Human rhinoviruses (HRV) are common cold viruses associated with exacerbations of lower airways diseases. Although viral induced epithelial damage mediates inflammation, the molecular mechanisms responsible for airway epithelial damage and dysfunction remain undefined. Using experimental HRV infection studies in highly differentiated human bronchial epithelial cells grown at air-liquid interface (ALI), we examine the links between viral host defense, cellular metabolism, and epithelial barrier function. We observe that early HRV-C15 infection induces a transitory barrier-protective metabolic state characterized by glycolysis that ultimately becomes exhausted as the infection progresses and leads to cellular damage. Pharmacological promotion of glycolysis induces ROS-dependent upregulation of the mitochondrial metabolic regulator, peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α), thereby restoring epithelial barrier function, improving viral defense, and attenuating disease pathology. Therefore, PGC-1α regulates a metabolic pathway essential to host defense that can be therapeutically targeted to rescue airway epithelial barrier dysfunction and potentially prevent severe respiratory complications or secondary bacterial infections.

Human rhinovirus (HRV) infections are the primary cause of the common cold and are a major trigger for exacerbations of lower airway diseases, such as asthma and chronic obstructive pulmonary diseases. Although human bronchial epithelial cells (HBE) are the natural host for HRV infections, much of our understanding of how HRV replicates and induces host antiviral responses is based on studies using non-airway cell lines (e.g. HeLa cells). The current study examines the replication cycle of HRV, and host cell responses, in highly differentiated cultures of HBE. Highly differentiated cultures of HBE were exposed to initial infectious doses ranging from 104 to 101 50% tissue culture-infective dose (TCID50) of purified HRV-16, and responses were monitored up to 144 h after infection. Viral genomic RNA and negative strand RNA template levels were monitored, along with levels of type I and II interferons and selected antivirals. Regardless of initial infectious dose, relatively constant levels of both genomic and negative strand RNA are generated during replication, with negative strand copy numbers being10,000-fold lower than those of genomic strands. Infections were limited to a small percentage of ciliated cells and did not result in any overt signs of epithelial death. Importantly, regardless of infectious dose, HRV-16 infections were cleared by HBE in the absence of immune cells. Levels of type I and type III interferons (IFNs) varied with initial infectious dose, implying that factors other than levels of double-stranded RNA regulate IFN induction, but the time-course of HRV-16 clearance HBE was the same regardless of levels of IFNs produced. Patterns of antiviral viperin and ISG15 expression suggest they may be generated in an IFN-independent manner during HRV-16 infections. These data challenge a number of aspects of dogma generated from studies in HeLa cells and emphasize the importance of appropriate cell context when studying HRV infections.

Virus–bacteria coinfections are associated with more severe exacerbations and increased risk of hospital readmission in patients with chronic obstructive pulmonary disease (COPD). The airway epithelium responds to such infections by releasing proinflammatory and antimicrobial cytokines, including IL-17C. However, the regulation and role of IL-17C is not well understood. In this study, we examine the mechanisms regulating IL-17C production and its potential role in COPD exacerbations. Human bronchial epithelial cells (HBE) obtained from normal, nontransplanted lungs or from brushings of nonsmokers, healthy smokers, or COPD patients were exposed to bacteria and/or human rhinovirus (HRV). RNA and protein were collected for analysis, and signaling pathways were assessed with pharmacological agonists, inhibitors, or small interfering RNAs. HBE were also stimulated with IL-17C to assess function. HRV–bacterial coinfections synergistically induced IL-17C expression. This induction was dependent on HRV replication and required NF-κB–mediated signaling. Synergy was lost in the presence of an inhibitor of the p38 MAP kinase pathway. HBE exposed to IL-17C show increased gene expression of CXCL1, CXCL2, NFKBIZ, and TFRC, and release CXCL1 protein, a neutrophil chemoattractant. Knockdown of IL-17C significantly reduced induction of CXCL1 in response to HRV–bacterial coinfection as well as neutrophil chemotaxis. HBE from healthy smokers release less IL-17C than cells from nonsmokers, but cells from COPD patients release significantly more IL-17C compared with either nonsmokers or healthy smokers. These data suggest that IL-17C may contribute to microbial-induced COPD exacerbations by promoting neutrophil recruitment.

Ligand-activated glucocorticoid receptor (GR) elicits variable glucocorticoid-modulated transcriptomes in different cell types. However, some genes, including Krüppel-like factor 9 (KLF9), a putative transcriptional repressor, demonstrate conserved responses. We show that glucocorticoids induce KLF9 expression in the human airways in vivo and in differentiated human bronchial epithelial (HBE) cells grown at air–liquid interface (ALI). In A549 and BEAS-2B pulmonary epithelial cells, glucocorticoids induce KLF9 expression with similar kinetics to primary HBE cells in submersion culture. A549 and BEAS-2B ChIP-seq data reveal four common glucocorticoid-induced GR binding sites (GBSs). Two GBSs mapped to the 5ʹ-proximal region relative to KLF9 transcription start site (TSS) and two occurred at distal sites. These were all confirmed in primary HBE cells. Global run-on (GRO) sequencing indicated robust enhancer RNA (eRNA) production from three of these GBSs in BEAS-2B cells. This was confirmed in A549 cells, plus submersion, and ALI culture of HBE cells. Cloning each GBS into luciferase reporters revealed glucocorticoid-induced activity requiring a glucocorticoid response element (GRE) within each distal GBS. While the proximal GBSs drove modest reporter induction by glucocorticoids, this region exhibited basal eRNA production, RNA polymerase II enrichment, and looping to the TSS, plausibly underlying constitutive KLF9 expression. Post glucocorticoid treatment, interactions between distal and proximal GBSs and the TSS correlated with KLF9 induction. CBP/P300 silencing reduced proximal GBS activity, but negligibly affected KLF9 expression. Overall, a model for glucocorticoid-mediated regulation of KLF9 involving multiple GBSs is depicted. This work unequivocally demonstrates that mechanistic insights gained from cell lines can translate to physiologically relevant systems.

Human rhinoviruses have been linked both to the susceptibility of asthma development and to the triggering of acute exacerbations. Given that the human airway epithelial cell is the primary site of human rhinovirus (HRV) infection and replication, the current review focuses on how HRV-induced modulation of several aspects of epithelial cell phenotype could contribute to the development of asthma or to the induction of exacerbations. Modification of epithelial proinflammatory and antiviral responses are considered, as are alterations in an epithelial barrier function and cell phenotype. The contributions of the epithelium to airway remodeling and to the potential modulation of immune responses are also considered. The potential interactions of each type of HRV-induced epithelial phenotypic changes with allergic sensitization and allergic phenotype are also considered in the context of asthma development and of acute exacerbations.

Triple-negative breast cancer (TNBC) is a highly aggressive and metastatic form of breast cancer that lacks an effective targeted therapy. To identify potential therapeutic targets, we investigated the phosphohistidine phosphatase, LHPP, which has been implicated in the development of several types of cancer. However, the full significance of LHPP in cancer progression remains unclear due to our limited understanding of its molecular mechanism. We found that levels of the LHPP phosphohistidine phosphatase were significantly increased in human breast cancer patients compared to normal adjacent tissues, with the highest levels in the TNBC subtype. When LHPP was knocked out in the MDA-MB-231 human TNBC cell line, cell proliferation, wound healing capacity, and invasion were significantly reduced. However, LHPP knockout in TNBC cells did not significantly affect overall phosphohistidine protein levels. Interestingly, LHPP knockout in MDA-MB-231 cells delayed tumor growth and reduced metastasis when orthotopically transplanted into mouse mammary glands. To investigate LHPP’s role in breast cancer progression, we used next-generation sequencing and proximity-labeling proteomics, and found that LHPP regulates gene expression in chemokine-mediated signaling and actin cytoskeleton organization. Depletion of LHPP reduced the presence of tumor-infiltrating macrophages in mouse xenografts. Our results support a tumor promoter role for LHPP phosphohistidine phosphatase in MDA-MB-231TNBC cells and suggest that targeting LHPP phosphatase could be a potential therapeutic strategy for TNBC.