Biology subjects

Ehre, C.

Publications and source records attributed to Ehre, C..

8 recordsLinked to original sources

bioRxiv2025

Airway Epithelial SARS-CoV-2 Infectious and Repair Responses: Relationships to Age, Sex, and Post-COVID Pulmonary Syndromes

The long-term pulmonary sequelae of SARS-CoV-2 respiratory infections reflect infection severity, innate and adaptive immunity, and respiratory epithelial repair. This study investigated the acute and reparative responses as a function of age and sex in primary human bronchial epithelial (HBE) cultures utilizing a 14-day SARS-CoV-2 infection protocol. SARS-CoV-2 infection peaked at 3 days post-infection (dpi) with an [~] 2 log titer suppression at 14 dpi. SARS-CoV-2 infection induced interferon, interferon-induced gene, and cell damage responses. No age- or sex-dependent effects on SARS-CoV-2 infection were detected. Airway epithelia repaired to an abnormal mucus metaplastic/inflammatory state that reflected potentially beneficial and adverse consequences at 14 dpi. Repair processes were infection severity-dependent, not sex-dependent, and were more robust in young donor cultures. Analyses of long-COVID subjects with persistent pulmonary fibrosis or persistent bronchitic airway diseases exhibited expression of HBE 14 dpi failed repair gene signatures, including ISG gene signatures. Human airway epithelial repair post-SARS-CoV-2 is prolonged and incomplete in vitro over 14 days, and persistently abnormal repair may contribute to phenotypes of people with long-COVID pulmonary syndrome.

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↗

bioRxiv2024

Profiling endogenous airway proteases and antiproteases and measuring proteolytic activation of Influenza HA using in vitro and ex vivo human airway surface liquid samples

Imbalance of airway proteases and antiproteases has been implicated in diseases such as COPD and environmental exposures including cigarette smoke and ozone. To initiate infection, endogenous proteases are commandeered by respiratory viruses upon encountering the airway epithelium. The airway proteolytic environment likely contains redundant antiproteases and proteases with diverse catalytic mechanisms, however a proteomic profile of these enzymes and inhibitors in airway samples has not been reported. The objective of this study was to first profile extracellular proteases and antiproteases using human airway epithelial cell cultures and ex vivo nasal epithelial lining fluid (NELF) samples. Secondly, we present an optimized method for probing the proteolytic environment of airway surface liquid samples (in vitro and ex vivo) using fluorogenic peptides modeling the cleavage sites of respiratory viruses. We detected 48 proteases in the apical wash of cultured human nasal epithelial cells (HNECs) (n=6) and 57 in NELF (n=13) samples from healthy human subjects using mass-spectrometry based proteomics. Additionally, we detected 29 and 48 antiproteases in the HNEC apical washes and NELF, respectively. We observed large interindividual variability in rate of cleavage of an Influenza H1 peptide in the ex vivo clinical samples. Since protease and antiprotease levels have been found to be altered in the airways of smokers, we compared proteolytic cleavage in ex vivo nasal lavage samples from male/female smokers and non-smokers. There was a statistically significant increase in proteolysis of Influenza H1 in NLF from male smokers compared to female smokers. Furthermore, we measured cleavage of the S1/S2 site of SARS-CoV, SARS-CoV-2, and SARS-CoV-2 Delta peptides in various airway samples, suggesting the method could be used for other viruses of public health relevance. This assay presents a direct and efficient method of evaluating the proteolytic environment of human airway samples in assessment of therapeutic treatment, exposure, or underlying disease.

immunology↗

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↗

bioRxiv2020

Increased Colonic Expression of ACE2 Associates with Poor Prognosis in Crohn's disease

Background and AimsThe host receptor for SARS-CoV-2, angiotensin-converting enzyme 2 (ACE2), is highly expressed in small intestine. Our aim was to study colonic ACE2 expression in Crohns disease (CD) and non-inflammatory bowel disease (non-IBD) controls. We hypothesized that the colonic expression levels of ACE2 impacts CD course. MethodsWe examined the expression of colon ACE2 using RNA-seq and quantitative (q) RT-PCR from 69 adult CD and 14 NIBD control patients. In a subset of this cohort we validated ACE2 protein expression and localization in formalin-fixed, paraffin-embedded matched colon and ileal tissues using immunohistochemistry. The impact of increased ACE2 expression in CD for the risk of surgery was evaluated by a multivariate regression analysis and a Kaplan-Meier estimator. To provide critical support for the generality of our findings, we analyzed previously published RNA-seq data from two large independent cohorts of CD patients. ResultsColonic ACE2 expression was significantly higher in a subset of adult CD patients (ACE2-high CD). IHC in a sampling of ACE2-high CD patients confirmed high ACE2 protein expression in the colon and ileum compared to ACE2-low CD and NIBD patients. Notably, we found that ACE2-high CD patients are significantly more likely to undergo surgery within 5 years of diagnosis, with a Cox regression analysis finding that high ACE2 levels is an independent risk factor (OR 2.18; 95%CI, 1.05-4.55; p=0.037). ConclusionIncreased intestinal expression of ACE2 is associated with deteriorated clinical outcomes in CD patients. These data point to the need for molecular stratification that may impact CD disease-related outcomes.

genomics↗

bioRxiv2020

SARS-CoV-2 D614G Variant Exhibits Enhanced Replication ex vivo and Earlier Transmission in vivo

The D614G substitution in the S protein is most prevalent SARS-CoV-2 strain circulating globally, but its effects in viral pathogenesis and transmission remain unclear. We engineered SARS-CoV-2 variants harboring the D614G substitution with or without nanoluciferase. The D614G variant replicates more efficiency in primary human proximal airway epithelial cells and is more fit than wildtype (WT) virus in competition studies. With similar morphology to the WT virion, the D614G virus is also more sensitive to SARS-CoV-2 neutralizing antibodies. Infection of human ACE2 transgenic mice and Syrian hamsters with the WT or D614G viruses produced similar titers in respiratory tissue and pulmonary disease. However, the D614G variant exhibited significantly faster droplet transmission between hamsters than the WT virus, early after infection. Our study demonstrated the SARS-CoV2 D614G substitution enhances infectivity, replication fitness, and early transmission.

microbiology↗

bioRxiv2020

BPIFB1 loss alters airway mucus properties and diminishes mucociliary clearance.

Airway mucociliary clearance (MCC) is required for host defense and often diminished in chronic lung diseases. Effective clearance depends upon coordinated actions of the airway epithelium and a mobile mucus layer. Dysregulation of the primary secreted airway mucin proteins, MUC5B and MUC5AC, is associated with a reduction in the rate of MCC; however, how other secreted proteins impact the integrity of the mucus layer and MCC remains unclear. We previously identified the gene Bpifb1/Lplunc1 as a regulator of airway MUC5B levels using genetic approaches. Here, we show that BPIFB1 is required for normal mucociliary clearance in vivo using Bpifb1 knockout (KO) mice. Reduced MCC in Bpifb1 KO mice occurred in the absence of defects in sodium or chloride ion transport or reduced ciliary beat frequency. BPIFB1 loss resulted in airway mucus flakes with significantly increased complex viscosity, a key biophysical property of mucus known to impact MCC. Finally, we detected colocalization of BPIFB1 and MUC5B in secretory granules in mice and in the protein mesh of secreted mucus in human airway cultures. Collectively, our findings demonstrate that BPIFB1 is an important component of the mucociliary apparatus in mice and a key component of the mucus protein network.

physiology↗

bioRxiv2020

Potentiating TMEM16A channel function has no effect on airway goblet cells or bronchial and pulmonary vascular smooth muscle function

The calcium-activated chloride channel TMEM16A enables chloride secretion across several transporting epithelia, including in the airway where it represents a therapeutic target for the treatment of cystic fibrosis. Additional roles for TMEM16A have also been proposed, including enhancing goblet cell exocytosis, increasing goblet cell numbers and stimulating smooth muscle contraction. The aim of the present study was to test whether the pharmacological regulation of TMEM16A channel function, both potentiation and inhibition, could affect any of these proposed biological roles. In vitro, a recently described potent and selective TMEM16A potentiator (ETX001) failed to stimulate mucin release from primary human bronchial epithelial (HBE) cells over a 24h exposure period using both biochemical and imaging endpoints. In addition, treatment of HBE cells with ETX001 or a potent and selective TMEM16A inhibitor (Ani9) for 4 days did not influence mucin release or goblet cell formation. In vivo, a TMEM16A potentiator was without effect on goblet cell emptying in an IL-13 driven goblet cell metaplasia model. Using freshly isolated human bronchi and pulmonary arteries, neither ETX001 or Ani9 had any effect on the contractile or relaxant responses of the tissues. In vivo, ETX001 also failed to influence either lung or cardiovascular function when delivered directly into the airways of telemetered rats. Together, these studies do not support a role for TMEM16A in the regulation of goblet cell numbers or mucin release, or on the regulation of airway or pulmonary artery smooth muscle contraction.

physiology↗