Pathogenesis Of Acute Lung Injury Research Articles

The calcineurin–NFATc pathway modulates the lipid mediators in BAL fluid extracellular vesicles, thereby regulating microvascular endothelial cell barrier function

Extracellular vesicles mediate intercellular communication by transporting biologically active macromolecules. Our prior studies have demonstrated that the nuclear factor of activated T cell cytoplasmic member 3 (NFATc3) is activated in mouse pulmonary macrophages in response to lipopolysaccharide (LPS). Inhibition of NFATc3 activation by a novel cell-permeable calcineurin peptide inhibitor CNI103 mitigated the development of acute lung injury (ALI) in LPS-treated mice. Although pro-inflammatory lipid mediators are known contributors to lung inflammation and injury, it remains unclear whether the calcineurin-NFATc pathway regulates extracellular vesicle (EV) lipid content and if this content contributes to ALI pathogenesis. In this study, EVs from mouse bronchoalveolar lavage fluid (BALF) were analyzed for their lipid mediators by liquid chromatography in conjunction with mass spectrometry (LC-MS/MS). Our data demonstrate that EVs from LPS-treated mice contained significantly higher levels of arachidonic acid (AA) metabolites, which were found in low levels by prior treatment with CNI103. The catalytic activity of lung tissue cytoplasmic phospholipase A2 (cPLA2) increased during ALI, correlating with an increased amount of arachidonic acid (AA) in the EVs. Furthermore, ALI is associated with increased expression of cPLA2, cyclooxygenase 2 (COX2), and lipoxygenases (5-LOX, 12-LOX, and 15-LOX) in lung tissue, and pretreatment with CNI103 inhibited the catalytic activity of cPLA2 and the expression of cPLA2, COX, and LOX transcripts. Furthermore, co-culture of mouse pulmonary microvascular endothelial cell (PMVEC) monolayer and NFAT-luciferase reporter macrophages with BALF EVs from LPS-treated mice increased the pulmonary microvascular endothelial cell (PMVEC) monolayer barrier permeability and luciferase activity in macrophages. However, EVs from CNI103-treated mice had no negative impact on PMVEC monolayer barrier integrity. In summary, BALF EVs from LPS-treated mice carry biologically active NFATc-dependent, AA-derived lipids that play a role in regulating PMVEC monolayer barrier function.

M1 Macrophage-Targeted Curcumin Nanocrystals with l-Arginine-Modified for Acute Lung Injury by Inhalation

Acute Lung Injury/Acute Respiratory Distress Syndrome (ALI/ARDS) with clinical manifestations of respiratory distress and hypoxemia remains a significant cause of respiratory failure, boasting a persistently high incidence and mortality rate. Given the central role of M1 macrophages in the pathogenesis of acute lung injury (ALI), this study utilized the anti-inflammatory agent curcumin as a model drug. l-arginine (L-Arg) was employed as a targeting ligand, and chitosan was initially modified with l-arginine. Subsequently, it was utilized as a surface modifier to prepare inhalable nano-crystals loaded with curcumin (Arg-CS-Cur), aiming for specific targeting of pulmonary M1 macrophages. Compared with unmodified chitosan-curcumin nanocrystals (CS-Cur), Arg-CS-Cur exhibited higher uptake in vitro by M1 macrophages, as evidenced by flow cytometry showing the highest fluorescence intensity in the Arg-CS-Cur group (P < 0.01). In vivo accumulation was greater in inflamed lung tissues, as indicated by small animal imaging demonstrating higher lung fluorescence intensity in the DiR-Arg-CS-Cur group compared to the DiR-CS-Cur group in the rat ALI model (P < 0.05), peaking at 12 h. Moreover, Arg-CS-Cur demonstrated enhanced therapeutic effects in both LPS-induced RAW264.7 cells and ALI rat models. Specifically, treatment with Arg-CS-Cur significantly suppressed NO release and levels of TNF-α and IL-6 in RAW264.7 cells (p < 0.01), while in ALI rat models, expression levels of TNF-α and IL-6 in lung tissues were significantly lower than those in the model group (P < 0.01). Furthermore, lung tissue damage was significantly reduced, with histological scores significantly lower than those in the CS-Cur group (P < 0.01). In conclusion, these findings underscore the targeting potential of l-arginine-modified nanocrystals, which effectively enhance curcumin concentration in inflammatory environments by selectively targeting M1 macrophages. This study thus introduces novel perspectives and theoretical support for the development of targeted therapeutic interventions for acute inflammatory lung diseases, including ALI/ARDS.

The involvement of oxidative stress and the TLR4/NF-κB/NLRP3 pathway in acute lung injury induced by high-altitude hypoxia

ObjectiveThis study investigated the effect of oxidative stress and the TLR4/NF-κB/NLRP3 pathway on the pathogenesis of acute lung injury (ALI) induced by high-altitude hypoxia. MethodsRats were placed in an animal hyperbaric oxygen chamber to establish a rat model of ALI induced by high-altitude hypoxia after treatment with N-acetylcysteine (NAC; a reactive oxygen species [ROS] inhibitor) or/and MCC950 (an NLPR3 inflammasome inhibitor). After modeling, the wet-to-dry weight ratio (W/D) of rat lung tissues was calculated. In lung tissues, ROS levels were detected with immunofluorescence, the enzyme activity was tested with the kit, and the expression of TLR4/NF-κB/NLRP3 pathway-related genes and proteins was measured with western blotting and qRT-PCR. The levels of inflammatory factors in the serum were quantified with ELISA. ResultsAfter modeling, rats showed significantly increased W/D, ROS levels, and Malondialdehyde (MDA) concentrations and markedly diminished Superoxide dismutase (SOD) and Glutathione (GSH) concentrations in lung tissues (all P < 0.01), accompanied by substantially enhanced serum levels of TNF-α, IL-6, and IL-1β, significantly reduced serum levels of IL-10, and remarkably augmented TLR4, NLRP3, p-NF-κB p65, NF-κB p65 mRNA, and Caspase-1 expression in lung tissues (all P < 0.01). Furthermore, treatment with NAC or MCC950 alone or in combination prominently lowered the W/D of lung tissues (P < 0.01), serum levels of TNF-α (P < 0.05), IL-6 (P < 0.05), and IL-1β (P < 0.01), and NF-κB p65 expression and phosphorylation (P < 0.05, P < 0.01) while significantly increasing SOD and GSH concentrations (P < 0.05, P < 0.01) and serum levels of IL-10 (P < 0.01) in modeled rats. Meanwhile, treatment of NAC alone or combined with MCC950 significantly reduced MDA concentration and ROS levels (P < 0.05, P < 0.01) in modeled rats, and treatment of MCC950 alone or combined with NAC considerably declined TLR4, NLRP3, and Caspase-1 expression in modeled rats (P < 0.05, P < 0.01). ConclusionInhibition of oxidative stress and the TLR4/NF-κB/NLRP3 pathway can ameliorate ALI in rats exposed to high-altitude hypoxia.

SIRT1-Rab7 axis attenuates NLRP3 and STING activation through late endosomal-dependent mitophagy during sepsis-induced acute lung injury.

Acute lung injury (ALI) is a leading cause of mortality in patients with sepsis due to proinflammatory endothelial changes and endothelial permeability defects. Mitochondrial dysfunction is recognized as a critical mediator in the pathogenesis of sepsis-induced ALI. Although mitophagy regulation of mitochondrial quality is well recognized, little is known about its role in lung ECs during sepsis-induced ALI. Sirtuin 1 (SIRT1) is a histone protein deacetylase involved in inflammation, mitophagy, and cellular senescence. Here, the authors show a type of late endosome-dependent mitophagy that inhibits NLRP3 and STING activation through SIRT1 signaling during sepsis-induced ALI. C57BL/6J male mice with or without administration of the SIRT1 inhibitor EX527 in the CLP model and lung ECs in vitro were developed to identify mitophagy mechanisms that underlie the cross-talk between SIRT1 signaling and sepsis-induced ALI. SIRT1 deficient mice exhibited exacerbated sepsis-induced ALI. Knockdown of SIRT1 interfered with mitophagy through late endosome Rab7, leading to the accumulation of damaged mitochondria and inducing excessive mitochondrial reactive oxygen species (mtROS) generation and cytosolic release of mitochondrial DNA (mtDNA), which triggered NLRP3 inflammasome and the cytosolic nucleotide sensing pathways (STING) over-activation. Pharmacological inhibition of STING and NLRP3 i n vivo or genetic knockdown in vitro reversed SIRT1 deficiency mediated endothelial permeability defects and endothelial inflammation in sepsis-induced ALI. Moreover, activation of SIRT1 with SRT1720 in vivo or overexpression of SIRT1 in vitro protected against sepsis-induced ALI. These findings suggest that SIRT1 signaling is essential for restricting STING and NLRP3 hyperactivation by promoting endosomal-mediated mitophagy in lung ECs, providing potential therapeutic targets for treating sepsis-induced ALI.

Targeting the TRAF3-ULK1-NLRP3 regulatory axis to control alveolar macrophage pyroptosis in acute lung injury.

Acute lung injury (ALI) is a serious condition characterized by damage to the lungs. Recent research has revealed that activation of the NLRP3 inflammasome in alveolar macrophages, a type of immune cell in the lungs, plays a key role in the development of ALI. This process, known as pyroptosis, contributes significantly to ALI pathogenesis. Researchers have conducted comprehensive bioinformatics analyses and identified 15 key genes associated with alveolar macrophage pyroptosis in ALI. Among these, NLRP3 has emerged as a crucial regulator. This study further reveal that the ULK1 protein diminishes the expression of NLRP3, thereby reducing the immune response of alveolar macrophages and mitigating ALI. Conversely, TRAF3, another protein, is found to inhibit ULK1 through a process called ubiquitination, leading to increased activation of the NLRP3 inflammasome and exacerbation of ALI. This TRAF3-mediated suppression of ULK1 and subsequent activation of NLRP3 are confirmed through various in vitro and in vivo experiments. The presence of abundant M0 and M1 alveolar macrophages in the ALI tissue samples further support these findings. This research highlights the TRAF3-ULK1-NLRP3 regulatory axis as a pivotal pathway in ALI development and suggests that targeting this axis could be an effective therapeutic strategy for ALI treatment.

Acute Lung Injury after Cardiopulmonary Resuscitation: A Narrative Review.

Although cardiopulmonary resuscitation (CPR) includes lifesaving maneuvers, it might be associated with a wide spectrum of iatrogenic injuries. Among these, acute lung injury (ALI) is frequent and yields significant challenges to post-cardiac arrest recovery. Understanding the relationship between CPR and ALI is determinant for refining resuscitation techniques and improving patient outcomes. This review aims to analyze the existing literature on ALI following CPR, emphasizing prevalence, clinical implications, and contributing factors. The review seeks to elucidate the pathogenesis of ALI in the context of CPR, assess the efficacy of CPR techniques and ventilation strategies, and explore their impact on post-cardiac arrest outcomes. CPR-related injuries, ranging from skeletal fractures to severe internal organ damage, underscore the complexity of managing post-cardiac arrest patients. Chest compression, particularly when prolonged and vigorous, i.e., mechanical compression, appears to be a crucial factor contributing to ALI, with the concept of cardiopulmonary resuscitation-associated lung edema (CRALE) gaining prominence. Ventilation strategies during CPR and post-cardiac arrest syndrome also play pivotal roles in ALI development. The recognition of CPR-related lung injuries, especially CRALE and ALI, highlights the need for research on optimizing CPR techniques and tailoring ventilation strategies during and after resuscitation.

Crtc1 deficiency protects against sepsis-associated acute lung injury through activating akt signaling pathway

BackgroundInterplay between systemic inflammation and programmed cell death contributes to the pathogenesis of acute lung injury (ALI). cAMP-regulated transcriptional coactivator 1 (CRTC1) has been involved in the normal function of the pulmonary system, but its role in ALI remains unclear.Methods and resultsWe generated a Crtc1 knockout (KO; Crtc1−/−) mouse line. Sepsis-induced ALI was established by cecal ligation and puncture (CLP) for 24 h. The data showed that Ctrc1 KO substantially ameliorated CLP-induced ALI phenotypes, including improved lung structure destruction, reduced pulmonary vascular permeability, diminished levels of proinflammatory cytokines and chemokines, compared with the wildtype mice. Consistently, in lipopolysaccharide (LPS)-treated RAW264.7 cells, Crtc1 knockdown significantly inhibited the expression of inflammatory effectors, including TNF-α, IL-1β, IL-6 and CXCL1, whereas their expressions were significantly enhanced by Crtc1 overexpression. Moreover, both Crtc1 KO in mice and its knockdown in RAW264.7 cells dramatically reduced TUNEL-positive cells and the expression of pro-apoptotic proteins. In contrast, Crtc1 overexpression led to an increase in the pro-apoptotic proteins and LPS-induced TUNEL-positive cells. Mechanically, we found that the phosphorylation of Akt was significantly enhanced by Crtc1 knockout or knockdown, but suppressed by Crtc1 overexpression. Administration of Triciribine, an Akt inhibitor, substantially blocked the protection of Crtc1 knockdown on LPS-induced inflammation and cell death in RAW264.7 cells.ConclusionsOur study demonstrates that CRTC1 contribute to the pathological processes of inflammation and apoptosis in sepsis-induced ALI, and provides mechanistic insights into the molecular function of CRTC1 in the lung. Targeting CRTC1 would be a promising strategy to treat sepsis-induced ALI in clinic.

STING mediates LPS-induced acute lung injury by regulating ferroptosis

The pathogenesis of acute lung injury is not fully understood. Stimulator of interferon genes (STING) and ferroptosis have been implicated in various pathological and physiological processes, including acute lung injury (ALI). However, the relationship between STING and ferroptosis in lipopolysaccharide (LPS)-induced ALI is unclear. We found that LPS stimulation activated STING and ferroptosis. Furthermore, STING knockout and ferroptosis inhibitor alleviated lung inflammation and epithelial cell damage. Also, STING knockout reduced inflammation injury and ferroptosis. Notably, the ferroptosis inducer reversed the alleviation of inflammation caused by STING knockout. These results show that STING participates in the inflammation injury of ALI by regulating ferroptosis. Results also showed that p-STAT3 levels increased after STING knockout, suggesting that STING negatively regulates STAT3 activation. Besides, STAT3 inhibitor aggravated ferroptosis after STING knockout, indicating that STING regulates ferroptosis through STAT3 signaling. In conclusion, STING mediates LPS-induced ALI by regulating ferroptosis, indicating that STING and ferroptosis may be new targets for ALI treatment.

Garlic oil supplementation blocks inflammatory pyroptosis-related acute lung injury by suppressing the NF-κB/NLRP3 signaling pathway via H2S generation.

Acute lung injury (ALI) is a major cause of acute respiratory failure with a high morbidity and mortality rate, and effective therapeutic strategies for ALI remain limited. Inflammatory response is considered crucial for the pathogenesis of ALI. Garlic, a globally used cooking spice, reportedly exhibits excellent anti-inflammatory bioactivity. However, protective effects of garlic against ALI have never been reported. This study aimed to investigate the protective effects of garlic oil (GO) supplementation on lipopolysaccharide (LPS)-induced ALI models. Hematoxylin and eosin staining, pathology scores, lung myeloperoxidase (MPO) activity measurement, lung wet/dry (W/D) ratio detection, and bronchoalveolar lavage fluid (BALF) analysis were performed to investigate ALI histopathology. Real-time polymerase chain reaction, western blotting, and enzyme-linked immunosorbent assay were conducted to evaluate the expression levels of inflammatory factors, nuclear factor-κB (NF-κB), NLRP3, pyroptosis-related proteins, and H2S-producing enzymes. GO attenuated LPS-induced pulmonary pathological changes, lung W/D ratio, MPO activity, and inflammatory cytokines in the lungs and BALF. Additionally, GO suppressed LPS-induced NF-κB activation, NLRP3 inflammasome expression, and inflammatory-related pyroptosis. Mechanistically, GO promoted increased H2S production in lung tissues by enhancing the conversion of GO-rich polysulfide compounds or by increasing the expression of H2S-producing enzymes in vivo. Inhibition of endogenous or exogenous H2S production reversed the protective effects of GO on ALI and eliminated the inhibitory effects of GO on NF-κB, NLRP3, and pyroptotic signaling pathways. Overall, these findings indicate that GO has a critical anti-inflammatory effect and protects against LPS-induced ALI by suppressing the NF-κB/NLRP3 signaling pathway via H2S generation.

S100A9 exacerbates sepsis-induced acute lung injury via the IL17-NFκB-caspase-3 signaling pathway

BackgroundSepsis-induced acute lung injury (ALI) is associated with considerable morbidity and mortality in critically ill patients. S100A9, a key endothelial injury factor, is markedly upregulated in sepsis-induced ALI; however, its specific mechanism of action has not been fully elucidated. MethodsThe Gene Expression Omnibus database transcriptome data for sepsis-induced ALI were used to screen for key differentially expressed genes (DEGs). Using bioinformatics analysis methods such as Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, and protein-protein interaction network analyses, the pathogenesis of sepsis-induced ALI was revealed. Intratracheal infusion of lipopolysaccharide (LPS, 10 mg/kg) induced ALI in wild-type (WT) and S100A9 knockout mice. Multiomics analyses (transcriptomics and proteomics) were performed to investigate the potential mechanisms by which S100A9 exacerbates acute lung damage. Hematoxylin-eosin, Giemsa, and TUNEL staining were used to evaluate lung injury and cell apoptosis. LPS (10 μg/mL)-induced murine lung epithelial MLE-12 cells were utilized to mimic ALI and were modulated by S100A9 lentiviral transfection. The impact of S100A9 on cell apoptosis and inflammatory responses were identified using flow cytometry and PCR. The expression of interleukin (IL)-17–nuclear factor kappa B (NFκB)–caspase-3 signaling components was identified using western blotting. ResultsSix common DEGs (S100A9, S100A8, IFITM6, SAA3, CD177, and MMP9) were identified in the six datasets related to ALI in sepsis. Compared to WT sepsis mice, S100A9 knockout significantly alleviated LPS-induced ALI in mice, with reduced lung structural damage and inflammatory exudation, decreased exfoliated cell and protein content in the lung lavage fluid, and reduced apoptosis and necrosis of pulmonary epithelial cells. Transcriptomic analysis revealed that knocking out S100A9 significantly affected 123 DEGs, which were enriched in immune responses, defense responses against bacteria or lipopolysaccharides, cytokine-cytokine receptor interactions, and the IL-17 signaling pathway. Proteomic analysis revealed that S100A9 knockout alleviated muscle contraction dysfunction and structural remodeling in sepsis-induced ALI. Multiomics analysis revealed that S100A9 may be closely related to interferon-induced proteins with tetratricopeptide repeats and oligoadenylate synthase-like proteins. LPS decreased MLE12 cell activity, accompanied by high expression of S100A9. The expression of IL-17RA, pNFκB, and cleaved-caspase-3 were increased by S100A9 overexpression and reduced by S100A9 knockdown in LPS-stimulated MLE12 cells. S100A9 knockdown decreases transcription of apoptosis-related markers Bax, Bcl and caspase-3, alleviating LPS-induced apoptosis. ConclusionsS100A9 as a key biomarker of sepsis-induced acute lung injury, and exacerbates lung damage and epithelial cell apoptosis induced by LPS via the IL-17–NFκB–caspase-3 signaling pathway.

Ischemia and reperfusion-injured liver-derived exosomes elicit acute lung injury through miR-122-5p regulated alveolar macrophage polarization

Acute lung injury (ALI) is a common postoperative complication, particularly in pediatric patients after liver transplantation. Hepatic ischemia–reperfusion (HIR) increases the release of exosomes (IR-Exos) in peripheral circulation. However, the role of IR-Exos in the pathogenesis of ALI induced by HIR remains unclear. Here, we explored the role of exosomes derived from the HIR-injured liver in ALI development. Intravenous injection of IR-Exos caused lung inflammation in naive rats, whereas pretreatment with an inhibitor of exosomal secretion (GW4869) attenuated HIR-related lung injury. In vivo and in vitro results show that IR-Exos promoted proinflammatory responses and M1 macrophage polarization. Furthermore, miRNA profiling of serum identified miR-122-5p as the exosomal miRNA with the highest increase in young rats with HIR compared with controls. Additionally, IR-Exos transferred miR-122-5p to macrophages and promoted proinflammatory responses and M1 phenotype polarization by targeting suppressor of cytokine signaling protein 1(SOCS-1)/nuclear factor (NF)-κB. Importantly, the pathological role of exosomal miR-122-5p in initiating lung inflammation was reversed by inhibition of miR-122-5p. Clinically, high levels of miR-122-5p were found in serum and correlated to the severity of lung injury in pediatric living-donor liver transplant recipients with ALI. Taken together, our findings reveal that IR-Exos transfer liver-specific miR-122-5p to alveolar macrophages and elicit ALI by inducing M1 macrophage polarization via the SOCS-1/NF-κB signaling pathway.

Roles of Macrophages and Endothelial Cells and Their Crosstalk in Acute Lung Injury.

Acute lung injury (ALI) and its severe form, acute respiratory distress syndrome (ARDS), present life-threatening conditions characterized by inflammation and endothelial injury, leading to increased vascular permeability and lung edema. Key players in the pathogenesis and resolution of ALI are macrophages (Mφs) and endothelial cells (ECs). The crosstalk between these two cell types has emerged as a significant focus for potential therapeutic interventions in ALI. This review provides a brief overview of the roles of Mφs and ECs and their interplay in ALI/ARDS. Moreover, it highlights the significance of investigating perivascular macrophages (PVMs) and immunomodulatory endothelial cells (IMECs) as crucial participants in the Mφ-EC crosstalk. This sheds light on the pathogenesis of ALI and paves the way for innovative treatment approaches.

GSTP alleviates acute lung injury by S-glutathionylation of KEAP1 and subsequent activation of NRF2 pathway

Oxidative stress plays an important role in the pathogenesis of acute lung injury (ALI). As a typical post-translational modification triggered by oxidative stress, protein S-glutathionylation (PSSG) is regulated by redox signaling pathways and plays diverse roles in oxidative stress conditions. In this study, we found that GSTP downregulation exacerbated LPS-induced injury in human lung epithelial cells and in mice ALI models, confirming the protective effect of GSTP against ALI both in vitro and in vivo. Additionally, a positive correlation was observed between total PSSG level and GSTP expression level in cells and mice lung tissues. Further results demonstrated that GSTP inhibited KEAP1-NRF2 interaction by promoting PSSG process of KEAP1. By the integration of protein mass spectrometry, molecular docking, and site-mutation validation assays, we identified C434 in KEAP1 as the key PSSG site catalyzed by GSTP, which promoted the dissociation of KEAP1-NRF2 complex and activated the subsequent anti-oxidant genes. In vivo experiments with AAV-GSTP mice confirmed that GSTP inhibited LPS-induced lung inflammation by promoting PSSG of KEAP1 and activating the NRF2 downstream antioxidant pathways. Collectively, this study revealed the novel regulatory mechanism of GSTP in the anti-inflammatory function of lungs by modulating PSSG of KEAP1 and the subsequent KEAP1/NRF2 pathway. Targeting at manipulation of GSTP level or activity might be a promising therapeutic strategy for oxidative stress-induced ALI progression.

Fortunellin ameliorates LPS-induced acute lung injury, inflammation, and collagen deposition by restraining the TLR4/NF-κB/NLRP3 pathway.

Acute lung injury (ALI) is the prevalent respiratory disease of acute inflammation with high morbidity and mortality. Fortunellin has anti-inflammation property, but its role in ALI remains elusive. Thus, this study clarified the function of fortunellin on ALI pathogenesis. The ALI mouse model was established by lipopolysaccharide (LPS) induction, and lung tissue damage was evaluated utilizing hematoxylin-eosin (HE) staining. The edema of lung tissue was measured by the lung wet/dry (W/D) ratio. The lung capillary permeability was reflected by the protein content in bronchoalveolar lavage fluid (BALF). Inflammatory cell infiltration was measured by the evaluation of the content of myeloperoxidase (MPO), neutrophils, and leukocytes in BALF. Cell apoptosis was measured by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. The secretions of inflammatory cytokines were quantified using enzyme-linked immunosorbent assay (ELISA) assays. Lung tissue collagen deposition was evaluated by Masson staining. Fortunellin attenuated LPS-induced lung tissue damage and reduced the W/D ratio, the content of MPO in lung tissue, the total protein contents in BALF, and the neutrophils and leukocytes number. Besides, fortunellin alleviated LPS-stimulated lung tissue apoptosis, inflammatory response, and collagen deposition. Furthermore, Fortunellin repressed the activity of the Toll-like receptor 4 (TLR4)/nuclear factor kappa-B (NF-κB)/NLR Family Pyrin Domain Containing 3 (NLRP3) pathway in the LPS-stimulated ALI model and LPS-induced RAW264.7 cells. Moreover, fortunellin attenuated LPS-stimulated tissue injury, apoptosis, inflammation, and collagen deposition of the lung via restraining the TLR4/NF-κB/NLRP3 pathway. Fortunellin attenuated LPS-stimulated ALI through repressing the TLR4/NF-κB/NLRP3 pathway. Fortunellin may be a valuable drug for ALI therapy.

The role and mechanism of PDZ binding kinase in hypobaric and hypoxic acute lung injury.

Acute lung injury (ALI) caused by hypobaric hypoxia (HH) is frequently observed in high-altitude areas, and it is one of the leading causes of death in high-altitude-related diseases due to its rapid onset and progression. However, the pathogenesis of HH-related ALI (HHALI) remains unclear, and effective treatment approaches are currently lacking. A new mouse model of HHALI developed by our laboratory was used as the study subject (Chinese patent No. ZL 2021 1 1517241 X). Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the messenger RNA (mRNA) expression levels of PDZ-binding kinase (PBK), sirtuin 1 (SIRT1), and PTEN-induced kinase 1 (PINK1) in mouse lung tissue. Hematoxylin and eosin staining was used to observe the main types of damage and damaged cells in lung tissue, and the lung injury score was used for quantification. The wet-dry (W/D) ratio was used to measure lung water content. Enzyme-linked immunosorbent assay was used to detect changes in inflammatory factors and oxidative stress markers in the lungs. Western blotting verified the expression of various mitochondrial autophagy-related proteins. The 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimi-dazoylcarbocyanine iodide (JC-1) method was used determined the health status of mitochondria based on changes in mitochondrial membrane potential. Transmission electron microscopy was used to directly observe the morphology of mitochondria. Multicolor immunofluorescence was used to observe the levels of mitochondrial autophagy markers. Other signaling pathways and molecular mechanisms that may play a role in epithelial cells were analyzed via through RNA sequencing. Low pressure and hypoxia caused pathological changes in mouse lung tissue, mainly ALI, leading to increased levels of inflammatory factors and intensified oxidative stress response in the lungs. Overexpression of PBK was found to alleviate HHALI, and activation of the p53 protein was shown to abrogate this therapeutic effect, while activation of SIRT1 protein reactivated this therapeutic effect. The therapeutic effect of PBK on HHALI is achieved via the activation of mitochondrial autophagy. Finally, RNA sequencing demonstrated that besides mitochondrial autophagy, PBK also exerts other functions in HHALI. Overexpression of PBK inhibits the expression of p53 and activates SIRT1-PINK1 axis mediated mitochondrial autophagy to alleviate HHALI.

Integrated network pharmacology and experimental validation-based approach to reveal the underlying mechanisms and key material basis of Jinhua Qinggan granules against acute lung injury

Ethnopharmacological relevanceJinhua Qinggan granules (JHQG), the traditional Chinese formula come into the market in 2016, has been proved clinically effective against coronavirus disease. Acute lung injury (ALI) is a major complication of respiratory infection such as coronavirus and influenza virus, with a high clinical fatality rate. Macrophage activation-induced inflammatory response plays a crucial role in the pathogenesis of ALI. However, the participation of inflammatory response in the efficacy of JHQG and its material basis against ALI is still unknown. Aim of the studyThe research aims to investigate the inflammatory response-involved efficacy of JHQG on ALI, explore the “ingredient-target-pathway” mechanisms, and searching for key material basis of JHQG by integrated network pharmacology and experimental validation-based approach. Materials and methodsLipopolysaccharide (LPS)-induced ALI mice was established to assess the protective impact of JHQG. Network pharmacology was utilized to identify potential targets of JHQG and investigate its action mechanisms related to inflammatory response in treating ALI. The therapeutic effect and mechanism of the primary active ingredient in JHQG was verified through high performance liquid chromatography (HPLC) and a combination of wet experiments. ResultsJHQG remarkably alleviated lung damage in mice model via suppressing macrophage activation, and inhibiting pro-inflammatory mediator level, p-ERK and p-STAT3 expression, TLR4/NF-κB activation. Network pharmacology combined with HPLC found luteolin is the main effective component of JHQG, and it could interact with TLR4/MD2 complex, further exerting the anti-inflammatory property and the protective role against ALI. ConclusionsIn summary, our finding clarified the underlying mechanisms and material basis of JHQG therapy for ALI by integrated network pharmacology and experimental validation-based strategy.

Suppressing Endoplasmic Reticulum Stress Alleviates LPS-Induced Acute Lung Injury via Inhibiting Inflammation and Ferroptosis.

Acute lung injury (ALI) is a life-threatening clinical disorder with high mortality rate. Ferroptosis is a new type of programmed cell death with lipid peroxidation and iron ion overloading as the main characteristics. Endoplasmic reticulum (ER) stress and ferroptosis play pivotal roles in the pathogenesis of ALI. The study aimed to investigate the underlying relationship between ER stress and ferroptosis in ALI. The ER stress inhibitor 4-phenylbutyric acid (4-PBA) alleviated LPS-induced inflammation, and decreased IL-1β, IL-6, and TNF-α levels in BALF and lungs. The increased MDA and decreased GSH induced by LPS were partially reversed by 4-PBA, which also inhibited the expressions of ferroptosis-related protein ACSL4, COX-2, and FTH1. TEM further confirmed the ferroptosis within airway epithelia cells was ameliorated by 4-PBA. Moreover, 4-PBA reduced the production of ROS and lipid ROS in LPS-exposed BEAS-2B cells in a concentration-dependent way. Meanwhile, 4-PBA mitigated LPS-induced cell apoptosis in vivo and in vitro. Mechanistically, the MAPK signaling pathway activated by LPS was downregulated by 4-PBA. Collectively, these findings suggested that 4-PBA protected against ALI by inhibiting inflammation and ferroptosis through downregulating ER stress, thus providing a potential intervention for ALI and revealing the possible interaction between ER stress and ferroptosis in ALI.

Role of biomarkers and molecular signaling pathways in acute lung injury.

Acute lung injury (ALI) is caused by bacterial, fungal, and viral infections. When pathogens invade the lungs, the immune system responds by producing cytokines, chemokines, and interferons to promote the infiltration of phagocytic cells, which are essential for pathogen clearance. Their excess production causes an overactive immune response and a pathological hyper-inflammatory state, which leads to ALI. Until now, there is no particular pharmaceutical treatment available for ALI despite known inflammatory mediators like neutrophil extracellular traps (NETs) and reactive oxygen species (ROS). Therefore, the primary objective of this review is to provide the clear overview on the mechanisms controlling NETs, ROS formation, and other relevant processes during the pathogenesis of ALI. In addition, we have discussed the significance of epithelial and endothelial damage indicators and several molecular signaling pathways associated with ALI. The literature review was done from Web of Science, Scopus, PubMed, and Google Scholar for ALI, NETs, ROS, inflammation, biomarkers, Toll- and nucleotide-binding oligomerization domain (NOD)-like receptors, alveolar damage, pro-inflammatory cytokines, and epithelial/endothelial damage alone or in combination. This review summarized the main clinical signs of ALI, including the regulation and distinct function of epithelial and endothelial biomarkers, NETs, ROS, and pattern recognition receptors (PRRs). However, no particular drugs including vaccine for ALI has been established. Furthermore, there is a lack of validated diagnostic tools and a poor predictive rationality of current therapeutic biomarkers. Hence, extensive and precise research is required to speed up the process of drug testing and development by the application of artificial intelligence technologies, structure-based drug design, in-silico approaches, and drug repurposing.

CPT1A as a potential therapeutic target for lipopolysaccharide-induced acute lung injury in mice

Acute lung injury (ALI) remains a high mortality rate with dramatic lung inflammation and alveolar epithelial cell death. Although fatty acid β-oxidation (FAO) impairment has been implicated in the pathogenesis of ALI, whether Carnitine palmitoyltransferase 1A (CPT1A), the rate-limiting enzyme for FAO, plays roles in lipopolysaccharide (LPS)-induced ALI remains unclear. Accordingly, we focused on exploring the effect of CPT1A in the context of ALI and the underlying mechanisms. We found that overexpression of CPT1A (AAV-CPT1A) effectively alleviated lung injury by reduction of lung wet-to-dry ratio, inflammatory cell infiltration, and protein levels in the BALF of ALI mice. Meanwhile, AAV-CPT1A significantly lessened histopathological changes and several cytokines’ secretions. In contrast, blocking CPT1A with etomoxir augmented inflammatory responses and lung injury in ALI mice. Furthermore, we found that overexpression of CPT1A with lentivirus reduced the apoptosis rates of alveolar epithelial cells and the expression of apoptosis-related proteins induced by LPS in MLE12 cells, while etomoxir increased the apoptosis of MLE12 cells. Overexpression of CPT1A prevented the drop in bioenergetics, palmitate oxidation, and ATP levels. In conclusion, the results rendered CPT1A worthy of further development into a pharmaceutical drug for the treatment of ALI.

Salecan ameliorates LPS-induced acute lung injury through regulating Keap1-Nrf2/HO-1 pathway in mice

Acute lung injury (ALI) is a severe clinical condition with high mortality, characterized by rapid onset and limited treatment options. The pathogenesis of ALI involves inflammation and oxidative stress. The polysaccharide salecan, a water-soluble β-(1,3)-D-glucan, has been found to possess numerous pharmaceutical effects, including anti-inflammatory properties, inhibition of oxidative stress, and anti-fatigue effects. This study aims to investigate the protective effect and underlying mechanism of salecan against LPS-induced ALI in mice. Using an in vivo LPS-induced ALI mouse model and an in vitro RAW264.7 cell system, we investigated the role of salecan in ALI with various experimental approaches, including histological staining, quantitative real-time PCR, flow cytometry, western blot analysis, and other relevant assays. Pre-treatment with salecan effectively attenuated LPS-induced ALI in vivo, reducing the severity of pulmonary edema, inflammation, and oxidative stress. NMR-based metabolomic profiling analysis revealed that salecan attenuated LPS-induced metabolic imbalances associated with ALI. Furthermore, salecan downregulated Keap1 and upregulated Nrf2 and HO-1 protein levels, indicating its modulation of the Keap1-Nrf2/HO-1 signaling pathway as a potential mechanism underlying its protective effects against ALI. In vitro studies on RAW264.7 cells revealed that salecan exhibited binding affinity towards macrophages, thereby alleviating LPS-induced apoptosis and inflammation, which underpin its therapeutic potential against ALI. Our study suggests that salecan can alleviate LPS-induced ALI by modulating oxidative stress, inflammatory response, and apoptosis through the activation of the Keap1-Nrf2/HO-1 pathway. These findings provide novel insights into the potential therapeutic use of salecan for the treatment of ALI.