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PubMed2025

IGF1R deficiency mitigates acute lung injury by promoting anti-inflammatory transcriptional profiles.

BACKGROUND: Acute lung injury (ALI), acute respiratory distress syndrome (ARDS) and COVID-19 are characterized by hyperinflammation, commonly referred to as "cytokine storm". The insulin-like growth factor (IGF) pathway, particularly the type 1 receptor (IGF1R), plays a critical role in lung homeostasis and has been implicated in the pathogenesis of pulmonary inflammatory diseases. In mice, widespread Igf1r deficiency attenuates lung inflammation and alveolar damage in bleomycin (BLM)-induced ALI. METHODS: We analyzed single-cell RNA sequencing datasets from lung tissue of COVID-19 cases and control donors as well as mouse lungs to determine Igf1r and IGF family expression across pulmonary cell types. Furthermore, we conducted bulk RNA sequencing on lungs from Igf1r-deficient mice three days after BLM or saline instillation, followed by differential expression and functional enrichment analyses. Findings were further tested through protein detection, assessment of DNA damage and methylation in lung tissues, and functional assays using Igf1r-deficient primary mouse embryonic fibroblasts (MEFs). RESULTS: IGF1R was broadly expressed across multiple cell types in both human and mouse lungs under normal and pathological conditions. Other IGF family members showed cell-type-specific expression, which was modulated by lung injury. Transcriptomic profiling revealed differentially expressed genes between BLM-challenged and control mouse lungs, detecting biological processes and signaling pathways involved in ALI pathobiology. Igf1r deficiency in BLM-challenged mice reversed a large fraction of the transcriptional changes triggered by BLM, including "cytokine storm"-related gene expression. Functional enrichment analysis additionally revealed significant modulation of pathways related to DNA damage, metabolic reprogramming, mitochondrial homeostasis, and epigenetic regulation. In vitro, Igf1r-deficient MEFs exhibited decreased mitochondrial respiration and glycolysis, protection against BLM-induced nuclear damage and mitochondrial accumulation, and decreased histone H3 acetylation. Moreover, Igf1r-deficient mouse lungs displayed increased global DNA methylation following BLM challenge. CONCLUSIONS: IGF1R is a key modulator of the inflammatory and molecular response to ALI pathogenesis. IGF1R deficiency dampens the "cytokine storm", modifies transcriptional and epigenetic profiles and promotes protective cellular responses. These findings highlight IGF1R signaling as a potential therapeutic target in ARDS and related lung injuries.

Animals↗
PubMed2026

Human airspace macrophage signatures are conserved during sterile lung injury and repair.

RATIONALE: Airspace macrophages (AM) are implicated in both persistent inflammation and tissue repair following acute lung injury. Distinct subsets of AM are associated with lung pathology in humans but whether unique AM signatures are specific to disease states or represent a conserved response to lung inflammation is unknown. OBJECTIVES: We sought to test the hypothesis that conserved subsets of inflammatory and reparative AM could be identified by transcriptional programing in a human model of self-resolving acute lung injury. METHODS: Fifteen subjects underwent bronchoscopic lavage (BAL) before and at a pre-assigned time point after endobronchial exposure to bacterial endotoxin. BAL cells were subjected to single cell RNA sequencing and longitudinal assessment of AM programing during resolution of inflammation and lung repair was performed. MEASUREMENTS AND MAIN RESULTS: We identify transcriptionally distinct subsets of tissue resident and recruited AM present at all time points, in all subjects. Two recruited AM populations increase following inflammation, one which aligns with classical monocytes (MoAM) and one with interstitial macrophages (IAM). AM subsets display unique patterns of gene expression throughout the time course. Comparison of subset-specific markers to those identified in disease states reveals that IAM express many so-called "pathogenic" markers during normal lung repair. CONCLUSIONS: By applying a uniform inflammatory stimulus to healthy adults and examining BAL cells obtained at precise time points thereafter, we construct a time-resolved kinetic of AM transcriptional programing during typical lung repair. Our data demonstrate that IAM share transcriptional similarity to AM identified in disease states and suggest they may reflect a conserved cellular response to tissue injury.

Journal Article↗
PubMed2026

Qingfei Dayuan granules alleviate the inflammatory response in lipopolysaccharide-induced acute lung injury mice by inhibiting the Nf-κB signaling pathway and regulating the complement pathway.

OBJECTIVES: The study aimed to explore the effects and mechanisms by which Qingfei Dayuan granules (QFDY) mitigate pulmonary inflammation in lipopolysaccharide (LPS)-induced acute lung injury (ALI). METHODS: We established an ALI mouse model by intraperitoneal injection of LPS. HE, Transmission electron microscopy, ELISA assay of inflammatory cytokines, and immunohistochemistry (IHC) were used to assess the degree of lung injury and inflammation. Utilizing network analysis and proteomics analysis, the potential targets and pathways of QFDY were identified. Western blot, IHC, and qRT-PCR analysis were used to evaluate the potential mechanism of QFDY. Additionally, the chemical composition of QFDY were performed using UPLC-MS/MS. KEY FINDINGS: QFDY reduced the pathologic changes and inflammatory cell infiltration in lung tissue inflammation. Network and proteomic analysis showed that the mechanism of QFDY protection against ALI is closely related to the Nuclear factor-kappa B (NF-κB) signaling pathway and complement pathway. Animal experiments showed that Qingfei Dayuan granules (QFDY) significantly reduced the levels of IL-1β, IL-6, TNF-α, and lung tissue F4/80-positive alveolar macrophages. Additionally, western blot and qRT-PCR analyses showed the inhibition of the NF-κB pathway. Notably, the levels of mannose-binding lectin (MBL2) were significantly increased, while complement C3a and complement C5a proteins were reduced in the QFDY group compared to the LPS group. CONCLUSIONS: QFDY suppressed the inflammation in LPS-induced ALI by inhibiting the NF-κB and complement pathway.

Animals↗
PubMed2026

Mechanism of Action of Hedyotis diffusa Extract in a Rat Model of Acute Lung Injury Based on Transcriptomic Analysis.

OBJECTIVE: This study established a rat model of lipopolysaccharide (LPS)-induced acute lung injury (ALI) to evaluate pathological damage, collagen deposition, inflammatory cytokine levels, and key gene/protein expression following Hedyotis diffusa water extract (HDWE) intervention. Combined with ultra-high-performance liquid chromatography-quadrupole Orbitrap high-resolution mass spectrometry (UHPLC-Q-Orbitrap HRMS), transcriptomic analysis, and molecular simulation, this study identified the bioactive components of HDWE, evaluated their potential interactions with ALI-related targets, and explored the multi-omics-based protective mechanisms of HDWE. METHODS: Thirty-six Sprague-Dawley (SD) rats were randomly divided into six groups: Control group, ALI group, DXMS group, HDWE-L group (100 mg/kg), HDWE-M group (200 mg/kg), and HDWE-H group (300 mg/kg). Hematoxylin and eosin (H&E) and Masson's trichrome staining were used to evaluate lung pathological changes and collagen deposition. Enzyme-linked immunosorbent assay (ELISA) was used to measure serum tumor necrosis factor-α TNF-α interleukin-1β IL-1β, erleukin-6 (IL-6), and interleukin-10 (IL-10) levels. Transcriptomic analysis identified differentially expressed genes (DEGs), followed by Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), receiver operating characteristic (ROC), and immune infiltration analyses. Quantitative real-time polymerase chain reaction (qRT-PCR) detected the mRNA expression levels of SPHK1, RELA, and NFKBIA. Immunohistochemistry evaluated the expression of eight hub targets, including endothelin-1 (EDN1), sphingosine kinase 1 (SPHK1), intercellular adhesion molecule 1 (ICAM1), interleukin-17 (IL-17), prostaglandin-endoperoxide synthase 2 (PTGS2/COX-2), NF-κB p65 (encoded by RELA), WT1-associated protein (WTAP), and myeloperoxidase (MPO). UHPLC-Q-Orbitrap HRMS characterized HDWE constituents. Molecular docking analysis was performed between 22 compounds and eight hub targets, followed by 100 ns molecular dynamics simulations and molecular mechanics-Poisson-Boltzmann surface area (MM/PBSA) binding free energy calculations for five core targets. Compared with the control group, the ALI group showed increased levels of TNF-α (86%), IL-1β (107%), and IL-6 (66%), accompanied by a 43% reduction in IL-10 and a 300% increase in lung collagen deposition. All HDWE doses alleviated inflammatory responses, with medium-dose HDWE showing the most pronounced effects. Specifically, medium-dose HDWE increased IL-10 levels by 52% and reduced IL-6, TNF-α, and IL-1β levels by 18%, 22%, and 11%, respectively. Transcriptomic analysis identified 2512 DEGs between the control group and ALI groups, 832 exclusive DEGs between the ALI group and HDWE-M groups, and 876 overlapping DEGs enriched in TNF, IL-17, and NF-κB signaling pathways. The eight-hub-gene diagnostic model achieved an area under the curve (AUC) of 0.969. RELA, SPHK1, and four other hub genes showed positive correlations with Th1, Th17, and neutrophil infiltration. In the ALI group, SPHK1, RELA, and NFKBIA mRNA expression levels were 1.30-, 0.96-, and 0.71-fold of those in the control group, respectively. Compared with the ALI group, high-dose HDWE treatment and low-dose HDWE treatment reduced SPHK1 expression to 0.62- and 0.57-fold, respectively, and increased NFKBIA expression to 1.68- and 1.58-fold, respectively. High-dose HDWE treatment reduced RELA expression to 0.43-fold. The expression levels of inflammation-related proteins were increased in the ALI group and were reduced after HDWE treatment. Twenty-two HDWE components were identified, 16 of which met the docking criteria. Asperulosidic acid exhibited favorable predicted binding affinities with all eight targets, with calculated binding free energies of -14.74, -14.92, -17.58, -23.04, and -16.10 kcal/mol for MPO, IL-17, NF-κB p65, PTGS2/COX-2, and SPHK1, respectively. CONCLUSIONS: This study provides systematic in vivo pharmacodynamic and in silico component-target evidence regarding the protective effects of HDWE against LPS-induced ALI. HDWE treatment increased NFKBIA expression and reduced SPHK1, RELA, and multiple inflammatory protein levels, suggesting that HDWE may regulate the IL-17/NF-κB-associated inflammatory network, although direct causal relationships require further validation. Asperulosidic acid may represent a key bioactive component with broad target-binding potential. This study was limited by the use of an LPS-induced rat ALI model without gene knockout or target inhibitor validation; therefore, further functional experiments are required to confirm the proposed regulatory mechanisms.

Hedyotis diffusa↗
PubMed2026

Matrine Alleviates Sepsis-Induced Acute Lung Injury by Reinforcing NQO1/SLC7A11/GPX4-Associated Anti-Ferroptotic Defenses and Attenuating NF-κB-Driven Inflammation.

BACKGROUND: Sepsis triggers dysregulated systemic inflammation and multiple-organ dysfunction, with the lungs being particularly susceptible to injury. Sepsis-induced acute respiratory distress syndrome (ARDS) is mainly driven by TLR4/NF-κB-mediated hyperinflammation and alveolar macrophage activation. Matrine, a bioactive alkaloid derived from Sophora flavescens, has been reported to modulate redox homeostasis and ferroptosis-associated lipid peroxidation. However, the target-specific mechanisms underlying its effects on ferroptosis and inflammatory signaling in sepsis-induced acute lung injury (SALI) remain incompletely understood. PURPOSE: This study aimed to evaluate the therapeutic effects of matrine in a cecal ligation and puncture (CLP)-induced SALI model and to determine whether its protective effects involve reinforcement of NQO1/SLC7A11/GPX4-associated anti-ferroptotic defenses and suppression of NF-κB-driven inflammation. METHODS: We analyzed the single-cell RNA-sequencing (scRNA-seq) dataset GSE273924 to characterize CD45-enriched pulmonary immune-cell subsets in sham mice and mice with intratracheal Escherichia coli-induced pneumonia. Network pharmacology and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were performed to predict Kushen (KS)-related targets and pathways associated with SALI. Differential expression analysis and weighted gene co-expression network analysis (WGCNA) of GSE245013 were used to identify candidate targets. Matrine-NQO1 binding and intracellular target engagement were evaluated using molecular docking, molecular dynamics simulations, surface plasmon resonance (SPR), and the cellular thermal shift assay (CETSA). The therapeutic effects of matrine were assessed in mice with CLP-induced SALI and in lipopolysaccharide (LPS)-stimulated MH-S cells. Lung histopathology, inflammatory cytokine production, target protein expression, ferroptosis-associated indicators, and NF-κB activation were evaluated using molecular, biochemical, and histological assays. The functional contribution of NQO1 was further examined using the NQO1 inhibitor ES936. RESULTS: scRNA-seq analysis of GSE273924 revealed substantial remodeling of the CD45-enriched pulmonary immune-cell landscape in mice with intratracheal E. coli-induced pneumonia, including macrophage transcriptional programs associated with ferroptosis and inflammatory signaling. Integrated network pharmacology and bioinformatics analyses prioritized NQO1 as a candidate target of matrine and identified NF-κB signaling as a potentially relevant pathway. Molecular docking, molecular dynamics simulations, SPR, and CETSA supported matrine-NQO1 binding and intracellular target engagement. Functionally, matrine improved survival, attenuated lung injury, reinforced NQO1/SLC7A11/GPX4-associated anti-ferroptotic defenses, and suppressed NF-κB activation in CLP mice. Similar protective effects were observed in LPS-stimulated MH-S cells. ES936 partially attenuated the matrine-mediated improvements in cell viability, redox homeostasis, ferroptosis-associated indicators, and NF-κB p65 phosphorylation, supporting a functional contribution of NQO1 to the protective effects of matrine. CONCLUSION: Matrine alleviates SALI by reinforcing NQO1/SLC7A11/GPX4-associated anti-ferroptotic defenses and attenuating NF-κB-driven inflammation.

Animals↗
PubMed2026

MASLD Exacerbates Chronic Low-dose PM2.5-induced Lung Injury, Inflammation, and Fibrosis.

BACKGROUND/AIM: Fine particulate matter (PM2.5) and metabolic dysfunction-associated steatotic liver disease (MASLD) are independent risk factors for respiratory disease. However, the combined impact of chronic, low-dose PM2.5 exposure and Western diet (WD)-induced metabolic dysfunction on pulmonary health remains poorly understood. We investigated whether this metabolic state exacerbates PM2.5-driven pathologies using an environmentally relevant PM2.5 dosage (~50 μg/m3). MATERIALS AND METHODS: C57BL/6J mice were fed a WD or normal diet (ND) for 28 weeks and concurrently received intratracheal instillations of PM2.5 (0.5 mg/kg diesel particulate matter) or vehicle three times per week. The MASLD phenotype was confirmed through metabolic and histological analyses. Pulmonary injury, fibrosis, and inflammation were assessed via histology (hematoxylin and eosin, and Masson's trichrome staining) and cytokine quantification in both bronchoalveolar lavage fluid using Luminex multiplex assay and lung tissue using enzyme-linked immunosorbent assay and quantitative polymerase chain reaction. RESULTS: The WD successfully induced MASLD characterized by weight gain, hepatic steatosis, and dyslipidemia. While PM2.5 exposure did not significantly worsen the primary features of MASLD, its combination with a WD markedly exacerbated pulmonary injury and fibrosis compared to PM2.5 exposure alone. This exacerbation was driven by a surge in pro-inflammatory chemokines, including C-X-C motif chemokine ligands 1 and 2 (CXCL1 and CXCL2), and C-C motif chemokine ligand 5 (CCL5), confirmed by Luminex analysis of lavage fluid and mRNA/protein quantification in lung tissue. CONCLUSION: Diet-induced metabolic dysfunction primes the lung for a hyper-inflammatory response to chronic PM2.5 exposure. These findings identify individuals with MASLD as a population with heightened susceptibility to air pollution-related respiratory diseases and underscore the critical interplay between metabolic health and environmental toxicology.

Animals↗
PubMed2025

Deep inflations maintain surfactant function and alveolar fluid balance in lungs with reduced surfactant protein B levels during mechanical ventilation.

Surfactant protein B (SP-B) is essential for surface tension reducing function of pulmonary surfactant and alveolar unfolding processes during inspiration. SP-B is reduced early in acute lung injury. Hence, we hypothesize that 1) reduced SP-B expression increases susceptibility to ventilation-induced lung injury (VILI), and 2) deep inflations (DI) are protective against VILI. Conditional SP-B knockout mice were randomized into OFF (reduced SP-B) and ON groups (normal SP-B) and subjected to mechanical ventilation at zero end-expiratory pressure. Over 4 h of ventilation, either 4 or 16 DI were administered. Lung mechanics were recorded, and pulmonary structure was quantified by design-based stereology. Inflammatory cells and bulk RNA sequencing were measured in bronchoalveolar lavage (BAL) and tissue, respectively. No differences in inflammatory cells in BAL were detected between ON and OFF groups. During ventilation, alveolar derecruitment-related increase in elastance was most pronounced in OFF-4DI but reversible by DI so that lung mechanics did not worsen. Finally, volumes of the alveolar liquid lining layer and the intracellular surfactant were largest, whereas the surface area of the apical plasma membrane of type II pneumocytes was smallest in OFF-4DI, suggesting impaired surfactant secretion. A higher frequency of DI prevented these abnormalities. Electron microscopy revealed disorganized tight junctions between alveolar epithelial cells in OFF-4DI, which was linked with decreased expression of genes relevant to the apical junctional complex. Reduced SP-B resulted in a progressive increase in surface tension and a disturbed fluid balance without triggering definite VILI. Maintenance of residual surfactant function is highly dependent on DI in conditions of reduced SP-B levels.NEW & NOTEWORTHY Surfactant protein B (SP-B) is critical for efficient surfactant function in the lung. Reduced SP-B levels occur at an early stage of acute lung injury and impair alveolar unfolding. In this study, we demonstrate that mechanical ventilation of lungs with reduced SP-B levels does not trigger ventilation-induced lung injury but results in disbalance of alveolar fluid volume and increase in surface tension due to failure of surfactant maintenance. Deep inflations prevent these ventilation-induced effects.

Animals↗
PubMed2026

Necroptosis in alveolar epithelium orchestrates lung ischemia-reperfusion injury: a multi-omics study.

BACKGROUND: Lung ischemia-reperfusion injury (LIRI) is a leading cause of early morbidity and mortality following lung transplantation and other cardiopulmonary procedures. It is characterized by acute sterile inflammation driven by regulated cell death (RCD). While various RCD modalities, including apoptosis, necroptosis, pyroptosis, and ferroptosis, have been implicated in lung injury, their relative contributions and distinct activation patterns in LIRI remain poorly defined. METHODS: We employed an integrated multi-omics approach combining transcriptomics and proteomics with histological and functional validations in a murine hilar clamping model of LIRI. Key findings were further corroborated using single-cell RNA sequencing (scRNA-seq) data from human lung transplant recipients. The functional role of necroptosis was validated using pharmacological inhibitors (Nec-1, GSK'872) and Mlkl-deficient (Mlkl-/-) mice. RESULTS: LIRI triggered acute, time-dependent lung injury peaking within 24 h of reperfusion. Although transcriptomic profiling suggested broad activation of multiple RCD pathways, proteomic and biochemical analyses revealed a distinct landscape in our experimental setting: markers of apoptosis, pyroptosis, and ferroptosis were either downregulated or showed no significant positive correlation with injury severity and inflammatory peaks. In contrast, the necroptotic pathway emerged as a highly activated modality. Specifically, necroptosis, marked by phosphorylated RIPK1, RIPK3, and MLKL, was localized primarily in alveolar epithelial cells, correlated strongly with cytokine release and histological lung injury, and preceded the inflammatory response. Pharmacological inhibition or genetic ablation of necroptosis significantly attenuated tissue damage and inflammation. This pronounced necroptotic signature appeared distinct from the broad multi-pathway activation observed in lipopolysaccharide (LPS)-induced lung injury. Translational analysis of human scRNA-seq data further confirmed the selective upregulation of necroptosis signatures in alveolar type 2 (AT2) cells following lung transplantation. CONCLUSION: Our multi-omics analysis identifies necroptosis, particularly in alveolar epithelial cells, as a critical driver of sterile inflammation and tissue injury in the early phase of LIRI. Targeting alveolar epithelial necroptosis may represent a precise and promising therapeutic strategy for lung transplantation and ischemia-reperfusion-associated pulmonary disorders.

Animals↗
PubMed2026

Developing Highly Effective Nanoparticle mRNA Therapeutic for Pediatric Acute Respiratory Distress Syndrome.

Sepsis-induced pediatric acute lung injury (ALI) and pediatric acute respiratory distress syndrome (PARDS) are life-threatening conditions with high mortality rates and no current cure. Most ALI/ARDS studies focus on adults, albeit the pediatric population has unique challenges often underrepresented. ALI/PARDS severely impacts pulmonary endothelial cells (ECs), causing endothelial dysfunction and vascular leakage. FOXF1 is a transcription factor critical for lung repair after injury, representing a viable target for ALI/PARDS. This study developed and tested a novel nanoparticle system for precise delivery of FOXF1 mRNA into lung ECs to reduce endothelial damage and improve lung function in mouse model of PARDS. Systemic inflammatory response was induced in neonatal mice after intraperitoneal administration of lipopolysaccharide (LPS). Specifically designed nanoparticles (NP) were used to intravenously deliver stabilized FOXF1 mRNA (FOXF1 NP) after LPS injury to restore FOXF1 expression in injured lung endothelial cells. FOXF1 NP selectively targeted pulmonary ECs without affecting other cell types or organs. FOXF1 NP treatment reduced vascular leakage, enhanced endothelial barrier function, and improved survival of neonatal mice after injury. FOXF1 NP decreased EC apoptosis by restoring the expression of BCL2, an anti-apoptotic FOXF1 target gene. Nanoparticle-based rescue of lung ECs has promise for future treatments of human ALI/PARDS.

endothelial cells↗
PubMed2026

miR-197 Targets NLRP3 3' UTR and Correlates with NLRP3/Caspase-1/IL-18 Signaling in Hyperoxia-Stimulated Neonatal BPD Mouse Model.

Reduced circulating miR-197 was previously observed in preterm infants who later developed bronchopulmonary dysplasia (BPD), but its relationship with NLRP3 inflammasome signaling remains unclear. This study examined miR-197 expression, NLRP3 inflammasome-related markers, and the interaction between miR-197 and the NLRP3 3' UTR in a neonatal hyperoxia model. Neonatal C57BL/6J mice were exposed to 60% oxygen or room air from birth, and lung tissues were collected on postnatal days 1, 7, 14, and 21. Lung injury and alveolar development were assessed by histology, radial alveolar count, mean linear intercept, and lung wet-to-dry ratio. miR-197 and NLRP3 expression and NF-κB-, caspase-1-, and IL-18-related proteins were evaluated by RT-qPCR and Western blotting. A dual-luciferase reporter assay in MLE12 cells tested the interaction between miR-197 and the NLRP3 3' UTR. Hyperoxia increased lung wet-to-dry ratios and mean linear intercept, reduced radial alveolar count, and caused progressive alveolar simplification. miR-197 expression decreased, whereas NLRP3 mRNA increased, at all examined time points; NLRP3 protein and inflammasome-related protein changes were most evident from postnatal day 7 onward. The miR-197 mimic reduced luciferase activity in the wild-type but not mutant NLRP3 3' UTR reporter. These findings show that neonatal hyperoxia is associated with reduced miR-197 and increased NLRP3/inflammasome-related signaling and support a sequence-specific interaction between miR-197 and the NLRP3 3' UTR, although causal regulation in vivo requires further investigation.

Animals↗
PubMed2025

Proteomics and Phosphoproteomics Characteristics of the Rhesus Macaque Lung Infected With Original SARS-CoV-2, Delta, and Omicron Variants.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strains mutate rapidly, making it crucial to study their molecular mechanisms for swift vaccine and drug development. Here, we utilized host lung proteomic and phosphoproteomic profiling to investigate the underlying pathology caused by the variants. Lung tissues infected with wild-type GD108, Delta, or Omicron BA.1 variants showed overexpression of proteins and phosphoproteins linked to the innate immune pathway, particularly in the Omicron group, with high activation of NOD-receptor and RIG-I like receptor signaling pathways. Protein-protein interaction (PPI) analysis revealed six key proteins, including antiviral innate immune response receptor RIG-I (DDX58), and five interferon-related proteins (IFIT2, ISG15, MX1, STAT1, and EIF2AK2), highlighting the importance of the innate immune response in combating all three variants. Kinase prediction analysis suggested that six kinases (DAPK1, DAPK2, DAPK3, PRACK, TTK, and MAP2K2), potentially inhibited by Fostamatinib, were activated across all three variants, and might be potential drug targets, pending further verification. Omicron infection, compared to other mutants, significantly disrupted proteins related to pulmonary structural support, like integrin and collagens, and inhibited efferocytosis, reducing the host's ability to eliminate the pathogen. These findings suggest that innate immune activation and structural disruption may contribute to Omicron-related pathology, potentially being useful for research into the molecular mechanisms underlying lung injury from SARS-CoV-2 variants.

Animals↗
PubMed2025

Dysregulation of lung epithelial cell homeostasis and immunity contributes to Middle East respiratory syndrome coronavirus disease severity.

Coronaviruses (CoV) emerge suddenly from animal reservoirs to cause novel diseases in new hosts. Discovered in 2012, the Middle East respiratory syndrome coronavirus (MERS-CoV) is endemic in camels in the Middle East and is continually causing local outbreaks and epidemics. While all three newly emerging human CoVs from the past 20 years (SARS-CoV, SARS-CoV-2, and MERS-CoV) cause respiratory disease, each CoV has unique host interactions that drive differential pathogeneses. To better understand the virus and host interactions driving lethal MERS-CoV infection, we performed a longitudinal multi-omics analysis of sublethal and lethal MERS-CoV infection in mice. Significant differences were observed in body weight loss, virus titers, and acute lung injury among lethal and sub-lethal virus doses. Virus-induced apoptosis of type I and II alveolar epithelial cells suggests that loss or dysregulation of these key cell populations was a major driver of severe disease. Omics analysis suggested differential pathogenesis was multi-factorial with clear differences among innate and adaptive immune pathways as well as those that regulate lung epithelial homeostasis. Infection of mice lacking functional T and B cells showed that adaptive immunity was important in controlling viral replication but also increased pathogenesis. In summary, we provide a high-resolution host response atlas for MERS-CoV infection and disease severity. Multi-omics studies of viral pathogenesis offer a unique opportunity to not only better understand the molecular mechanisms of disease but also to identify genes and pathways that can be exploited for therapeutic intervention all of which is important for our future pandemic preparedness.IMPORTANCEEmerging coronaviruses like SARS-CoV, SARS-CoV-2, and MERS-CoV cause a range of disease outcomes in humans from an asymptomatic, moderate, and severe respiratory disease that can progress to death but the factors causing these disparate outcomes remain unclear. Understanding host responses to mild and life-threatening infections provides insight into virus-host networks within and across organ systems that contribute to disease outcomes. We used multi-omics approaches to comprehensively define the host response to moderate and severe MERS-CoV infection. Severe respiratory disease was associated with dysregulation of the immune response. Key lung epithelial cell populations that are essential for lung function get infected and die. Mice lacking key immune cell populations experienced greater virus replication but decreased disease severity implicating the immune system in both protective and pathogenic roles in response to MERS-CoV. These data could be utilized to design new therapeutic strategies targeting specific pathways that contribute to severe disease.

Animals↗
PubMed2026

Paternal exposure to polystyrene nanoplastics induces inter- and transgenerational bronchopulmonary dysplasia-like damage in male offspring by FtMt hypermethylation-mediated ferroptosis.

Bronchopulmonary dysplasia (BPD) is a major cause of chronic lung disease in both preterm infants and adults, but its etiology remains incompletely understood. In this study, F0 generation mice were exposed to polystyrene nanoplastics (PS-NPs), and F1 to F3 generations were obtained by breeding. Multi-omics sequencing including whole genome methylation sequencing, single cell transcriptome sequencing and transcriptome sequencing was performed on the lungs of offspring. The levels of Fe2+, lipid peroxidation products and key gene expression were determined. Male mice exposed to PS-NPs at environmentally relevant doses produced offspring (F1 and F2) that exhibited a typical BPD-like phenotype. Meanwhile, the F0 males showed diminished sperm motility, demonstrating that paternal PS-NPs exposure constituted an etiological factor for BPD in descendants. Mechanistic studies showed that PS-NPs exposure upregulated the expression of DNA methyltransferase Dnmt3a, leading to global hypermethylation of the sperm genome. Importantly, the hypermethylated promoter signature of the mitochondrial ferritin (FtMt) gene partially resisted epigenetic reprogramming and was transmitted to the lungs of offspring, resulting in persistently low FtMt expression in F1 and F2 lungs. This led to increased intracellular Fe2+ levels, subsequently triggered ferroptosis in alveolar epithelial cells, and ultimately impaired alveolarization. Knockdown of FtMt confirmed that FtMt deficiency was sufficient to induce ferroptosis and BPD-like lung injury both in vitro and in vivo. Furthermore, using in vitro fertilization of F0 sperm combined with Dnmt3a siRNA microinjection, we directly demonstrated that Dnmt3a is a key driver for FtMt to escape reprogramming and maintain its hypermethylation. In summary, this study reveals for the first time that paternal PS-NPs exposure causes BPD through a Dnmt3a-FtMt hypermethylation intergenerational and transgenerational axis, providing an epigenetic basis for understanding paternal derived chronic lung disease and potential targets for early intervention.

Animals↗
PubMed2025

ITIH4 alleviates OVA-induced asthma by regulating lung-gut microbiota.

BACKGROUND: Inter-alpha-trypsin inhibitor heavy chain 4 (ITIH4), a Type 2 acute phase protein, is critical for resolving inflammation and promoting tissue repair. While its role in chronic respiratory diseases is recognized, its effects on asthma remain unclear. This study investigated the effects of ITIH4 on the modulation of lung and gut microbiota, the attenuation of allergic inflammation, and the improvement of respiratory outcomes in an asthma mouse model. METHODS: Six-week-old male Balb/c mice were divided into five groups: control, ITIH4, ovalbumin (OVA), and two OVA&#x2009;+&#x2009;ITIH4 treatment groups at different doses. Lung function and oxygen saturation were measured, and bronchoalveolar lavage fluid (BALF) was analyzed for white blood cell counts and cytokines. Lung and gut microbiota were profiled using 16&#xa0;S rRNA gene sequencing, and short-chain fatty acids (SCFAs) were measured using gas chromatography-mass spectrometry (GC-MS). Proteomic profiling of intestinal tissues was conducted to identify ITIH4-associated signaling pathways. RESULTS: ITIH4 administration significantly mitigated OVA-induced asthma symptoms by reducing weight loss, airway resistance, and tissue damping (p&#x2009;<&#x2009;0.05). Histological analysis showed decreased airway wall thickening and lung injury scores (p&#x2009;<&#x2009;0.05). ITIH4 also lowered BALF eosinophils and lymphocytes, IgE, and Th2 cytokines (IL-4, IL-5, and IL-13) (p&#x2009;<&#x2009;0.05). ITIH4 treatment modulated microbiome composition, enriching Gram-positive taxa (Nocardioidaceae and Acholeplasmataceae) and depleting Gram-negative Helicobacteraceae (p&#x2009;<&#x2009;0.05). SCFAs correlated with microbiome alterations, notably reduced 4-methylpentanoic acid levels (p&#x2009;<&#x2009;0.05). Proteomic analysis revealed a dose-dependent activation of granzyme A signaling and suppression of metabolic and solute transport pathways. CONCLUSIONS: ITIH4 ameliorates asthma symptoms by modulating lung and gut microbiota, dampening Th2-driven inflammation, and restoring mucosal immune balance. These findings support ITIH4 as a potential candidate for microbiome-targeted asthma therapy.

Animals↗
PubMed2026

Charge-switching ionizable lipids lower the toxicity of lipid nanoparticles.

Lipid nanoparticles (LNPs) have great potential as nucleic acid delivery vehicles; however, they trigger the production of inflammatory cytokines, which limits their medical applications. Developing non-inflammatory LNPs is challenging because the LNP's ionizable lipid and the process of endosomal disruption are the major sources of LNP toxicity but are also essential for delivering nucleic acids. Here we demonstrate that ionizable lipids containing a carboxylic acid and an amine (termed S-lipid) switch their charged state between the pHs of 7.4 and 4.0, allowing them to generate LNPs (termed switchable nanoparticles) that efficiently encapsulate nucleic acid and trigger endosomal release without activation of the TLR4, complement, galectin-8 and platelet activating factor signalling pathways. Finally, we demonstrate that switchable nanoparticles are better at treating lipopolysaccharide-induced acute lung injury than traditional LNPs because they do not exacerbate pre-existing inflammation. Collectively, these results demonstrate that negatively charged ionizable lipids can mitigate the toxicity of LNPs.

Journal Article↗
PubMed2025

Single-dose cathepsin L CRISPR nanotherapy mitigates PASC-like lung damage in hamsters.

Respiratory post-acute sequelae of COVID-19 (PASC) persists in many SARS-CoV-2 survivors, yet no therapies specifically address its long-term pulmonary damage. We demonstrate that a single-dose CRISPR-CasRx nanotherapy targeting the host enzyme cathepsin L (SCNC) effectively reduces acute SARS-CoV-2 infection in Syrian hamsters, with antiviral efficacy comparable to Paxlovid. Importantly, SCNC outperforms Paxlovid in alleviating alveolar epithelial hyperplasia and lung inflammation at 31 days post-infection, a recognized PASC time point. Single-cell RNA sequencing reveals that SCNC enhances alveolar repair by promoting the differentiation of alveolar type 2 cells into alveolar type 1 cells and by reducing inflammatory infiltration through multiple signaling pathways. Thus, SCNC exerts a dual mechanism: host-directed viral inhibition and promotion of epithelial repair with reduced inflammation. This distinguishes it from therapies focused solely on viral suppression or symptom relief. These findings support SCNC as a promising therapeutic candidate for acute infection and, particularly, for PASC-related lung injury, where options remain limited.

alveolar epithelial regeneration↗
PubMed2024

A proteomics-based survey reveals thrombospondin-4 as a ligand regulated by the mannose receptor in the injured lung.

Receptor-mediated cellular uptake of specific ligands constitutes an important step in the dynamic regulation of individual protein levels in extracellular fluids. With a focus on the inflammatory lung, we here performed a proteomics-based search for novel ligands regulated by the mannose receptor (MR), a macrophage-expressed endocytic receptor. WT and MR-deficient mice were exposed to lipopolysaccharide, after which the protein content in their lung epithelial lining fluid was compared by tandem mass tag-based mass spectrometry. More than 1200 proteins were identified in the epithelial lining fluid using this unbiased approach, but only six showed a statistically different abundance. Among these, an unexpected potential new ligand, thrombospondin-4 (TSP-4), displayed a striking 17-fold increased abundance in the MR-deficient mice. Experiments using exogenous addition of TSP-4 to MR-transfected CHO cells or MR-positive alveolar macrophages confirmed that TSP-4 is a ligand for MR-dependent endocytosis. Similar studies revealed that the molecular interaction with TSP-4 depends on both the lectin activity and the fibronectin type-II domain of MR and that a closely related member of the TSP family, TSP-5, is also efficiently internalized by the receptor. This was unlike the other members of this protein family, including TSPs&#xa0;-1 and&#xa0;-2, which are ligands for a close MR homologue known as urokinase plasminogen activator receptor-associated protein. Our study shows that MR takes part in the regulation of TSP-4, an important inflammatory component in the injured lung, and that two closely related endocytic receptors, expressed on different cell types, undertake the selective endocytosis of distinct members of the TSP family.

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PubMed2025

Hypoxia-inducible factor 2 regulates alveolar regeneration after repetitive injury in three-dimensional cellular and in vivo models.

Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial lung disease in which repetitive epithelial injury and incomplete alveolar repair result in accumulation of profibrotic intermediate/transitional "aberrant" epithelial cell states. The mechanisms leading to the emergence and persistence of aberrant epithelial populations in the distal lung remain incompletely understood. By interrogating single-cell RNA sequencing (scRNA-seq) data from patients with IPF and a mouse model of repeated lung epithelial injury, we identified persistent activation of hypoxia-inducible factor (HIF) signaling in these aberrant epithelial cells. Using mouse genetic lineage-tracing strategies together with scRNA-seq, we found that these disease-emergent aberrant epithelial cells predominantly arose from airway-derived (Scgb1a1-CreER-traced) progenitors and exhibited transcriptional programs of Hif2a activation. In mice treated with repetitive intratracheal bleomycin, deletion of Epas1 (Hif2a) but not Hif1a, from airway-derived progenitors, or administration of the small-molecule HIF2 inhibitor PT-2385, using both prevention and rescue approaches, attenuated experimental lung fibrosis, reduced the appearance of aberrant epithelial cells, and promoted alveolar repair. In mouse alveolar organoids, genetic or pharmacologic inhibition of Hif2 promoted alveolar differentiation of airway-derived epithelial progenitors. In addition, treatment of human distal lung organoids with PT-2385 increased colony-forming efficiency, enhanced protein and transcriptional markers of alveolar type 2 epithelial cell maturation, and prevented the emergence of aberrant epithelial cells. Together, these studies showed that HIF2 activation drives the emergence of aberrant epithelial populations after repetitive injury and that targeted HIF2 inhibition may represent an effective therapeutic strategy to promote functional alveolar repair in IPF and other interstitial lung diseases.

Animals↗