Accumulating evidence suggests that altered cellular metabolism is systemic in pulmonary hypertension (PH) and central to disease pathogenesis. However, bioenergetic changes in PH patients and their association with disease severity remain unclear. Here, we hypothesize that alteration in bioenergetic function is present in platelets from PH patients and correlates with clinical parameters of PH. Platelets isolated from controls and PH patients (
Quyen L. Nguyen, Catherine Corey, Pamela White, Annie Watson, Mark T. Gladwin, Marc A. Simon, Sruti Shiva
Ciliary motion defects cause defective mucociliary transport (MCT) in primary ciliary dyskinesia (PCD). Current diagnostic tests do not assess how MCT is affected by perturbation of ciliary motion. In this study, we sought to use micro-optical coherence tomography (μOCT) to delineate the mechanistic basis of cilia motion defects of PCD genes by functional categorization of cilia motion. Tracheae from three PCD mouse models were analyzed using μOCT to characterize ciliary motion and measure MCT. We developed multiple measures of ciliary activity, integrated these measures, and quantified dyskinesia by the angular range of the cilia effective stroke (ARC).
George M. Solomon, Richard Francis, Kengyeh K. Chu, Susan E. Birket, George Gabriel, John E. Trombley, Kristi L. Lemke, Nikolai Klena, Brett Turner, Guillermo J. Tearney, Cecilia W. Lo, Steven M. Rowe
Idiopathic pulmonary fibrosis (IPF) is a severe fibrotic lung disease associated with fibroblast activation that includes excessive proliferation, tissue invasiveness, myofibroblast transformation, and extracellular matrix (ECM) production. To identify inhibitors that can attenuate fibroblast activation, we queried IPF gene signatures against a library of small-molecule-induced gene-expression profiles and identified Hsp90 inhibitors as potential therapeutic agents that can suppress fibroblast activation in IPF. Although Hsp90 is a molecular chaperone that regulates multiple processes involved in fibroblast activation, it has not been previously proposed as a molecular target in IPF. Here, we found elevated Hsp90 staining in lung biopsies of patients with IPF. Notably, fibroblasts isolated from fibrotic lesions showed heightened Hsp90 ATPase activity compared with normal fibroblasts. 17-
Vishwaraj Sontake, Yunguan Wang, Rajesh K. Kasam, Debora Sinner, Geereddy B. Reddy, Anjaparavanda P. Naren, Francis X. McCormack, Eric S. White, Anil G. Jegga, Satish K. Madala
Decreased noradrenergic excitation of hypoglossal motoneurons during sleep causing hypotonia of pharyngeal dilator muscles is a major contributor to the pathogenesis of obstructive sleep apnea (OSA), a widespread disease for which treatment options are limited. Previous OSA drug candidates targeting various excitatory/inhibitory receptors on hypoglossal motoneurons have proved unviable in reactivating these neurons, particularly during rapid-eye-movement (REM) sleep. To identify a viable drug target, we show that the repurposed α2-adrenergic antagonist yohimbine potently reversed the depressant effect of REM sleep on baseline hypoglossal motoneuron activity (a first-line motor defense against OSA) in rats. Remarkably, yohimbine also restored the obstructive apnea–induced long-term facilitation of hypoglossal motoneuron activity (hLTF), a much-neglected form of noradrenergic-dependent neuroplasticity that could provide a second-line motor defense against OSA but was also depressed during REM sleep. Corroborating immunohistologic, optogenetic, and pharmacologic evidence confirmed that yohimbine’s beneficial effects on baseline hypoglossal motoneuron activity and hLTF were mediated mainly through activation of pontine A7 and A5 noradrenergic neurons. Our results suggest a 2-tier (impaired first- and second-line motor defense) mechanism of noradrenergic-dependent pathogenesis of OSA and a promising pharmacotherapy for rescuing both these intrinsic defenses against OSA through disinhibition of A7 and A5 neurons by α2-adrenergic blockade.
Gang Song, Chi-Sang Poon
Excessive ROS promote allergic asthma, a condition characterized by airway inflammation, eosinophilic inflammation, and increased airway hyperreactivity (AHR). The mechanisms by which airway ROS are increased and the relationship between increased airway ROS and disease phenotypes are incompletely defined. Mitochondria are an important source of cellular ROS production, and our group discovered that Ca2+/calmodulin-dependent protein kinase II (CaMKII) is present in mitochondria and activated by oxidation. Furthermore, mitochondrial-targeted antioxidant therapy reduced the severity of allergic asthma in a mouse model. Based on these findings, we developed a mouse model of CaMKII inhibition targeted to mitochondria in airway epithelium. We challenged these mice with OVA or
Sara C. Sebag, Olha M. Koval, John D. Paschke, Christopher J. Winters, Omar A. Jaffer, Ryszard Dworski, Fayyaz S. Sutterwala, Mark E. Anderson, Isabella M. Grumbach
Idiopathic pulmonary fibrosis (IPF) is a fatal progressive fibrotic lung disease characterized by the presence of invasive myofibroblasts in the lung. Currently, there are only two FDA-approved drugs (pirfenidone and nintedanib) for the treatment of IPF. There are no defined criteria to guide specific drug therapy. New methodologies are needed not only to predict personalized drug therapy, but also to screen novel molecules that are on the horizon for treatment of IPF. We have developed a model system that exploits the invasive phenotype of IPF lung tissue. This ex vivo 3D model uses lung tissue from patients to develop pulmospheres. Pulmospheres are 3D spheroids composed of cells derived exclusively from primary lung biopsies and inclusive of lung cell types reflective of those in situ, in the patient. We tested the pulmospheres of 20 subjects with IPF and 9 control subjects to evaluate the responsiveness of individual patients to antifibrotic drugs. Clinical parameters and outcomes were also followed in the same patients. Our results suggest that pulmospheres simulate the microenvironment in the lung and serve as a personalized and predictive model for assessing responsiveness to antifibrotic drugs in patients with IPF.
Ranu Surolia, Fu Jun Li, Zheng Wang, Huashi Li, Gang Liu, Yong Zhou, Tracy Luckhardt, Sejong Bae, Rui-ming Liu, Sunad Rangarajan, Joao de Andrade, Victor J. Thannickal, Veena B. Antony
Oxidation of calmodulin-dependent protein kinase II (ox-CaMKII) by ROS has been associated with asthma. However, the contribution of ox-CaMKII to the development of asthma remains to be fully characterized. Here, we tested the effect of ox-CaMKII on IgE-mediated mast cell activation in an allergen-induced mouse model of asthma using oxidant-resistant CaMKII MMVVδ knockin (MMVVδ) mice. Compared with WT mice, the allergen-challenged MMVVδ mice displayed less airway hyperresponsiveness (AHR) and inflammation. These MMVVδ mice exhibited reduced levels of ROS and diminished recruitment of mast cells to the lungs. OVA-activated bone marrow–derived mast cells (BMMCs) from MMVVδ mice showed a significant inhibition of ROS and ox-CaMKII expression. ROS generation was dependent on intracellular Ca2+ concentration in BMMCs. Importantly, OVA-activated MMVVδ BMMCs had suppressed degranulation, histamine release, leukotriene C4, and IL-13 expression. Adoptive transfer of WT, but not MMVVδ, BMMCs, reversed the alleviated AHR and inflammation in allergen-challenged MMVVδ mice. The CaMKII inhibitor KN-93 significantly suppressed IgE-mediated mast cell activation and asthma. These studies support a critical but previously unrecognized role of ox-CaMKII in mast cells that promotes asthma and suggest that therapies to reduce ox-CaMKII may be a novel approach for asthma.
Jingjing Qu, Danh C. Do, Yufeng Zhou, Elizabeth Luczak, Wayne Mitzner, Mark E. Anderson, Peisong Gao
Maladaptive epithelial repair from chronic injury is a common feature in fibrotic diseases, which in turn activates a pathogenic fibroblast response that produces excessive matrix deposition. Dysregulated microRNAs (miRs) can regulate expression of multiple genes and fundamentally alter cellular phenotypes during fibrosis. Although several miRs have been shown to be associated with lung fibrosis, the mechanisms by which miRs modulate epithelial behavior in lung fibrosis are lacking. Here, we identified miR-323a-3p to be downregulated in the epithelium of lungs with bronchiolitis obliterans syndrome (BOS) after lung transplantation, idiopathic pulmonary fibrosis (IPF), and murine bleomycin-induced fibrosis. Antagomirs for miR-323a-3p augment, and mimics suppress, murine lung fibrosis after bleomycin injury, indicating that this miR may govern profibrotic signals. We demonstrate that miR-323a-3p attenuates TGF-α and TGF-β signaling by directly targeting key adaptors in these important fibrogenic pathways. Moreover, miR-323a-3p lowers caspase-3 expression, thereby limiting programmed cell death from inducers of apoptosis and ER stress. Finally, we find that epithelial expression of miR-323a-3p modulates inhibitory crosstalk with fibroblasts. These studies demonstrate that miR-323a-3p has a central role in lung fibrosis that spans across murine and human disease, and downregulated expression by the lung epithelium releases inhibition of various profibrotic pathways to promote fibroproliferation.
Lingyin Ge, David M. Habiel, Phil M. Hansbro, Richard Y. Kim, Sina A. Gharib, Jeffery D. Edelman, Melanie Königshoff, Tanyalak Parimon, Rena Brauer, Ying Huang, Jenieke Allen, Dianhua Jiang, Adrianne A. Kurkciyan, Takako Mizuno, Barry R. Stripp, Paul W. Noble, Cory M. Hogaboam, Peter Chen
The epigenome provides a substrate through which environmental exposures can exert their effects on gene expression and disease risk, but the relative importance of epigenetic variation on human disease onset and progression is poorly characterized. Asthma is a heterogeneous disease of the airways, for which both onset and clinical course result from interactions between host genotype and environmental exposures, yet little is known about the molecular mechanisms for these interactions. We assessed genome-wide DNA methylation using the Infinium Human Methylation 450K Bead Chip and characterized the transcriptome by RNA sequencing in primary airway epithelial cells from 74 asthmatic and 41 nonasthmatic adults. Asthma status was based on doctor’s diagnosis and current medication use. Genotyping was performed using various Illumina platforms. Our study revealed a regulatory locus on chromosome 17q12-21 associated with asthma risk and epigenetic signatures of specific asthma endotypes and molecular networks. Overall, these data support a central role for DNA methylation in lung cells, which promotes distinct molecular pathways of asthma pathogenesis and modulates the effects of genetic variation on disease risk and clinical heterogeneity.
Jessie Nicodemus-Johnson, Rachel A. Myers, Noburu J. Sakabe, Debora R. Sobreira, Douglas K. Hogarth, Edward T. Naureckas, Anne I. Sperling, Julian Solway, Steven R. White, Marcelo A. Nobrega, Dan L. Nicolae, Yoav Gilad, Carole Ober
Chronic inflammation with mucous metaplasia and airway remodeling are hallmarks of allergic asthma, and these outcomes have been associated with enhanced expression and activation of EGFR signaling. Here, we demonstrate enhanced expression of EGFR ligands such as amphiregulin as well as constitutive EGFR activation in cultured nasal epithelial cells from asthmatic subjects compared with nonasthmatic controls and in lung tissues of mice during house dust mite–induced (HDM-induced) allergic inflammation. EGFR activation was associated with cysteine oxidation within EGFR and the nonreceptor tyrosine kinase Src, and both amphiregulin production and oxidative EGFR activation were diminished by pharmacologic or genetic inhibition of the epithelial NADPH oxidase dual oxidase 1 (DUOX1). DUOX1 deficiency also attenuated several EGFR-dependent features of HDM-induced allergic airway inflammation, including neutrophilic inflammation, type 2 cytokine production (IL-33, IL-13), mucous metaplasia, subepithelial fibrosis, and central airway resistance. Moreover, targeted inhibition of airway DUOX1 in mice with previously established HDM-induced allergic inflammation, by intratracheal administration of DUOX1-targeted siRNA or pharmacological NADPH oxidase inhibitors, reversed most of these outcomes. Our findings indicate an important function for DUOX1 in allergic inflammation related to persistent EGFR activation and suggest that DUOX1 targeting may represent an attractive strategy in asthma management.
Aida Habibovic, Milena Hristova, David E. Heppner, Karamatullah Danyal, Jennifer L. Ather, Yvonne M.W. Janssen-Heininger, Charles G. Irvin, Matthew E. Poynter, Lennart K. Lundblad, Anne E. Dixon, Miklos Geiszt, Albert van der Vliet
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