ABSTRACTIntroductionRecurrent wheezing disorders including asthma are complex and heterogeneous d... more ABSTRACTIntroductionRecurrent wheezing disorders including asthma are complex and heterogeneous diseases that affect up to 30% of all children, contributing to a major burden on children, their families, and global healthcare systems. It is now recognized that a dysfunctional airway epithelium plays a central role in the pathogenesis of recurrent wheeze, although the underlying mechanisms are still not fully understood. This prospective birth cohort aims to bridge this knowledge gap by investigating the influence of intrinsic epithelial dysfunction on the risk for developing respiratory disorders and the modulation of this risk by maternal morbidities,in uteroexposures, and respiratory exposures in the first year of life.Methods and AnalysisThe Airway Epithelium Respiratory Illnesses and Allergy (AERIAL) study is nested within the ORIGINS Project and will monitor 400 infants from birth to five years. The primary outcome of the AERIAL study will be the identification of epithelial en...
Introduction: Junctional protein complexes such as tight junctions (TJ) provide a physical barrie... more Introduction: Junctional protein complexes such as tight junctions (TJ) provide a physical barrier against external insults. Although various aspects of TJs have been extensively assessed, few studies have directly addressed whether these are intrinsically different in asthmatic epithelium and the consequence on barrier function of disassembly post- human rhinovirus (HRV) infection. This study aimed to assess basal TJ expression and barrier function prior to and post-HRV infection in both monolayer and air-liquid interface (ALI) cultures of healthy and asthmatic epithelium. Methods: Primary airway epithelial cells from healthy and asthmatic children obtained via bronchial brushings and cultured as previously described (Kicic et al. 2006) were also differentiated into ALI and subsequently infected with HRV-1B over 24h. Tight junction gene expression was assessed via qPCR, barrier integrity measured by TJ protein expression of claudin-1, occludin and zonula occluden-1 (ZO-1) via immun...
s Urinary Biomarkers In Patients With Cystic Fibrosis Correlate With FEV1 & Lung Microbiome Diver... more s Urinary Biomarkers In Patients With Cystic Fibrosis Correlate With FEV1 & Lung Microbiome Diversity Emilie Vallieres, M.D. MBW & MRI Differentiate Disease Severity In Stable CF Lung Disease & Detect Response to Therapy in Children & Adolescents Mirjam Stahl Personalized Treatment For CF-Patients With Extremely Rare Mutations Based On The Intestinal Organoid Model Gitte Berkers, M.D. Safety & Efficacy of Treatment With Lumacaftor in Combination With Ivacaftor In Patients With CF Homozygous For F508del-CFTR Michael Konstan, M.D. Correlation Of Sweat Chloride & Percent Predicted FEV1 In Cystic Fibrosis Patients Treated With Ivacaftor Fred Van Goor, Ph.D. Qr-010 Treatment For Cystic Fibrosis: Assessing The Airway-Mucus Barrier In Delivery Vera Brinks Correlation Between Sweat Chloride Concentration & Lung Function In CF Patients Patrick Sosnay, M.D. End-Expiratory Lung Volumes On CT Are Not Elevated in The Presence Of "Gas Trapping" in Young Children With CF Tim Rosenow, B.S...
The airway epithelium of children with asthma is characterized by aberrant repair that may be the... more The airway epithelium of children with asthma is characterized by aberrant repair that may be therapeutically modifiable. The development of epithelial-targeting therapeutics that enhance airway repair could provide a novel treatment avenue for childhood asthma. Drug discovery efforts utilizing high-throughput live cell imaging of patient-derived airway epithelial culture-based wound repair assays can be used to identify compounds that modulate airway repair in childhood asthma. Manual cell tracking has been used to determine cell trajectories and wound closure rates, but is time consuming, subject to bias, and infeasible for high-throughput experiments. We therefore developed software, EPIC, that automatically tracks low-resolution low-framerate cells using artificial intelligence, analyzes high-throughput drug screening experiments and produces multiple wound repair metrics and publication-ready figures. Additionally, unlike available cell trackers that perform cell segmentation, ...
A significant proportion of chronic obstructive pulmonary disease exacerbations are strongly asso... more A significant proportion of chronic obstructive pulmonary disease exacerbations are strongly associated with rhinovirus infection (HRV). In this study, we combined long-term cigarette smoke exposure with HRV infection in a mouse model. Our aim was to better understand the effects of HRV infection on such exacerbations, using a realistic method for generating a COPD-like phenotype. After 12-weeks of cigarette smoke exposure, adult female BALB/c mice were infected with HRV-1A and three days later we assessed a range of outcomes including lung volume and function, collected lung tissue for measurement of viral titre, bronchoalveolar lavage for assessment of pulmonary inflammation and levels of key mediators, and fixed lungs for stereological structural analyses. Cigarette smoke exposure alone significantly increased total cells and macrophages, and reduced MIP-2 in bronchoalveolar lavage. HRV infection alone increased neutrophilic inflammation, IP-10 and total protein in lavage and also increased specific airway resistance measured at functional residual capacity. Cigarette smoke and HRV together impacted various lung structural parameters including increasing stereological lung volume. Our results show that long-term cigarette smoke exposure and HRV-1A infection both individually impact respiratory outcomes and combine to alter aspects of lung structure in a mouse model, thus providing insight into the development of future mechanistic studies and appropriate interventions in human disease.
The airway epithelium of children with wheeze is characterized by defective repair that contribut... more 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 altere...
Antimicrobial resistance is a global health crisis, partly contributed by inappropriate use of an... more Antimicrobial resistance is a global health crisis, partly contributed by inappropriate use of antibiotics. The increasing emergence of multidrug resistant infections has led to the resurgent interest in bacteriophages as an alternative treatment. Current procedures assessing susceptibility and breadth of host range to bacteriophage are conducted using large-scale manual processes that are labor-intensive. The aim here was to establish and validate a scaled down methodology for high-throughput screening in order to reduce procedural footprint. Bacteriophages were isolated from wastewater samples and screened for specificity against 29 clinical Pseudomonas aeruginosa isolates and PA01 using a spot test (2 μL/ drop). Host range assessment was performed on four representative P. aeruginosa isolates using both double agar overlay assay on petri dishes and 24-well culture plates. The breadth of host range of bacteriophages that exhibited lytic activity on P. aeruginosa isolates were corr...
Neutrophil elastase (NE) is the most significant predictor of bronchiectasis in early life cystic... more Neutrophil elastase (NE) is the most significant predictor of bronchiectasis in early life cystic fibrosis (CF), however, the causal link between NE and airway damage is not well understood. Matrix metalloproteinases (MMPs) play a crucial role in extracellular matrix modelling and are activated by NE. The aim of this study was to assess if MMP activation positively correlates with NE activity, disease severity and bronchiectasis in young children with CF. Total MM-1, MMP-2, MMP-7, MMP-9 and TIMP-2, TIMP-1 levels were measured in bronchoalveolar lavage fluid, collected from young children with CF during annual clinical assessment. Active/pro-enzyme ratio of MMP-9 was determined by gelatin zymography. Annual chest computed tomography imaging was scored for bronchiectasis. A higher MMP-9/TIMP-1 ratio was associated with free NE activity. In contrast, MMP2/TIMP-2 ratio decreased and MMP-1 and MMP-7 were not detected in the majority of samples. Ratio of active/pro-enzyme MMP-9 was also h...
ABSTRACTIntroductionRecurrent wheezing disorders including asthma are complex and heterogeneous d... more ABSTRACTIntroductionRecurrent wheezing disorders including asthma are complex and heterogeneous diseases that affect up to 30% of all children, contributing to a major burden on children, their families, and global healthcare systems. It is now recognized that a dysfunctional airway epithelium plays a central role in the pathogenesis of recurrent wheeze, although the underlying mechanisms are still not fully understood. This prospective birth cohort aims to bridge this knowledge gap by investigating the influence of intrinsic epithelial dysfunction on the risk for developing respiratory disorders and the modulation of this risk by maternal morbidities,in uteroexposures, and respiratory exposures in the first year of life.Methods and AnalysisThe Airway Epithelium Respiratory Illnesses and Allergy (AERIAL) study is nested within the ORIGINS Project and will monitor 400 infants from birth to five years. The primary outcome of the AERIAL study will be the identification of epithelial en...
Introduction: Junctional protein complexes such as tight junctions (TJ) provide a physical barrie... more Introduction: Junctional protein complexes such as tight junctions (TJ) provide a physical barrier against external insults. Although various aspects of TJs have been extensively assessed, few studies have directly addressed whether these are intrinsically different in asthmatic epithelium and the consequence on barrier function of disassembly post- human rhinovirus (HRV) infection. This study aimed to assess basal TJ expression and barrier function prior to and post-HRV infection in both monolayer and air-liquid interface (ALI) cultures of healthy and asthmatic epithelium. Methods: Primary airway epithelial cells from healthy and asthmatic children obtained via bronchial brushings and cultured as previously described (Kicic et al. 2006) were also differentiated into ALI and subsequently infected with HRV-1B over 24h. Tight junction gene expression was assessed via qPCR, barrier integrity measured by TJ protein expression of claudin-1, occludin and zonula occluden-1 (ZO-1) via immun...
s Urinary Biomarkers In Patients With Cystic Fibrosis Correlate With FEV1 & Lung Microbiome Diver... more s Urinary Biomarkers In Patients With Cystic Fibrosis Correlate With FEV1 & Lung Microbiome Diversity Emilie Vallieres, M.D. MBW & MRI Differentiate Disease Severity In Stable CF Lung Disease & Detect Response to Therapy in Children & Adolescents Mirjam Stahl Personalized Treatment For CF-Patients With Extremely Rare Mutations Based On The Intestinal Organoid Model Gitte Berkers, M.D. Safety & Efficacy of Treatment With Lumacaftor in Combination With Ivacaftor In Patients With CF Homozygous For F508del-CFTR Michael Konstan, M.D. Correlation Of Sweat Chloride & Percent Predicted FEV1 In Cystic Fibrosis Patients Treated With Ivacaftor Fred Van Goor, Ph.D. Qr-010 Treatment For Cystic Fibrosis: Assessing The Airway-Mucus Barrier In Delivery Vera Brinks Correlation Between Sweat Chloride Concentration & Lung Function In CF Patients Patrick Sosnay, M.D. End-Expiratory Lung Volumes On CT Are Not Elevated in The Presence Of "Gas Trapping" in Young Children With CF Tim Rosenow, B.S...
The airway epithelium of children with asthma is characterized by aberrant repair that may be the... more The airway epithelium of children with asthma is characterized by aberrant repair that may be therapeutically modifiable. The development of epithelial-targeting therapeutics that enhance airway repair could provide a novel treatment avenue for childhood asthma. Drug discovery efforts utilizing high-throughput live cell imaging of patient-derived airway epithelial culture-based wound repair assays can be used to identify compounds that modulate airway repair in childhood asthma. Manual cell tracking has been used to determine cell trajectories and wound closure rates, but is time consuming, subject to bias, and infeasible for high-throughput experiments. We therefore developed software, EPIC, that automatically tracks low-resolution low-framerate cells using artificial intelligence, analyzes high-throughput drug screening experiments and produces multiple wound repair metrics and publication-ready figures. Additionally, unlike available cell trackers that perform cell segmentation, ...
A significant proportion of chronic obstructive pulmonary disease exacerbations are strongly asso... more A significant proportion of chronic obstructive pulmonary disease exacerbations are strongly associated with rhinovirus infection (HRV). In this study, we combined long-term cigarette smoke exposure with HRV infection in a mouse model. Our aim was to better understand the effects of HRV infection on such exacerbations, using a realistic method for generating a COPD-like phenotype. After 12-weeks of cigarette smoke exposure, adult female BALB/c mice were infected with HRV-1A and three days later we assessed a range of outcomes including lung volume and function, collected lung tissue for measurement of viral titre, bronchoalveolar lavage for assessment of pulmonary inflammation and levels of key mediators, and fixed lungs for stereological structural analyses. Cigarette smoke exposure alone significantly increased total cells and macrophages, and reduced MIP-2 in bronchoalveolar lavage. HRV infection alone increased neutrophilic inflammation, IP-10 and total protein in lavage and also increased specific airway resistance measured at functional residual capacity. Cigarette smoke and HRV together impacted various lung structural parameters including increasing stereological lung volume. Our results show that long-term cigarette smoke exposure and HRV-1A infection both individually impact respiratory outcomes and combine to alter aspects of lung structure in a mouse model, thus providing insight into the development of future mechanistic studies and appropriate interventions in human disease.
The airway epithelium of children with wheeze is characterized by defective repair that contribut... more 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 altere...
Antimicrobial resistance is a global health crisis, partly contributed by inappropriate use of an... more Antimicrobial resistance is a global health crisis, partly contributed by inappropriate use of antibiotics. The increasing emergence of multidrug resistant infections has led to the resurgent interest in bacteriophages as an alternative treatment. Current procedures assessing susceptibility and breadth of host range to bacteriophage are conducted using large-scale manual processes that are labor-intensive. The aim here was to establish and validate a scaled down methodology for high-throughput screening in order to reduce procedural footprint. Bacteriophages were isolated from wastewater samples and screened for specificity against 29 clinical Pseudomonas aeruginosa isolates and PA01 using a spot test (2 μL/ drop). Host range assessment was performed on four representative P. aeruginosa isolates using both double agar overlay assay on petri dishes and 24-well culture plates. The breadth of host range of bacteriophages that exhibited lytic activity on P. aeruginosa isolates were corr...
Neutrophil elastase (NE) is the most significant predictor of bronchiectasis in early life cystic... more Neutrophil elastase (NE) is the most significant predictor of bronchiectasis in early life cystic fibrosis (CF), however, the causal link between NE and airway damage is not well understood. Matrix metalloproteinases (MMPs) play a crucial role in extracellular matrix modelling and are activated by NE. The aim of this study was to assess if MMP activation positively correlates with NE activity, disease severity and bronchiectasis in young children with CF. Total MM-1, MMP-2, MMP-7, MMP-9 and TIMP-2, TIMP-1 levels were measured in bronchoalveolar lavage fluid, collected from young children with CF during annual clinical assessment. Active/pro-enzyme ratio of MMP-9 was determined by gelatin zymography. Annual chest computed tomography imaging was scored for bronchiectasis. A higher MMP-9/TIMP-1 ratio was associated with free NE activity. In contrast, MMP2/TIMP-2 ratio decreased and MMP-1 and MMP-7 were not detected in the majority of samples. Ratio of active/pro-enzyme MMP-9 was also h...
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