Human Tracheobronchial Basal Cells. Normal versus Remodeling/Repairing PhenotypesIn VivoandIn Vitro

Abstract

Human tracheobronchial epithelial (TBE) basal cells (BCs) function as progenitors in normal tissue. However, mechanistic studies are typically performed in vitro and frequently use BCs recovered from patients who die of nonrespiratory disease. It is not knownwhether the cadaveric epithelium (1) is undergoing homeostatic remodeling and/or repair, or (2) yields BC clones that represent homeostatic processes identifiedin tissue. We sought to compare the phenotype of TBE-BCs with that of BCs cultured under optimal clone-forming conditions. TBE pathology was evaluated using quantitative histomorphometry. The cultured BC phenotype was determined by fluorescence-activated cell sorter analysis. Clone organization and cell phenotype were determined by immunostaining. The cadaveric TBE is 20% normal. In these regions, BCs are keratin (K)-5 1 and tetraspanin CD1511 , and demonstrate a low mitotic index. In contrast, 80% of the cadaveric TBE exhibits homeostatic remodeling/repair processes. In these regions, BCs are K5 1 /K15 1 /K14 1 on Recovery Days 3-13 (5). These injury/repair studies suggested that the human BC phenotype may also vary as a function of tissue homeostasis or wounding. Our primary aim involved determining the K5/K14 profile of human TBE BCs.Keratins are cytoplasmic proteins (6, 7). Consequently, viable respiratory BCs cannot be isolated on the basis of K expression. To overcome this issue, several groups identified cell-surface markers that can be used to separate BCs into subsets, using a fluorescence-activated cell sorter (FACS) (2,3,(8)(9)(10). One of these markers, tissue factor (TF), is a component of the extrinsic coagulation cascade (11), and was originally developed for the isolation of nasal polyp BCs (12). We showed that all Passage 1 TBE BCs that were cultured in bronchial epithelial cell growth medium (BEGM) were TF 1 , and that TF activity was necessary for BC survival in vitro (13). Other TBE BC markers include nerve growth factor receptor (2), podoplanin (14), CD49f (a6 integrin) (8), the tetraspanin CD151 1 (12), and Trp63 (p63) (15). Our secondary aim involved determining whether TF and CD151 were (1) expressed by TBE BCs, and (2) could be used to identify TBE BC subsets.Our previous studies and those of others indicate that TBE repair in mice is mediated by BCs that proliferate and then differentiate to replace ciliated and secretory cell types (2,5,8,14,(16)(17)(18)(19). Human BC progenitor functions can also be evaluated in vitro (20). However, most analyses of human BCs have focused on the BC population as a whole, rather than on the CLINICAL RELEVANCEHuman basal cells (BCs) function as progenitors for tracheobronchial epithelial repair and regeneration. However, the mechanisms regulating BC function are typically evaluated in vitro. The impact of these mechanistic studies is limited by a lack of cross-referencing in tissue and cultured BC phenotype and function. We show that human tracheobronchial BC phenotypes are heterogeneous in vivo, and become more homogeneous in vitro. Highly ...

IL-13 Augments Compressive Stress–Induced Tissue Factor Expression in Human Airway Epithelial Cells

“…Our data are the first to show enhanced TF expression under allergic inflammatory conditions in mice, indicating that this model will be useful for studying the function of TF in asthma. The distribution of TF in mouse lung epithelial cells was unknown, although, in humans, it is considered a basal cell marker (47,48). We show that, as in humans, TF is abundantly expressed in basal epithelial cells in the mouse airway.…”

Lung organoids: current uses and future promise

“…Lineage tracing and other studies have shown that in the adult mouse lung there is only very slow turnover of the mucociliary epithelium and replacement of luminal cells from basal cells and their immediate progeny (Ghosh et al, 2013(Ghosh et al, , 2011Hong et al, 2004;Rock et al, , 2009Watson et al, 2015). However, following cell damage by agents typically used experimentally -for example naphthalene, which kills secretory Club cells, or SO 2 gas, which kills all luminal cells -or viral infection there are rapid changes in the behavior and proliferation of the basal cells so that they quickly regenerate the epithelium and restore barrier function.…”

Airway Progenitor Clone Formation Is Enhanced by Y-27632–Dependent Changes in the Transcriptome

“…The number of progenitor cells was determined using the limiting dilution assay (23) modified for lung (24)(25)(26).…”

“…Cell surface phenotype was determined using antibodies and sorting methods as previously described (24,27). Cell cycle analysis was performed using the method of Krishan (28) and assayed on a Gallios 561 instrument equipped with a Hypercyte 96-well plate reader (Beckman Coulter, Indianapolis, IN).…”

“…Cultures were generated and analyzed according to published methods (5,24,29), and are described in detail in the online supplement.…”