Biology subjects

Kudrna, K.

Publications and source records attributed to Kudrna, K..

3 recordsLinked to original sources

bioRxiv2026

Autonomous MTOR activation in COPD airway epithelium impairs resolution through persistent mucus cell metaplasia and lysosome dysfunction

Background: COPD is associated with persistent airway epithelial mucous cell metaplasia (MCM), mucin hypersecretion, and airway obstruction. Cytokines, such as IL-13, are well-known inflammatory drivers of airway MCM development. However, less is known about the factors that contribute to the impaired resolution of MCM in COPD. We hypothesized that MTOR activation in COPD airway epithelial cells (AECs) impairs autophagy and contributes to the delayed resolution of MCM. Methods: AEC from COPD and non-diseased donors were grown under air-liquid interface (ALI) conditions and treated with IL-13 to promote MCM. Airway sections from COPD lung explants or non-diseased lung donors were utilized for immunohistochemical and immunostaining. Lysosomes were assessed using molecular probes and immunostaining. Results: There was persistent MTOR-dependent MUC5AC immunostaining and epithelial hypertrophy in COPD AEC, which did not resolve after IL-13 withdrawal. MTOR substrate phosphorylation of RBS6, S6K1, and ULK1 was significantly elevated under baseline conditions, and at multiple timepoints independent of IL-13. Pharmacologic activation of MTOR led to increased IL-13-mediated MUC5AC levels in normal AEC but less robustly in COPD derived AEC. We hypothesized that persistent MTOR activation would reduce lysosome function and abundance in disease. COPD airways had reduced lysosome markers, LAMP1 and LAMP2 and fewer functional lysosomes by live-cell reporter probes. This lysosome deficiency in COPD AEC was partially rescued with MTOR inhibition. Conclusions: We provide evidence to support an axis of autonomous MTOR activation and impaired lysosome function in the COPD airway epithelium. Persistent MTOR signaling is a molecular driver that contributes to persistent MCM.

pathology↗

bioRxiv2024

mTOR signaling regulates aberrant epithelial cell proliferative and migratory behaviors characteristic of airway mucous metaplasia in asthma

In asthma, the airway epithelium is hyperplastic, hypertrophied, and lined with numerous large MUC5AC-containing goblet cells (GC). Furthermore, the normal epithelial architecture is disorganized with numerous, what we here describe as, ectopic goblet cells (eGC) deep within the thickened epithelial layer disconnected from the lumenal surface. mTOR is a highly conserved pathway that regulates cell size and proliferation. We hypothesized that the balance between mTOR and autophagy signaling regulates key features of the asthma epithelial layer. Airway histological sections from subjects with asthma had increased frequency of eGC and increased levels of mTOR phosphorylation target-Ribosomal S6. Using human airway epithelial cells (hAECs) with IL-13 stimulation and timed withdrawal to stimulate resolution, we found that multiple key downstream phosphorylation targets downstream from the mTOR complex were increased during early IL-13-mediated mucous metaplasia, and then significantly declined during resolution. The IL-13-mediated changes in mTOR signaling were paralleled by morphologic changes with airway epithelial hypertrophy, hyperplasia, and frequency of eGC. We then examined the relationship between mTOR and autophagy using mice deficient in autophagy protein Atg16L1. Despite having increased cytoplasmic mucins, mouse AECs from Atg16L1 deficient mice had no significant difference in mTOR downstream signaling. mTOR inhibition with rapamycin led to a loss of IL-13-mediated epithelial hypertrophy, hyperplasia, ectopic GC distribution, and reduction in cytoplasmic MUC5AC levels. mTOR inhibition was also associated with a reduction in aberrant IL-13-mediated hAEC proliferation and migration. Our findings demonstrate that mTOR signaling is associated with mucous metaplasia and is crucial to the disorganized airway epithelial structure and function characteristic of muco-obstructive airway diseases such as asthma. Graphical Abstract Key ConceptsO_LIThe airway epithelium in asthma is disorganized and characterized by cellular proliferation, aberrant migration, and goblet cell mucous metaplasia. C_LIO_LImTOR signaling is a dynamic process during IL-13-mediated mucous metaplasia, increasing with IL-13 stimulation and declining during resolution. C_LIO_LImTOR signaling is strongly increased in the asthmatic airway epithelium. C_LIO_LImTOR signaling is associated with the development of key features of the metaplastic airway epithelium including cell proliferation and ectopic distribution of goblet cells and aberrant cellular migration. C_LIO_LIInhibition of mTOR leads to decreased epithelial hypertrophy, reduced ectopic goblet cells, and cellular migration. C_LI

cell biology↗

bioRxiv2020

Mucin granules are degraded in the autophagosome-lysosome pathway as a means of resolving airway mucous cell metaplasia

Exacerbations of muco-obstructive airway diseases such as COPD and asthma are associated with epithelial changes termed mucous cell metaplasia (MCM). The molecular pathways triggering MCM have been identified; however, the factors that regulate resolution are less well understood. We hypothesized that the autophagosome-lysosome pathway is required for resolution of MCM by degrading cytoplasmic mucins. We found increased intracellular levels of Muc5ac and Muc5b in autophagy-deficient mice. This difference was not due to defective mucin secretion. Instead, we found that Lamp1-labeled lysosomes surrounded mucin granules of mucous cells indicating that granules were being degraded. Using a model of resolution of mucous cell metaplasia in mice, we found increased lysosomal proteolytic activity that peaked in the days after inflammation. Autophagy-deficient mice had persistent accumulation of mucin granules that failed to decline due to reduced mucin degradation. We applied these findings in vitro to human airway epithelial cells (AECs). Activation of autophagy by mTOR inhibition led to degradation of mucin granules in AECs. Our findings indicate that during peak and resolution phases of MCM, mucin granules can be degraded by autophagy. The addition of mucin degradation to the existing paradigm of production and secretion may more fully explain how the secretory cells handle excess amounts of cytoplasmic mucin and offers a therapeutic target to speed resolution of MCM in airway disease exacerbations. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/130534v2_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@94239org.highwire.dtl.DTLVardef@1dac6dforg.highwire.dtl.DTLVardef@368161org.highwire.dtl.DTLVardef@680b7d_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗