopreserved and maintain phenotypes over at least 5 passages. Population doublings of CRAEC cultures were significantly greater than standard cultures, but maintained their lineage characteristics. CRAECs from all phenotypes were also capable of fully differentiating at air-liquid interface (ALI) and maintained disease specific characteristics including; defective CFTR channel function cultures and the inability to repair wounds. Our findings indicate that CRAECs derived from children maintain lineage, phenotypic and importantly disease-specific functional characteristics over a specified passage range.
- Published
- 2017
47. A Senescence Bystander Effect in Human Lung Fibroblasts
- Author
-
Kaj E C Blokland, Glen P. Westall, Cecilia M. Prêle, Lan Wei, Prabuddha S. Pathinayake, David W Waters, Michael Schuliga, Jade Jaffar, Darryl A. Knight, Hui Ying Tan, Janette K. Burgess, Steven E. Mutsaers, Christopher Grainge, Groningen Research Institute for Asthma and COPD (GRIAC), and Restoring Organ Function by Means of Regenerative Medicine (REGENERATE)
- Subjects
collagen ,A549 cell ,Senescence ,senescence ,QH301-705.5 ,lung fibroblasts ,Medicine (miscellaneous) ,respiratory system ,Biology ,medicine.disease ,Phenotype ,Article ,General Biochemistry, Genetics and Molecular Biology ,In vitro ,respiratory tract diseases ,Idiopathic pulmonary fibrosis ,Parenchyma ,medicine ,Bystander effect ,Cancer research ,idiopathic pulmonary fibrosis (IPF) ,Biology (General) ,Etoposide ,medicine.drug - Abstract
Idiopathic pulmonary fibrosis (IPF) is a chronic disease characterised by a dense fibrosing of the lung parenchyma. An association between IPF and cellular senescence is well established and several studies now describe a higher abundance of senescent fibroblasts and epithelial cells in the lungs of IPF patients compared with age-matched controls. The cause of this abnormal accumulation of senescent cells is unknown but evidence suggests that, once established, senescence can be transferred from senescent to non-senescent cells. In this study, we investigated whether senescent human lung fibroblasts (LFs) and alveolar epithelial cells (AECs) could induce a senescent-like phenotype in “naïve” non-senescent LFs in vitro. Primary cultures of LFs from adult control donors (Ctrl-LFs) with a low baseline of senescence were exposed to conditioned medium (CM) from: (i) Ctrl-LFs induced to become senescent using H2O2 or etoposide, (ii) LFs derived from IPF patients (IPF-LFs) with a high baseline of senescence, or (iii) senescence-induced A549 cells, an AEC line. Additionally, ratios of non-senescent Ctrl-LFs and senescence-induced Ctrl-LFs (100:0, 0:100, 50:50, 90:10, 99:1) were co-cultured and their effect on induction of senescence measured. We demonstrated that exposure of naïve non-senescent Ctrl-LFs to CM from senescence-induced Ctrl-LFs and AECs and IPF-LFs increased the markers of senescence including nuclear localisation of phosphorylated-H2A histone family member X (H2AXγ) and expression of p21, IL-6 and IL-8 in Ctrl-LFs. Additionally, co-cultures of non-senescent and senescence-induced Ctrl-LFs induced a senescent-like phenotype in the non-senescent cells. These data suggest that the phenomenon of “senescence-induced senescence” can occur in vitro in primary cultures of human LFs, and provides a possible explanation for the abnormal abundance of senescent cells in the lungs of IPF patients.
- Published
- 2021
48. Ageing mechanisms that contribute to tissue remodeling in lung disease
- Author
-
Michael Schuliga, Jane Read, and Darryl A. Knight
- Subjects
Lung Diseases ,Aging ,ARDS ,Lung injury ,Biochemistry ,Idiopathic pulmonary fibrosis ,Fibrosis ,Humans ,Medicine ,Lung ,Pandemics ,Molecular Biology ,Aged ,COPD ,SARS-CoV-2 ,business.industry ,COVID-19 ,Immunosenescence ,medicine.disease ,medicine.anatomical_structure ,Neurology ,Ageing ,Immunology ,business ,Biotechnology - Abstract
Age is a major risk factor for chronic respiratory diseases such as idiopathic pulmonary fibrosis (IPF), chronic obstructive pulmonary disease (COPD) and certain phenotypes of asthma. The recent COVID-19 pandemic also highlights the increased susceptibility of the elderly to acute respiratory distress syndrome (ARDS), a diffuse inflammatory lung injury with often long-term effects (ie parenchymal fibrosis). Collectively, these lung conditions are characterized by a pathogenic reparative process that, rather than restoring organ function, contributes to structural and functional tissue decline. In the ageing lung, the homeostatic control of wound healing following challenge or injury has an increased likelihood of being perturbed, increasing susceptibility to disease. This loss of fidelity is a consequence of a diverse range of underlying ageing mechanisms including senescence, mitochondrial dysfunction, proteostatic stress and diminished autophagy that occur within the lung, as well as in other tissues, organs and systems of the body. These ageing pathways are highly interconnected, involving localized and systemic increases in inflammatory mediators and damage associated molecular patterns (DAMPs); along with corresponding changes in immune cell function, metabolism and composition of the pulmonary and gut microbiomes. Here we comprehensively review the roles of ageing mechanisms in the tissue remodeling of lung disease.
- Published
- 2021
49. Previous Influenza Infection Exacerbates Allergen Specific Response and Impairs Airway Barrier Integrity in Pre-Sensitized Mice
- Author
-
Luke J. Berry, Stephen M. Stick, Kara L. Perks, Paul Rigby, Darryl A. Knight, Anthony Kicic, Alexander N. Larcombe, Kevin Looi, and Graeme R. Zosky
- Subjects
tight junctions ,Immunoglobulin E ,Mice ,Airway resistance ,Claudin-1 ,BALB/c mice ,Medicine ,Biology (General) ,house dust mite ,Methacholine Chloride ,Spectroscopy ,Sensitization ,Mice, Inbred BALB C ,biology ,medicine.diagnostic_test ,Pyroglyphidae ,General Medicine ,respiratory system ,Computer Science Applications ,Chemistry ,Treatment Outcome ,medicine.anatomical_structure ,Influenza A virus ,Female ,Bronchial Hyperreactivity ,medicine.symptom ,influenza ,medicine.drug ,QH301-705.5 ,Ovalbumin ,Down-Regulation ,Inflammation ,Article ,Catalysis ,Inorganic Chemistry ,Orthomyxoviridae Infections ,Animals ,Physical and Theoretical Chemistry ,QD1-999 ,Molecular Biology ,business.industry ,epithelial barrier integrity ,Airway Resistance ,Organic Chemistry ,lung function ,respiratory tract diseases ,Bronchoalveolar lavage ,Immunology ,biology.protein ,Respiratory epithelium ,Methacholine ,business - Abstract
In this study we assessed the effects of antigen exposure in mice pre-sensitized with allergen following viral infection on changes in lung function, cellular responses and tight junction expression. Female BALB/c mice were sensitized to ovalbumin and infected with influenza A before receiving a second ovalbumin sensitization and challenge with saline, ovalbumin (OVA) or house dust mite (HDM). Fifteen days post-infection, bronchoalveolar inflammation, serum antibodies, responsiveness to methacholine and barrier integrity were assessed. There was no effect of infection alone on bronchoalveolar lavage cellular inflammation 15 days post-infection, however, OVA or HDM challenge resulted in increased bronchoalveolar inflammation dominated by eosinophils/neutrophils or neutrophils, respectively. Previously infected mice had higher serum OVA-specific IgE compared with uninfected mice. Mice previously infected, sensitized and challenged with OVA were most responsive to methacholine with respect to airway resistance, while HDM challenge caused significant increases in both tissue damping and tissue elastance regardless of previous infection status. Previous influenza infection was associated with decreased claudin-1 expression in all groups and decreased occludin expression in OVA or HDM-challenged mice. This study demonstrates the importance of the respiratory epithelium in pre-sensitized individuals, where influenza-infection-induced barrier disruption resulted in increased systemic OVA sensitization and downstream effects on lung function.
- Published
- 2021
50. Long‐chain fatty acids are bad in <scp>IPF</scp> , or are they?
- Author
-
Darryl A. Knight and Bruce M. McManus
- Subjects
Pulmonary and Respiratory Medicine ,medicine.medical_specialty ,Idiopathic pulmonary fibrosis ,business.industry ,Internal medicine ,Fatty Acids ,medicine ,Humans ,business ,medicine.disease ,Long chain ,Gastroenterology - Published
- 2020
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