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The Science of the Total Environment Jun 2024Crystalline silica (CS) particles are ubiquitously present in the environment, particularly in occupational settings, and exposure to respirable CS causes silicosis,...
Crystalline silica (CS) particles are ubiquitously present in the environment, particularly in occupational settings, and exposure to respirable CS causes silicosis, imposing a significant disease burden. However, the pathogenesis of silicosis remains unclear. Exposure to external stimuli, such as CS, leads to the accumulation of unfolded proteins and triggers endoplasmic reticulum (ER) stress, disrupting tissue immune homeostasis and accelerating pathological progression. While pulmonary macrophages phagocytose CS particles to initiate the immune response, the role of ER stress in this process is unknown. Herein, we used a murine model of silicosis to simulate the pathological progression from acute inflammation to fibrosis in silicosis and conducted in vivo pharmacological inhibition of ER stress to explore the underlying mechanism. Using flow cytometry, we further classified pulmonary macrophages into monocyte-like macrophages (monocytes), interstitial macrophages (IMs), and alveolar macrophages (AMs). Our results showed that CS-induced ER stress primarily contributed to the augmentation of IMs and thereby exerted a significant impact on pulmonary macrophages. Despite coexpressing M1- and M2-like markers, IMs predominantly exhibited an M1-like polarization state and played a proinflammatory role by expressing the cytokines pro-IL-1β and TNF-α during the pathological progression of silicosis. Additionally, IMs recruited by CS-induced ER stress also exhibited high expression of MHCII and exerted active immunomodulatory effects. Overall, our study demonstrates that ER stress induced by CS particles triggers a proinflammatory immune microenvironment dominated by IMs and reveals novel insights into the pulmonary toxicological effects of CS particles.
PubMed: 38936737
DOI: 10.1016/j.scitotenv.2024.174299 -
Molecular Immunology Jun 2024Immune cells in the human lung are associated with idiopathic pulmonary fibrosis. However, the contribution of different immune cell subpopulations to the pathogenesis...
Immune cells in the human lung are associated with idiopathic pulmonary fibrosis. However, the contribution of different immune cell subpopulations to the pathogenesis of pulmonary fibrosis remains unclear. We used single-cell RNA sequencing data to investigate the transcriptional profiles of immune cells in the lungs of 5 IPF patients and 3 subjects with non-fibrotic lungs. In an identifiable population of immune cells, we found increased percentage of CD8 T cells in the T cell subpopulation in IPF. Monocle analyzed the dynamic immune status and cell transformation of CD8 T cells, as well as the cytotoxicity and exhausted status of CD8 T cell subpopulations at different stages. Among CD8 T cells, we found differences in metabolic pathways in IPF and Ctrl, including lipid, amino acid and carbohydrate metabolic. By analyzing the metabolites of CD8 T cells, we found that different populations of CD8 T cells in IPF have unique metabolic characteristics, but they also have multiple identical up-regulated or down-regulated metabolites. In IPF, signaling pathways associated with fibrosis were enriched in CD8 T cells, suggesting that CD8 T cells may have an important contribution to fibrosis. Finally, we analyzed the interactions between CD8 T cells and other cells. Together, these studies highlight key features of CD8 T cells in the pathogenesis of IPF and help to develop effective therapeutic targets.
PubMed: 38936318
DOI: 10.1016/j.molimm.2024.06.008 -
Assessment of Global and Regional Lung Compliance in Pulmonary Fibrosis With Hyperpolarized Gas MRI.Journal of Magnetic Resonance Imaging :... Jun 2024Lung compliance, a biomarker of pulmonary fibrosis, is generally measured globally. Hyperpolarized Xe gas MRI offers the potential to evaluate lung compliance...
BACKGROUND
Lung compliance, a biomarker of pulmonary fibrosis, is generally measured globally. Hyperpolarized Xe gas MRI offers the potential to evaluate lung compliance regionally, allowing for visualization of changes in lung compliance associated with fibrosis.
PURPOSE
To assess global and regional lung compliance in a rat model of pulmonary fibrosis using hyperpolarized Xe gas MRI.
STUDY TYPE
Prospective.
ANIMAL MODEL
Twenty Sprague-Dawley male rats with bleomycin-induced fibrosis model (N = 10) and saline-treated controls (N = 10).
FIELD STRENGTH/SEQUENCE
7-T, fast low-angle shot (FLASH) sequence.
ASSESSMENT
Lung compliance was determined by fitting lung volumes derived from segmented Xe MRI with an iterative selection method, to corresponding airway pressures. Similarly, lung compliance was obtained with computed tomography for cross-validation. Direction-dependencies of lung compliance were characterized by regional lung compliance ratios (R) in different directions. Pulmonary function tests (PFTs) and histological analysis were used to validate the pulmonary fibrosis model and assess its correlation with Xe lung compliance.
STATISTICAL TESTS
Shapiro-Wilk tests, unpaired and paired t-tests, Mann-Whitney U and Wilcoxon signed-rank tests, and Pearson correlation coefficients. P < 0.05 was considered statistically significant.
RESULTS
For the entire lung, the global and regional lung compliance measured with Xe gas MRI showed significant differences between the groups, and correlated with the global lung compliance measured using PFTs (global: r = 0.891; regional: r = 0.873). Additionally, for the control group, significant difference was found in mean regional compliance between areas, eg, 0.37 (0.32, 0.39) × 10 mL/cm HO and 0.47 (0.41, 0.56) × 10 mL/cm HO for apical and basal lung, respectively. The apical-basal direction R was 1.12 ± 0.09 and 1.35 ± 0.13 for fibrosis and control groups, respectively, indicating a significant difference.
DATA CONCLUSION
Our findings demonstrate the feasibility of using hyperpolarized gas MRI to assess regional lung compliance.
EVIDENCE LEVEL
2 TECHNICAL EFFICACY: Stage 1.
PubMed: 38935670
DOI: 10.1002/jmri.29497 -
American Journal of Respiratory and... Jun 2024
PubMed: 38935633
DOI: 10.1164/rccm.202405-1075LE -
Therapeutic Drug Monitoring Jun 2024The highly effective Cystic Fibrosis Transmembrane conductance Regulator (CFTR) modulator, elexacaftor-tezacaftor-ivacaftor, is now widely being used by people with...
Dried Blood Spot Method Development and Clinical Validation for the Analysis of Elexacaftor, Elexacaftor-M23, Tezacaftor, Tezacaftor-M1, Ivacaftor, Ivacaftor Carboxylate, and Hydroxymethyl Ivacaftor Using LC-MS/MS.
BACKGROUND
The highly effective Cystic Fibrosis Transmembrane conductance Regulator (CFTR) modulator, elexacaftor-tezacaftor-ivacaftor, is now widely being used by people with cystic fibrosis. However, few independent studies have detailed the pharmacokinetics (PK) of CFTR modulators. Blood collection by venipuncture is the gold standard for PK measurements, but it is invasive. The aim of this study was to develop and clinically validate a quantification method for elexacaftor, tezacaftor, ivacaftor, and their main metabolites in dried blood spots (DBSs) using liquid chromatography with tandem mass spectrometry.
METHODS
Linearity, accuracy, precision, stability, hematocrit (Hct), spot-to-spot carryover, spot volume, and extraction efficiency were validated in DBS for all analytes. The clinical validation of elexacaftor-tezacaftor-ivacaftor in patients was performed by comparing 21 DBS samples with matched plasma samples.
RESULTS
The preset requirements for linearity, within-run and between-run accuracy, precision, Hct, spot volume, and extraction efficiency were met. Puncher carryover was observed and resolved by punching 3 blanks after each sample. The samples remained stable and showed no notable degradation across the tested temperatures and time intervals. Corrected DBS values with the Passing-Bablok regression equation showed good agreement in Bland-Altman plots, and acceptance values were within 20% of the mean for a minimum of 67% of the repeats, according to the EMA guidelines.
CONCLUSIONS
A quantification method for the analysis of elexacaftor, tezacaftor, ivacaftor, and their main metabolites was developed and clinically validated in DBS. This method could be valuable in both clinical care and research to address unanswered PK questions regarding CFTR modulators.
PubMed: 38935410
DOI: 10.1097/FTD.0000000000001231 -
Lung Jun 2024This study aimed to evaluate the hypothesis that active smoking impacts upon mediators and abundance of circulating fibrocyte cells in smoking-related disease...
PURPOSE
This study aimed to evaluate the hypothesis that active smoking impacts upon mediators and abundance of circulating fibrocyte cells in smoking-related disease characterised by fibrosis.
METHODS
Flow cytometry and enzyme-linked immunosorbent assays were used to investigate blood from five patient groups: healthy never-smokers, healthy current smokers, stable chronic obstructive pulmonary disease (COPD) active smokers, idiopathic pulmonary fibrosis (IPF) never-smokers, and IPF active smokers.
RESULTS
A significant inverse dose-response relationship was observed in healthy smokers among cumulative smoking burden (pack-years) and fibrocyte abundance (p = 0.006, r = -0.86). Among serum profibrotic fibrocyte chemokines measured, CCL18 rose significantly alongside fibrocyte numbers in all five subject groups, while having an inverse dose-response relationship with pack-year burden in healthy smokers (p = 0.003, r = -0.89). In IPF, CCL2 rose in direct proportion to fibrocyte abundance irrespective of smoking status but had lower serum levels in those currently smoking (p = < 0.001). For the study population, CXCL12 was decreased in pooled current smokers versus never-smokers (p = 0.03).
CONCLUSION
The suppressive effect of current, as distinct from former, chronic smoking on circulating fibrocyte abundance in healthy smokers, and modulation of regulatory chemokine levels by active smoking may have implications for future studies of fibrocytes in smoking-related lung diseases as a potential confounding variable.
PubMed: 38935158
DOI: 10.1007/s00408-024-00720-3 -
Naunyn-Schmiedeberg's Archives of... Jun 2024The dreaded nosocomial pathogen Clostridioides difficile causes diarrhea and severe inflammation of the colon, especially after the use of certain antibiotics. The...
The dreaded nosocomial pathogen Clostridioides difficile causes diarrhea and severe inflammation of the colon, especially after the use of certain antibiotics. The bacterium releases two deleterious toxins, TcdA and TcdB, into the gut, which are mainly responsible for the symptoms of C. difficile-associated diseases (CDADs). Both toxins are capable of entering independently into various host cells, e.g., intestinal epithelial cells, where they mono-O-glucosylate and inactivate Rho and/or Ras GTPases, important molecular switches for various cellular functions. We have shown recently that the cellular uptake of the Clostridioides difficile toxins TcdA and TcdB (TcdA/B) is inhibited by the licensed class III antiarrhythmic drug amiodarone (Schumacher et al. in Gut Microbes 15(2):2256695, 2023). Mechanistically, amiodarone delays the cellular uptake of both toxins into target cells most likely by lowering membrane cholesterol levels and by interfering with membrane insertion and/or pore formation of TcdA/B. However, serious side effects, such as thyroid dysfunction and severe pulmonary fibrosis, limit the clinical use of amiodarone in patients with C. difficile infection (CDI). For that reason, we aimed to test whether dronedarone, an amiodarone derivative with a more favorable side effect profile, is also capable of inhibiting TcdA/B. To this end, we tested in vitro with various methods the impact of dronedarone on the intoxication of Vero and CaCo-2 cells with TcdA/B. Importantly, preincubation of both cell lines with dronedarone for 1 h at concentrations in the low micromolar range rendered the cells less sensitive toward TcdA/B-induced Rac1 glucosylation, collapse of the actin cytoskeleton, cell rounding, and cytopathic effects, respectively. Our study points toward the possibility of repurposing the already approved drug dronedarone as the preferable safer-to-use alternative to amiodarone for inhibiting TcdA/B in the (supportive) therapy of CDADs.
PubMed: 38935126
DOI: 10.1007/s00210-024-03248-8 -
Pediatric Pulmonology Jun 2024A primary palliative care model for cystic fibrosis (CF) recommends using the Integrated Palliative Care Outcome Scale (IPOS) for screening. Validation of the IPOS is...
BACKGROUND
A primary palliative care model for cystic fibrosis (CF) recommends using the Integrated Palliative Care Outcome Scale (IPOS) for screening. Validation of the IPOS is needed.
METHODS
This secondary analysis utilized baseline data from a multisite trial of the palliative care model, Improving Life with CF. Adults with CF completed the IPOS, the Memorial Symptom Assessment Scale-CF (MSAS-CF), the CF Questionnaire-Revised (CFQ-R), the Patient Health Questionnaire (PHQ-8), the Generalized Anxiety Disorder (GAD-7), and the Perceived Stress Scale (PSS). IPOS structure was assessed using Cronbach α coefficients and a factor analysis. Construct validity was evaluated through bivariate relationships between IPOS scores and other questionnaire scores, and linear regressions assessing the extent to which the IPOS explains variance in quality-of-life domains.
RESULTS
The sample comprised 256 adults with complete IPOS data. α coefficients were .86 for the IPOS total score, .81 for the Physical Symptoms subscale, .79 for the Emotional Symptoms subscale, and .63 for the Communication/Practical Issues subscale. A two-component factor structure best aligned with the current subscales. IPOS scores were significantly associated with other measures; associations with MSAS-CF and CFQ-R subscales differentiated the IPOS Physical and Emotional subscales. The IPOS total score provided unique information about the variance in the CFQ-R Physical Functioning and Respiratory Symptoms domain scores.
CONCLUSIONS
In adults with CF, the IPOS has acceptable internal consistency and there is evidence of construct validity. These findings support adoption of the IPOS in the primary palliative care model for CF.
PubMed: 38934771
DOI: 10.1002/ppul.27143 -
MSystems Jun 2024Airway microbiota are known to contribute to lung diseases, such as cystic fibrosis (CF), but their contributions to pathogenesis are still unclear. To improve our...
Airway microbiota are known to contribute to lung diseases, such as cystic fibrosis (CF), but their contributions to pathogenesis are still unclear. To improve our understanding of host-microbe interactions, we have developed an integrated analytical and bioinformatic mass spectrometry (MS)-based metaproteomics workflow to analyze clinical bronchoalveolar lavage (BAL) samples from people with airway disease. Proteins from BAL cellular pellets were processed and pooled together in groups categorized by disease status (CF vs. non-CF) and bacterial diversity, based on previously performed small subunit rRNA sequencing data. Proteins from each pooled sample group were digested and subjected to liquid chromatography tandem mass spectrometry (MS/MS). MS/MS spectra were matched to human and bacterial peptide sequences leveraging a bioinformatic workflow using a metagenomics-guided protein sequence database and rigorous evaluation. Label-free quantification revealed differentially abundant human peptides from proteins with known roles in CF, like neutrophil elastase and collagenase, and proteins with lesser-known roles in CF, including apolipoproteins. Differentially abundant bacterial peptides were identified from known CF pathogens (e.g., ), as well as other taxa with potentially novel roles in CF. We used this host-microbe peptide panel for targeted parallel-reaction monitoring validation, demonstrating for the first time an MS-based assay effective for quantifying host-microbe protein dynamics within BAL cells from individual CF patients. Our integrated bioinformatic and analytical workflow combining discovery, verification, and validation should prove useful for diverse studies to characterize microbial contributors in airway diseases. Furthermore, we describe a promising preliminary panel of differentially abundant microbe and host peptide sequences for further study as potential markers of host-microbe relationships in CF disease pathogenesis.IMPORTANCEIdentifying microbial pathogenic contributors and dysregulated human responses in airway disease, such as CF, is critical to understanding disease progression and developing more effective treatments. To this end, characterizing the proteins expressed from bacterial microbes and human host cells during disease progression can provide valuable new insights. We describe here a new method to confidently detect and monitor abundance changes of both microbe and host proteins from challenging BAL samples commonly collected from CF patients. Our method uses both state-of-the art mass spectrometry-based instrumentation to detect proteins present in these samples and customized bioinformatic software tools to analyze the data and characterize detected proteins and their association with CF. We demonstrate the use of this method to characterize microbe and host proteins from individual BAL samples, paving the way for a new approach to understand molecular contributors to CF and other diseases of the airway.
PubMed: 38934598
DOI: 10.1128/msystems.00929-23 -
Histology and Histopathology Jun 2024Five cases of patients with systemic connective tissue diseases (CTD) who developed connective tissue disease-associated interstitial lung disease (CTD-ILD) with...
Five cases of patients with systemic connective tissue diseases (CTD) who developed connective tissue disease-associated interstitial lung disease (CTD-ILD) with progressive pulmonary fibrosis (PPF) are reported here. Unspecified ILD was diagnosed using high-resolution computed tomography (HRCT). Histologically, all cases were usual interstitial pneumonia (UIP) with findings of advanced (3/5) to diffuse (2/5) fibrosis, with a partially (4/5) to completely (1/5) formed image of a honeycomb lung. The fibrosis itself spread subpleurally and periseptally to more central parts (2/5) of the lung, around the alveolar ducts (2/5), or even without predisposition (1/5). Simultaneously, there was architectural reconstruction based on the mutual fusion of fibrosis without compression of the surrounding lung parenchyma (1/5), or with its compression (4/5). The whole process was accompanied by multifocal (1/5), dispersed (2/5), or organized inflammation in aggregates and lymphoid follicles (2/5). As a result of continuous fibroproduction and maturation of the connective tissue, the alveolar septa thickened, delimiting groups of alveoli that merged into air bullae. Few indistinctly visible (2/5), few clearly visible (1/5), multiple indistinctly visible (1/5), and multiple clearly visible (1/5) fibroblastic foci were present. Among the concomitant changes, areas of emphysema, bronchioloectasia, and bronchiectasis, as well as bronchial and vessel wall hypertrophy, and mucostasis in the alveoli and edema were observed. The differences in the histological appearance of usual interstitial pneumonia associated with systemic connective tissue diseases (CTD-UIP) versus the pattern associated with idiopathic pulmonary fibrosis (IPF-UIP) are discussed here. The main differences lie in spreading lung fibrosis, architectural lung remodeling, fibroblastic foci, and inflammatory infiltrates.
PubMed: 38934227
DOI: 10.14670/HH-18-777