Biology subjects

Kesimer, M.

Publications and source records attributed to Kesimer, M..

6 recordsLinked to original sources

bioRxiv2026

Region-specific molecular regulatory programs define epithelial identity, progenitor states, and mucus homeostasis in human distal airways

Small distal airways differ from proximal large airways in structure, airflow dynamics, and epithelial composition, and represent a central site of muco-obstructive lung disease pathogenesis. However, due in part to their inaccessibility, the molecular mechanisms that establish regional epithelial identity and govern mucociliary defense in distal airway epithelia remain poorly defined. Here, we integrate transcriptomic, secretomic, and chromatin accessibility analyses of matched primary human large and small airway epithelial cultures to define region-specific regulatory networks. We identify distal airway-specific transcriptional and chromatin programs required for maintaining epithelial identity and mucus homeostasis. Loss of NKX2-1 impairs distal airway secretory cell (DASC) differentiation and shifts mucus properties toward a disease-associated state. Lineage-resolved organoid assays identify an NKX2-1-high distal airway basal cell population with hybrid basal-secretory features as a selective progenitor for DASCs. Collectively, these findings establish a molecular framework for distal airway epithelial biology and define mechanisms regulating region-specific mucociliary host defense.

Cell Biology↗

bioRxiv2024

Dysregulated Airway Host Defense in Hyper IgE Syndrome due to STAT3 Mutations

RationaleHyper IgE syndrome (STAT3-HIES), also known as Jobs syndrome, is a rare immunodeficiency disease typically caused by dominant-negative STAT3 mutations. STAT3-HIES syndrome is characterized by chronic pulmonary infection and inflammation, suggesting impairment of pulmonary innate host defense. ObjectivesTo identify airway epithelial host defense defects consequent to STAT3 mutations that, in addition to reported mutant STAT3 immunologic abnormalities, produce pulmonary infection. MethodsSTAT3-HIES sputum was evaluated for biochemical/biophysical properties. STAT3-HIES excised lungs were harvested for histology; bronchial brush samples were collected for RNA sequencing and in vitro culture. A STAT3-HIES-specific mutation (R382W), expressed by lentiviruses, and a STAT3 knockout, generated by CRISPR/Cas9, were maintained in normal human bronchial epithelia under basal or inflammatory (IL1{beta}) conditions. Effects of STAT3 deficiency on transcriptomics, and epithelial ion channel, secretory, antimicrobial, and ciliary functions were assessed. Measurements and Main ResultsMucus concentrations and viscoelasticity were increased in STAT3-HIES sputum. STAT3-HIES excised lungs exhibited mucus obstruction and elevated IL1{beta} expression. STAT3 deficiency impaired CFTR-dependent fluid and mucin secretion, inhibited expression of antimicrobial peptides, cytokines, and chemokines, and acidified airway surface liquid at baseline and post-IL1{beta} exposure in vitro. Notably, mutant STAT3 suppressed IL1R1 expression. STAT3 mutations also inhibited ciliogenesis in vivo and impaired mucociliary transport in vitro, a process mediated via HES6 suppression. Administration of a {gamma}-secretase inhibitor increased HES6 expression and improved ciliogenesis in STAT3 R382W mutant cells. ConclusionsSTAT3 dysfunction leads to multi-component defects in airway epithelial innate defense, which, in conjunction with STAT3-HIES immune deficiency, contributes to chronic pulmonary infection.

cell biology↗

bioRxiv2023

Mucus polymer concentration and in vivo adaptation converge to define the antibiotic response of Pseudomonas aeruginosa during chronic lung infection

The airway milieu of individuals with muco-obstructive airway diseases (MADs) is defined by the accumulation of dehydrated mucus due to hyperabsorption of airway surface liquid and defective mucociliary clearance. Pathological mucus becomes progressively more viscous with age and disease severity due to the concentration and overproduction of mucin and accumulation of host-derived extracellular DNA (eDNA). Respiratory mucus of MADs provides a niche for recurrent and persistent colonization by respiratory pathogens, including Pseudomonas aeruginosa, which is responsible for the majority of morbidity and mortality in MADs. Despite high concentration inhaled antibiotic therapies and the absence of antibiotic resistance, antipseudomonal treatment failure in MADs remains a significant clinical challenge. Understanding the drivers of antibiotic recalcitrance is essential for developing more effective treatments that eradicate persistent infections. The complex and dynamic environment of diseased airways makes it difficult to model antibiotic efficacy in vitro. We aimed to understand how mucin and eDNA concentrations, the two dominant polymers in respiratory mucus, alter the antibiotic tolerance of P. aeruginosa. Our results demonstrate that polymer concentration and molecular weight affect P. aeruginosa survival post antibiotic challenge. Polymer-driven antibiotic tolerance was not explicitly associated with reduced antibiotic diffusion. Lastly, we established a robust and standardized in vitro model for recapitulating the ex vivo antibiotic tolerance of P. aeruginosa observed in expectorated sputum across age, underlying MAD etiology, and disease severity, which revealed the inherent variability in intrinsic antibiotic tolerance of host-evolved P. aeruginosa populations. ImportanceAntibiotic treatment failure in Pseudomonas aeruginosa chronic lung infections is associated with increased morbidity and mortality, illustrating the clinical challenge of bacterial infection control. Understanding the underlying infection environment, as well as the host and bacterial factors driving antibiotic tolerance and the ability to accurately recapitulate these factors in vitro, is crucial for improving antibiotic treatment outcomes. Here, we demonstrate that increasing concentration and molecular weight of mucin and host eDNA drive increased antibiotic tolerance to tobramycin. Through systematic testing and modeling, we identified a biologically relevant in vitro condition that recapitulates antibiotic tolerance observed in ex vivo treated sputum. Ultimately, this study revealed a dominant effect of in vivo evolved bacterial populations in defining inter-subject ex vivo antibiotic tolerance and establishes a robust and translatable in vitro model for therapeutic development.

microbiology↗

bioRxiv2022

Genetic Regulators of Sputum Mucin Concentration and Their Associations with COPD Phenotypes

Hyper-secretion and/or hyper-concentration of mucus is a defining feature of multiple obstructive lung diseases, including chronic obstructive pulmonary disease (COPD). Mucus itself is composed of a mixture of water, ions, salt and proteins, of which the gel-forming mucins, MUC5AC and MUC5B, are the most abundant. Recent studies have linked the concentrations of these proteins in sputum to COPD phenotypes, including chronic bronchitis (CB) and acute exacerbations (AE). We sought to determine whether common genetic variants influence sputum mucin concentrations and whether these variants are also associated with COPD phenotypes, specifically CB and AE. We performed a GWAS to identify quantitative trait loci for sputum mucin protein concentration (pQTL) in the Sub-Populations and InteRmediate Outcome Measures in COPD Study (SPIROMICS, n=708 for total mucin, n=215 for MUC5AC, MUC5B). Subsequently, we tested for associations of mucin pQTL with CB and AE using regression modeling (n=822-1300). Replication analysis was conducted using data from COPDGene (n =5740) and by examining results from the UK Biobank. We identified one genome-wide significant pQTL for MUC5AC (rs75401036) and two for MUC5B (rs140324259, rs10001928). The strongest association for MUC5B, with rs140324259 on chromosome 11, explained 14% of variation in sputum MUC5B. Despite being associated with lower MUC5B, the C allele of rs140324259 conferred increased risk of CB (odds ratio (OR) = 1.42; 95% confidence interval (CI): 1.10-1.80) as well as AE ascertained over three years of follow up (OR=1.41; 95% CI: 1.02-1.94). Associations between rs140324259 and CB or AE did not replicate in COPDGene. However, in the UK Biobank, rs140324259 was associated with phenotypes that define CB, namely chronic mucus production and cough, again with the C allele conferring increased risk. We conclude that sputum MUC5AC and MUC5B concentrations are associated with common genetic variants, and the top locus for MUC5B may influence COPD phenotypes, in particular CB. Author SummaryChronic obstructive pulmonary disease (COPD) is characterized by presence of emphysema and/or chronic bronchitis. Excessive mucus production is a defining phenotype of chronic bronchitis, and is associated with several important features of COPD, including exacerbations and loss of lung function. Recent studies have demonstrated that the amount of mucus produced in COPD patients is an important marker of disease state. We investigated whether common genetic variants are associated with the concentration of two key proteins in mucus, MUC5AC and MUC5B, and whether the variants we identified are also associated with COPD outcomes. We identified multiple genetic variants that were associated with MUC5AC or MUC5B concentration. The strongest association we detected, for MUC5B on chromosome 11, was also associated with features of COPD, including chronic bronchitis and acute exacerbations, in one COPD study population but not another. Results from a much larger study, the UK Biobank, indicate that this variant is associated with chronic mucus production and chronic cough, which are key features of chronic bronchitis. Thus, we conclude that the concentration of key proteins in mucus are influenced by genetic variation, and that a variant on chromosome 11 that affects MUC5B may in turn alter COPD outcomes.

genetics↗

bioRxiv2021

Hyperconcentrated Mucus Unifies Submucosal Gland and Superficial Airway Dysfunction in Cystic Fibrosis

Cystic fibrosis (CF) is characterized by abnormal transepithelial ion transport. However, a description of CF lung disease pathophysiology unifying superficial epithelial and submucosal gland (SMG) dysfunctions has remained elusive. We hypothesized that biophysical abnormalities associated with CF mucus hyperconcentration provide a unifying mechanism. Studies of the anion secretion-inhibited pig airway CF model revealed elevated SMG mucus concentrations, osmotic pressures, and SMG mucus accumulation. Human airway studies revealed hyperconcentrated CF SMG mucus with raised osmotic pressures and cohesive forces predicted to limit SMG mucus secretion/release. Utilizing proline-rich protein 4 (PRR4) as a biomarker of SMG secretion, proteomics analyses of CF sputum revealed markedly lower PRR4 levels compared to healthy and bronchiectasis controls, consistent with a failure of CF SMGs to secrete mucus onto airway surfaces. Raised mucus osmotic/cohesive forces, reflecting mucus hyperconcentration, provide a unifying mechanism that describes disease-initiating mucus accumulation on airway surfaces and within SMGs of the CF lung.

pathology↗

bioRxiv2021

Membrane-Tethered Mucin 1 is Stimulated by Interferon in Multiple Cell Types and Antagonizes Influenza A Virus Infection in Human Airway Epithelium

Influenza A virus (IAV) causes seasonal epidemics and periodic pandemics, resulting in significant morbidity and mortality in the human population. Tethered mucin 1 (MUC1) is highly expressed in airway epithelium, the primary site of IAV replication, and also by other cell types that influence IAV infection, including macrophages. MUC1 has the potential to influence infection dynamics through physical interactions and/or signaling activity, and recent work suggests MUC1 acts as a releasable decoy receptor and anti-inflammatory molecule during IAV infection. Still, the modulation of MUC1 and its impact during viral pathogenesis remains unclear. Thus, we sought to further investigate the interplay between MUC1 and IAV in an in vitro model of primary human airway epithelium (HAE). Our data indicate that a recombinant IAV hemagglutinin (H3) and H3N2 virus can bind endogenous HAE MUC1. We find that infection of HAE cultures with H1N1 or H3N2 IAV strains does not trigger enhanced MUC1 shedding, but instead stimulates an increase in cell-associated MUC1 protein. We observed a similar increase after stimulation with either type I or type III interferon (IFN); however, inhibition of IFN signaling during H1N1 infection only partially abrogated this increase, indicating multiple soluble factors contribute to MUC1 upregulation during the antiviral response. We expanded these findings and demonstrate that in addition to HAE, primary human monocyte-derived macrophages also upregulate MUC1 protein in response to both IFN treatment and conditioned media from IAV-infected HAE cultures. We then developed HAE genetically depleted for MUC1 to determine its impact on IAV pathogenesis, finding that MUC1 knock-out cultures exhibited enhanced viral growth compared to control cultures for several IAV strains. Together, our data support a model whereby MUC1 antagonizes productive uptake of IAV in HAE. Infection then stimulates MUC1 expression on multiple cell types through IFN-dependent and -independent mechanisms that may further impact infection dynamics. Author SummaryThe mucosal surface of the respiratory epithelium is an important site of first contact for viral respiratory pathogens. Large and heavily glycosylated molecules known as tethered mucins extend from the cell surface and may physically restrict access to underlying cells. Recently, one of these tethered mucins, MUC1, has also been shown to influence cell signaling and inflammation. Still, despite its abundance in the airway and multifunctional capability, the role of MUC1 during influenza virus infection in the human respiratory tract remains unclear. Here, we demonstrate that influenza virus directly interacts with MUC1 in a physiologically-relevant model of human airway epithelium and find that MUC1 protein expression is elevated throughout the epithelium and in primary human monocyte-derived macrophages in response to important antiviral signals produced during infection. Using genetically-modified human airway cultures lacking MUC1, we then provide evidence of more efficient influenza virus infection in the absence of this mucin. Our data suggest that MUC1 not only physically restricts influenza virus uptake, but also represents a dynamic component of the host response that acts to further stem viral spread.

microbiology↗