Pulmonology
Abstract
Preclinical studies suggest beneficial effects of GLP-1 agonists in pulmonary arterial hypertension (PAH). This first-in-disease study evaluated acute hemodynamic effects of GLP-1 agonist, exenatide administered i.v. in patients with idiopathic PAH and CTEPH as well as in a PAH rodent model. Seventeen patients (9 idiopathic PAH) received an exenatide infusion during right heart catheterization, which included multisite sampling for circulating metabolites. Acute effects of exenatide were also assessed by cardiac magnetic resonance imaging in monocrotaline (MCT) PAH and control rats. In the clinical study, exenatide was well tolerated, reduced mean pulmonary artery pressure (45 ± 15 mmHg versus 40 ± 18 mmHg), and improved cardiac index (2.1 ± 0.6 L/min versus 2.4 ± 0.9 L/min/m2) and pulmonary vascular resistance (7.8 ± 8.0 WU versus 5.9 ± 5.0 WU) across all patients. Right ventricular (RV) contractility and afterload improved in a subset of patients undergoing pressure-volume measurements. In an exploratory metabolomics analysis, 47 metabolite levels changed after exenatide infusion, predominantly in free fatty acid pathways. Six metabolites with prognostic relevance in PAH within myocardial glycolytic and lipid oxidation pathways were also altered after exenatide. In MCT rats, exenatide improved RV stroke-volume, RV ejection fraction, and RV-arterial coupling. These findings support the further evaluation of exenatide within chronic studies as a potentially novel pulmonary vasodilator therapy.
Authors
Chinthaka B. Samaranayake, Marili Niglas, Nicoleta Baxan, Alexander Kempny, Ali Ashek, Michael Gatzoulis, Laura C. Price, Konstantinos Dimopoulos, Martin R. Wilkins, Stephen Wort, Christopher J. Rhodes, Lan Zhao, Colm McCabe
Abstract
Acute kidney injury (AKI) is a common and fatal complication of severe pneumonia, yet the mechanisms linking pulmonary inflammation to remote kidney injury remain poorly understood. Multicenter cohort data (n = 300) revealed that the incidence of severe pneumonia–associated AKI (SP-AKI) was 53.6%, with a mortality rate of 24.2%. SP-AKI was associated with elevated circulating levels of HMGB1, NETs, and IL-33. Murine experiments demonstrated that alveolar HMGB1 triggers the formation of IL-33–enriched NETs, which migrate to the kidney and activate tubular ST2/NF-κB signaling, driving inflammation and apoptosis. Genetic knockout of IL-33, ST2, or the NET-forming key enzyme PAD4, as well as pharmacological inhibition of HMGB1, IL-33, or NETs, all attenuated lung and kidney injury. Exogenous HMGB1 amplified NET-mediated IL-33 release, establishing a self-sustaining HMGB1/NET/IL-33 feed-forward loop. PAD4 deficiency completely blocked NET generation and disrupted HMGB1/IL-33 signaling. This study identified and validated a damage-associated molecular pattern–driven (DAMP-driven) HMGB1/NET/IL-33 signaling axis that mediates remote kidney injury in SP-AKI, redefining NETs from local effectors to cross-organ pathogenic carriers, thereby providing potential DAMP-targeted therapeutic avenues for SP-AKI.
Authors
Mengqing Ma, Hao Zhang, Weijuan Deng, Xia Du, Mengxing Chen, Dawei Chen, Binbin Pan, Zhaowei Wang, Ting Chen, Caimei Chen, Xin Wan, Changchun Cao
Abstract
Idiopathic pulmonary fibrosis (IPF) is characterized by parenchymal scarring reflecting an imbalance between collagen deposition by myofibroblasts (MFs) and its turnover. Although collagen clearance is essential for fibrosis resolution, this process and its potential for therapeutic modulation in IPF are poorly understood. Here we evaluated internalization of degraded collagen and the role of its requisite endocytic receptor mannose receptor C-type 2 (MRC2), in lung tissue and MFs from IPF patients and bleomycin-injured mice. Fibrotic human and murine lung tissue exhibited an accumulation of degraded collagen, highlighting a failure of its clearance. MFs from fibrotic lung demonstrated a reduced capacity to internalize extracellular degraded collagen, with a concomitant reduction in MRC2 expression and endolysosomal activity. Both diminished collagen uptake and MRC2 expression recovered to baseline levels during spontaneous resolution of bleomycin fibrosis. In vitro treatment of IPF or TGF-β-elicited MFs with a variety of mechanistically distinct agents known to effect phenotypic dedifferentiation restored defective collagen internalization. Although enhanced uptake was MRC2-dependent, it involved increased endolysosomal activity rather than increased MRC2 expression. These results implicate defective MRC2-dependent collagen internalization and endolysosomal function in MFs as important factors contributing to fibrosis that may be therapeutically targeted to promote resolution.
Authors
Natalie M. Walker, Sean M. Fortier, Jennifer Speth, Steven K. Huang, Sergey Gutor, Timothy S. Blackwell, Marc Peters-Golden
Abstract
The lungs have a remarkable capacity to undergo homoeostatic repair and regeneration after injury, which often occurs in patients with acute respiratory distress syndrome (ARDS) and in the single-dose bleomycin mouse model. Fibroblasts are critical mediators of fibrotic disease and RNA sequencing has identified significant heterogeneity within pulmonary fibroblast populations. However, the contribution of distinct fibroblast subsets to the repair process has been understudied compared to their role in fibrosis initiation and progression. Therefore, we sought to define the transcriptional landscape of three phenotypically-defined fibroblast subsets that occupy discrete spatial locations in naïve lungs. Using TdTomato-lineage tracing approaches, we identified and interrogated collagen1a1+ (Col1a1) fibroblasts, perilipin 2+ (Plin2) alveolar fibroblasts, and a-smooth muscle actin+ (Acta2) myofibroblasts during fibrosis development and resolution after single-dose bleomycin. Quantification of fibroblast numbers showed that all three subsets expand during fibrosis and contract towards naïve levels with resolution. Principal component and gene-set enrichment analyses indicated that each subset undergoes major transcriptomic shifts during fibrosis development, converging on a similar pro-fibrotic transcriptional profile. However, during resolution, Plin2+ and Acta2+ fibroblasts revert towards a pre-fibrotic transcriptional state, whereas Col1a1+ fibroblasts acquire a distinct program that suggests suggesting an active role in mediating the repair processes.
Authors
Daniel G. Foster, Nomin Javkhlan, Bart P. Black, Brian E. Vestal, David W.H. Riches, Elizabeth F. Redente
Abstract
CHI3L1, a chitinase-like protein, is implicated in pulmonary fibrosis, yet its mechanisms incompletely understood. In this study, we demonstrated that CHI3L1 coordinates profibrotic macrophage activation and invasive myofibroblast differentiation, and their crosstalk. In vitro, CHI3L1 drove M2-like macrophage polarization as evidenced by increased CD163, CD206, and PD-L1, and amplified TGF-β1-induced fibroblast responses, including myofibroblast transformation, migration, and invasion. Mechanistically, CHI3L1 enhanced TGF-β1 signaling through SMAD, AKT, and ERK pathways, and PD-L1 was required for CHI3L1/TGF-β1-driven myofibroblast transformation. Co-culture studies further demonstrated the ability of CHI3L1 to induce profibrotic macrophage activation that enhanced myofibroblast transformation mediated via a CD44–PD-L1 axis. In vivo, following bleomycin challenge, CHI3L1 transgenic mice exhibited increased PD-L1+ M2 macrophages, PD-L1+/PDGFRα+ fibroblasts, and PD-1+ immune cells compared with wild-type controls. Therapeutically, combined anti-CHI3L1 and anti-PD-1 antibodies, as well as a bispecific anti-CHI3L1-anti-PD-1 antibody, produced greater anti-fibrotic efficacy than monotherapy. These findings demonstrate crosstalk between CHI3L1 and the PD-1/PD-L1 pathway that promotes profibrotic macrophage activation and invasive fibroblast differentiation and support dual targeting of CHI3L1 and PD-1/PD-L1 as a promising therapeutic strategy for pulmonary fibrosis.
Authors
Han-Seok Jeong, Takayuki Sadanaga, Joyce H. Lee, Suchitra Kamle, Bing Ma, Yang Zhou, Sung Jae Shin, Jack A. Elias, Chun Geun Lee
Abstract
Heterozygous TBX4 variants are the second most common genetic cause of pediatric pulmonary hypertension (PH), yet mechanisms underlying TBX4-related lung disease remain poorly understood. This study developed a lung mesenchyme-specific Tbx4 loss-of-function (Tbx4cKO) mouse model that bypasses embryonic lethality to investigate this condition. Adult Tbx4cKO mice demonstrated significantly impaired pulmonary flow acceleration consistent with PH. Three-dimensional analysis of embryonic lungs revealed reduced lobe volumes and decreased distance between pleural edges and muscularized vessels. In adult Tbx4cKO lungs, we identified extensive vascular remodeling characterized by medial thickening and the extension of muscularized arteries into normally non-muscularized subpleural parenchymal zones. Contrary to previous reports suggesting vascular simplification, three-dimensional analysis demonstrated an elaborated pulmonary artery (PA) tree in addition to pathologic wall muscularization. Depletion of a single Tbx5 allele in addition to both Tbx4 alleles exacerbated histologic phenotypes with worsened right ventricular dilation. This model also demonstrated dysregulated airway smooth muscle patterning and prominent subpleural smooth muscle bands, similar to those in human TBX4 syndrome. We identify TBX4 as a critical regulator of smooth muscle differentiation and patterning across multiple lung compartments. Our model recapitulates key features of human TBX4 syndrome and identifies dysregulated smooth muscle differentiation as a potential future therapeutic target.
Authors
Lea C. Steffes, Kaylie A. Chiles, Sehar R. Masud, Aleen Rahman, Madeline Dawson, Csaba Galambos, Maya E. Kumar, Ripla Arora
Abstract
Survival after lung transplantation is limited by chronic, progressive graft failure, termed chronic lung allograft dysfunction (CLAD). Graft-resident mesenchymal cells (MCs) drive CLAD pathogenesis and exhibit stable dysregulated signaling, yet the transcriptomic and epigenomic drivers underlying this fibrogenic transformation remain elusive. We used single-cell multi-omic profiling to characterize gene expression and chromatin accessibility in MCs isolated from lavage fluid of lung transplant recipients with and without CLAD, collected early post-transplantation or after disease onset. MCs obtained after CLAD onset demonstrated a distinct transcriptomic signature compared with non-CLAD controls, enabling classification of disease status at the single-cell level with > 98% accuracy using signature genes. Chromatin accessibility analyses identified enrichment of CCAAT-enhancer-binding protein family transcription factors, specifically CEBPD, in CLAD MCs. Early post-transplant MCs showed minimal accessibility differences, suggesting that CEBPD-associated regulatory changes emerge over time. Integration analyses identified eight MC states and a CLAD-specific shift towards a fibrotic state. CEBPD, SOX4, and FOXP2 were identified as putative regulators of this state with substantial overlap in predicted targets. Targeting CEBPD reversed fibrotic phenotypes of CLAD MCs (decreased ECM expression, contractility, proliferation, and migration). Together, these data provide insights into transcriptomic and epigenomic changes in post-transplant MCs, nominating biomarkers and therapeutic targets.
Authors
Lu Lu, A. Patrick McLinden, Natalie M. Walker, Ragini Vittal, Yichen Wang, Fatemeh Fattahi, Stephen T. Russell, Michael P. Combs, Joshua D. Welch, Vibha N. Lama
Abstract
Pulmonary fibrosis is frequently accompanied by pulmonary hypertension, which can occur disproportionate to the extent of fibrosis, suggesting a fibrosis-independent vascular remodeling process. Here, we demonstrated that plasma growth differentiation factor 15 (GDF15) is elevated across diverse fibrotic lung disease subtypes and correlates with markers of elevated right heart pressures, but not pulmonary function indices, indicating a possible link to endothelial cell dysfunction. To investigate the import of endothelial GDF15 as a modifier of lung fibrosis pathogenesis, we generated endothelial cell-specific Gdf15 knockout mice, which showed protection from bleomycin-induced lung injury and fibrosis, with preserved lung function. RNA sequencing of human pulmonary microvascular endothelial cells revealed altered expression of barrier-regulatory genes in GDF15-deficient endothelial cells compared to controls. Functional studies confirmed that GDF15 knockdown attenuates thrombin-induced barrier disruption by reducing cytosolic Ca2+ responses. Together, these findings implicate endothelial GDF15 as a modifier of vascular permeability and Ca2+ signaling, and a contributor to lung injury and fibrosis.
Authors
Kristen Raffensperger, Marta Bueno, Brian J. Philips, Megan Miller, Máté Katona, Shuai Yuan, Adriana Estrada-Bernal, Byron Chuan, Pavan Suresh, Stephanie Taiclet, Scott Hahn, Yingze Zhang, Jonathan K. Alder, Seyed Mehdi Nouraie, Daniel J. Kass, Oliver Eickelberg, Adam C. Straub
Abstract
Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial lung disease driven by aberrant fibroblast-to-myofibroblast differentiation, which requires metabolic reprogramming. Here, we identify alanine as an essential metabolite for myofibroblast differentiation. Transforming growth factor–β1 (TGF-β) increases intracellular alanine levels through enhanced synthesis and import in both normal and IPF lung fibroblasts. Alanine synthesis is primarily mediated by glutamate-pyruvate transaminase 2 (GPT2), whose expression is regulated by the glutamine–glutamate–α-ketoglutarate axis. Inhibition of GPT2 depletes alanine and suppresses TGF-β-induced α-SMA and COL1A1 expression, which are rescued by exogenous alanine. We also identify solute carrier family 38 member 2 (SLC38A2) as a transporter for both alanine and glutamine, upregulated by TGF-β or alanine deprivation. SLC38A2 and GPT2 form a coordinated regulatory axis sustaining intracellular alanine levels to support myofibroblast differentiation. Mechanistically, alanine deficiency impairs glycolytic flux and depletes tricarboxylic acid cycle intermediates, while alanine supplementation provides carbon and nitrogen for intracellular glutamate and proline biosynthesis, particularly under glutamine deprivation. Combined inhibition of alanine synthesis and uptake suppresses fibrogenic responses in fibroblasts and human precision-cut lung slices, highlighting dual metabolic targeting as a potential therapeutic strategy for fibrotic lung disease.
Authors
Fei Li, Niv Vigder, David R. Ziehr, Mari Kamiya, Hung N. Nguyen, Diana E. Ferreyra Faustino, Aseel H. Khalil, Hilaire C. Lam, Matthew L. Steinhauser, Edy Y. Kim, William M. Oldham
Abstract
Systemic sclerosis (SSc) is characterized by fibrosis and vasculopathy affecting the skin and internal organs, leading to multiorgan dysfunction. Injury of microvascular endothelial cells (ECs) in SSc impairs blood flow and causes tissue ischemia, leading to vascular complications such as Raynaud’s, digital ulcers, and pulmonary hypertension (PH). PH in SSc presents as group 1 pulmonary arterial hypertension or as group 3 PH related to hypoxia and interstitial lung disease (ILD), both major causes of mortality. Analysis of multiome data from SSc ILD-PH lungs inferred transcription factors regulating EC phenotype, including FOSL2. Overexpression of FOSL2 in transgenic mice (Fosl2tg) leads to vascular changes mirroring human SSc-PH, such as intimal thickening and fibrosis. scRNA-Seq analysis of altered EC gene expression in Fosl2tg mice showed strong overlap with altered EC gene expression in SSc-ILD-PH. Overlapping as well as discrete EC gene expression in Sugen/hypoxia- and hypoxia-treated mice suggested that FOSL2 regulates both hypoxia-dependent and -independent pathways in Fosl2tg mice and SSc-ILD-PH. A deep learning model, ChromBPNet, inferred increased AP-1 binding at base pair resolution in SSc-ILD-PH ECs, and binding to the same motifs was found upon FOSL2 overexpression in primary vascular ECs, highlighting FOSL2’s key role in driving the pathological changes seen in SSc-ILD-PH.
Authors
Rithika Behera, Yuechen Zhou, Peter H. Gerges, Jingyu Fan, Tracy Tabib, Alyxzandria M. Gaydosik, Mengqi Huang, Jishnu Das, Elena Pachera, Amela Hukara, Ying Tang, Florian Renoux, Miranda Tai, Oliver Distler, Gabriela Kania, Stephen Y. Chan, Harinder Singh, Eleanor Valenzi, Robert Lafyatis
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