Idiopathic pulmonary fibrosis (IPF) is a progressive disease with unremitting extracellular matrix deposition, leading to a distortion of pulmonary architecture and impaired gas exchange. Fibroblasts from IPF patients acquire an invasive phenotype that is essential for progressive fibrosis. Here, we performed RNA-seq analysis on invasive and non-invasive fibroblasts and found that the immune checkpoint ligand CD274 (PD-L1) was up-regulated on invasive lung fibroblasts and was required for the invasive phenotype of lung fibroblasts, is regulated by P53 and FAK, and drives lung fibrosis in a humanized IPF model in mice. Activating CD274 in IPF fibroblasts promoted invasion in vitro and pulmonary fibrosis in vivo. CD274 knockout in IPF fibroblasts and targeting CD274 by FAK inhibition or CD274 neutralizing antibodies blunted invasion and attenuated fibrosis, suggesting that CD274 may be a novel therapeutic target in IPF.
Yan Geng, Xue Liu, Jiurong Liang, David M. Habiel, Kulur Vrishika, Ana Lucia Coelho, Nan Deng, Ting Xie, Yizhou Wang, Ningshan Liu, Guanling Huang, Adrianne Kurkciyan, Zhenqiu Liu, Jie Tang, Cory M. Hogaboam, Dianhua Jiang, Paul W. Noble
Recovery from acute lung injury (ALI) is an active process. Foxp3+ regulatory T cells (Tregs) contribute to recovery from ALI through modulating immune responses and enhancing alveolar epithelial proliferation and tissue repair. The current study investigates Treg transcriptional profiles during resolution of ALI in mice. Tregs from either lung or splenic tissue were isolated from uninjured mice or mice recovering from ALI and then examined for differential gene expression between these conditions. In mice with ALI, Tregs isolated from the lungs had hundreds of differentially expressed transcripts compared to those from the spleen, indicating that organ-specificity and microenvironment are critical in Treg function. These regulated transcripts suggest which intracellular signaling pathways modulate Treg behavior. Interestingly, several transcripts having no prior recognized function in Tregs were differentially expressed by lung Tregs during resolution. Further investigation into two identified transcripts, Mmp12 and Sik1, revealed that Treg-specific expression of each play a role in Treg-promoted ALI resolution. This study provides novel information describing the signals that may expand resident Tregs, recruit or retain them to the lung during ALI, and modulate their function. The results provide insight into both tissue- and immune microenvironment-specific transcriptional differences through which Tregs direct their effects.
Jason R. Mock, Catherine F. Dial, Miriya K. Tune, Dustin L. Norton, Jessica R. Martin, John C. Gomez, Robert S. Hagan, Hong Dang, Claire M. Doerschuk
Although Type-2 (T2) induced epithelial dysfunction is likely to profoundly alter epithelial differentiation and repair in asthma, the mechanisms for these effects are poorly understood. A role for specific mucins, heavily N-glycosylated epithelial glycoproteins, in orchestrating epithelial cell fate in response to T2 stimuli has not previously been investigated. Levels of a sialylated MUC4β isoform were found to be increased in airway specimens from asthmatic patients, in association with T2 inflammation. We hypothesized that IL-13 would increase sialylation of MUC4β, thereby altering its function and that the β-galactoside α-2,6-sialyltranferase 1 (ST6GAL1) would regulate the sialylation. Using human biologic specimens and cultured primary human airway epithelial cells (HAECs), we demonstrated that IL-13 increased sialylation of MUC4β under control of ST6GAL1, and that both were increased in asthma, particularly in those with elevated T2 biomarkers. ST6GAL1 induced sialylation of MUC4β altered its lectin binding and secretion. Both ST6GAL1 and MUC4β inhibited epithelial cell proliferation while promoting goblet cell differentiation. These in vivo and in vitro data provide strong evidence for a critical role for ST6GAL1 induced sialylation of MUC4β in epithelial dysfunction associated with T2-High asthma, thereby identifying specific sialylation pathways as potential targets in asthma.
Xiuxia Zhou, Carol L. Kinlough, Rebecca P. Hughey, Mingzhu Jin, Hideki Inoue, Emily Etling, Brian D. Modena, Naftali Kaminski, Eugene R. Bleecker, Deborah A. Meyers, Nizar N. Jarjour, John B. Trudeau, Fernando Holguin, Anuradha Ray, Sally E. Wenzel
Lung transplantation (LTx) is the only therapeutic option for many patients with chronic lung disease. However, long-term survival after LTx is severely compromised by chronic rejection (chronic lung allograft dysfunction [CLAD]), which affects 50% of recipients after 5 years. The underlying mechanisms for CLAD are poorly understood, largely due to a lack of clinically relevant animal models, but lymphocytic bronchiolitis is an early sign of CLAD. Here, we report that lymphocytic bronchiolitis occurs early in a long-term murine orthotopic LTx model, based on a single mismatch (grafts from HLA-A2:B6–knockin donors transplanted into B6 recipients). Lymphocytic bronchiolitis is followed by formation of B cell–dependent lymphoid follicles that induce adjacent bronchial epithelial cell dysfunction in a spatiotemporal fashion. B cell deficiency using recipient μMT–/– mice prevented intrapulmonary lymphoid follicle formation and lymphocytic bronchiolitis. Importantly, selective inhibition of the follicle-organizing receptor EBI2, using genetic deletion or pharmacologic inhibition, prevented functional and histological deterioration of mismatched lung grafts. In sum, we provided what we believe to be a mouse model of chronic rejection and lymphocytic bronchiolitis after LTx and identified intrapulmonary lymphoid follicle formation as a target for pharmacological intervention of long-term allograft dysfunction after LTx.
Natalia F. Smirnova, Thomas M. Conlon, Carmela Morrone, Peter Dorfmuller, Marc Humbert, Georgios Stathopoulos, Stephan Umkehrer, Franz Pfeiffer, Ali Ö. Yildirim, Oliver Eickelberg
Acid aspiration, which can result from several etiologies, including postoperative complications, leads to direct contact of concentrated hydrochloric acid (HCl) with the alveolar epithelium. As a result, rapid endothelial activation induces alveolar inflammation, leading to life-threatening pulmonary edema. Because mechanisms underlying the rapid endothelial activation are not understood, here we determined responses in real time through optical imaging of alveoli of live mouse lungs. By alveolar micropuncture, we microinfused concentrated HCl in the alveolar lumen. As expected, acid contact with the epithelium caused rapid, but transient, apical injury. However, there was no concomitant membrane injury to the endothelium. Nevertheless, H2O2-mediated epithelial-endothelial paracrine signaling induced endothelial barrier failure, as detected by microvascular dextran leakage and lung water quantification. Remarkably, endothelial mitochondria regulated the barrier failure by activating uncoupling protein 2 (UCP2), thereby inducing transient mitochondrial depolarization that led to cofilin-induced actin depolymerization. Knockdown, or endothelium-targeted deletion of UCP2 expression, blocked these responses, including pulmonary edema. To our knowledge, these findings are the first to mechanistically implicate endothelial mitochondria in acid-induced barrier deterioration and pulmonary edema. We suggest endothelial UCP2 may be a therapeutic target for acid-induced acute lung injury.
Rebecca F. Hough, Mohammad N. Islam, Galina A. Gusarova, Guangchun Jin, Shonit Das, Jahar Bhattacharya
Increased airway vagal sensory C-fiber activity contributes to the symptoms of inflammatory airway diseases. The KCNQ/Kv7/M-channel is a well-known determinant of neuronal excitability, yet whether it regulates the activity of vagal bronchopulmonary C-fibers and airway reflex sensitivity remain unknown. Here we addressed this issue using single-cell RT-PCR, patch clamp technique, extracellular recording of single vagal nerve fibers innervating the mouse lungs, and telemetric recording of cough in free-moving mice. Single-cell mRNA analysis and biophysical properties of M-current (IM) indicate that KCNQ3/Kv7.3 is the major M-channel subunit in mouse nodose neurons. The M-channel opener retigabine negatively shifted the voltage-dependent activation of IM, leading to membrane hyperpolarization, increased rheobase and suppression of both evoked and spontaneous action potential (AP) firing in nodose neurons in the M-channel inhibitor XE991-sensitive manner. Retigabine also markedly suppressed the α,β-methylene ATP-induced AP firing in nodose C-fiber terminals innervating the mouse lungs, and irritant gases-evoked coughing in awake mice. In conclusion, KCNQ/M-channels play a role in regulating the excitability of vagal airway C-fibers at both the cell soma and nerve terminals. Drugs that open M-channels in airway sensory afferents may relieve the sufferings associated with pulmonary inflammatory diseases such as chronic coughing.
Hui Sun, An-Hsuan Lin, Fei Ru, Mayur J. Patil, Sonya Meeker, Lu-Yuan Lee, Bradley J. Undem
Macrophages are well-recognized for their dual roles in orchestrating inflammatory responses and regulating tissue repair. In almost all acutely inflamed tissues, two main subclasses of macrophages co-exist. These include embryonically-derived resident tissue macrophages and bone marrow-derived recruited macrophages. While it is clear that macrophage subsets categorized in this fashion display distinct transcriptional and functional profiles, whether all cells within these categories and in the same inflammatory microenvironment share similar functions or whether further specialization exists has not been determined. To investigate inflammatory macrophage heterogeneity on a more granular level, we induced acute lung inflammation in mice and performed single cell RNA sequencing of macrophages isolated from the airspaces during health, peak inflammation, and resolution of inflammation. In doing so, we confirm that cell origin is the major determinant of AM programing and describe two previously uncharacterized, transcriptionally distinct subdivisions of AMs based on proliferative capacity and inflammatory programing.
Kara J. Mould, Nathan D. Jackson, Peter M. Henson, Max A. Seibold, William J. Janssen
Airway mucin secretion is necessary for ciliary clearance of inhaled particles and pathogens, but can be detrimental in pathologies such as asthma and cystic fibrosis. Exocytosis in mammals requires a Munc18 scaffolding protein, and airway secretory cells express all three Munc18 isoforms. Using conditional airway epithelial deletant mice, we found that Munc18a has the major role in baseline mucin secretion, Munc18b has the major role in stimulated mucin secretion, and Munc18c does not function in mucin secretion. In an allergic asthma model, Munc18b deletion reduced airway mucus occlusion and airflow resistance. In a cystic fibrosis model, Munc18b deletion reduced airway mucus occlusion and emphysema. Munc18b deficiency in the airway epithelium did not result in any abnormalities of lung structure, particle clearance, inflammation, or bacterial infection. Our results show that regulated secretion in a polarized epithelial cell may involve more than one exocytic machine at the apical plasma membrane, and that the protective roles of mucin secretion can be preserved while therapeutically targeting its pathologic roles.
Ana M. Jaramillo, Lucia Piccotti, Walter V. Velasco, Anna Sofia Huerta Delgado, Zoulikha Azzegagh, Felicity S. Chung, Usman I. Nazeer, Junaid Farooq, Joshua M. Brenner, Jan Parker-Thornburg, Brenton L. Scott, Christopher M. Evans, Roberto Adachi, Alan R. Burns, Silvia M. Kreda, Michael J. Tuvim, Burton F. Dickey
Alveolar type 2 (AT2) cell endoplasmic reticulum (ER) stress is a prominent feature in adult and pediatric interstitial lung disease (ILD and ChILD), but in vivo models linking AT2 cell ER stress to ILD have been elusive. Based on a clinical ChILD case we identified a critical cysteine residue in the Surfactant Protein C gene (SFTPC) BRICHOS domain whose mutation induced ER stress in vitro. To model this in vivo, we generated a knock-in model expressing a cysteine-to-glycine substitution at codon 121 (C121G) in the Sftpc gene. SftpcC121G expression during fetal development resulted in a toxic gain of function resulting in fatal post-natal respiratory failure from disrupted lung morphogenesis. Induced SftpcC121G expression in adult mice resulted in an ER retained pro-protein causing AT2 cell ER stress. SftpcC121G AT2 cells were a source of cytokines expressed in concert with development of a polycellular alveolitis. These cytokines were subsequently found in a high-dimensional proteomic screen of bronchoalveolar lavage fluid from ChILD patients with the same class of SFTPC mutations. Following alveolitis resolution, SftpcC121G mice developed spontaneous pulmonary fibrosis and restrictive lung impairment. This model provides proof of concept linking AT2 cell ER stress to fibrotic lung disease coupled with translationally relevant biomarkers.
Jeremy Katzen, Brandie D. Wagner, Alessandro Venosa, Meghan Kopp, Yaniv Tomer, Scott J. Russo, Alvis C. Headen, Maria C. Basil, James M. Stark, Surafel Mulugeta, Robin R. Deterding, Michael F. Beers
Macrophage activation, i.e., the classical M1 and the alternative M2, plays a critical role in many pathophysiological processes, such as inflammation and tissue injury and repair. Although the regulation of macrophage activation has been under extensive investigations, there is little knowledge about the role of long non-coding RNAs (lncRNAs) in the event. In this study, we found that lncRNA Malat1 expression is distinctly regulated in differentially activated macrophages in that it is upregulated in LPS-, whereas downregulated in IL-4-treated cells. Malat1 knockdown attenuates LPS induced M1 macrophage activation. In contrast, Malat1 knockdown enhanced IL-4 activated M2 differentiation as well as macrophage pro-fibrotic phenotype. Mechanistically, Malat1 knockdown led to decreased expression of Clec16a, of which silencing phenocopied the regulatory effect of Malat1 on M1 activation. Interestingly, Malat1 knockdown promoted IL-4 induction of mitochondrial pyruvate carriers (MPCs) and their mediation of glucose derived oxidative phosphorylation (OxPhos), which was crucial to the Malat1 regulation of M2 differentiation and pro-fibrotic phenotype. Furthermore, mice with either global or conditional myeloid knockout of Malat1 demonstrated diminished LPS induced systemic and pulmonary inflammation and injury. Conversely, these mice developed more severe bleomycin induced lung fibrosis, accompanied by alveolar macrophages displaying augmented M2 and pro-fibrotic phenotype. In summary, we have identified a previously unrecognized role of Malat1 in regulation of macrophage polarization. Our data demonstrate that Malat1 is involved in pulmonary pathogeneses in association with aberrant macrophage activation.
Huachun Cui, Sami Banerjee, Sijia Guo, Na Xie, Jing Ge, Dingyuan Jiang, Martin Zörnig, Victor J. Thannickal, Gang Liu
No posts were found with this tag.