Expression In Lung Tissue Research Articles

Obesity-induced downregulation of miR-192 exacerbates lipopolysaccharide-induced acute lung injury by promoting macrophage activation

BackgroundMacrophage activation may play a crucial role in the increased susceptibility of obese individuals to acute lung injury (ALI). Dysregulation of miRNA, which is involved in various inflammatory diseases, is often observed in obesity. This study aimed to investigate the role of miR-192 in lipopolysaccharide (LPS)-induced ALI in obese mice and its mechanism of dysregulation in obesity.MethodsHuman lung tissues were obtained from obese patients (BMI ≥ 30.0 kg/m2) and control patients (BMI 18.5–24.9 kg/m2). An obese mouse model was established by feeding a high-fat diet (HFD), followed by intratracheal instillation of LPS to induce ALI. Pulmonary macrophages of obese mice were depleted through intratracheal instillation of clodronate liposomes. The expression of miR-192 was examined in lung tissues, primary alveolar macrophages (AMs), and the mouse alveolar macrophage cell line (MH-S) using RT-qPCR. m6A quantification and RIP assays helped determine the cause of miR-192 dysregulation. miR-192 agomir and antagomir were used to investigate its function in mice and MH-S cells. Bioinformatics and dual-luciferase reporter gene assays were used to explore the downstream targets of miR-192.ResultsIn obese mice, depletion of macrophages significantly alleviated lung tissue inflammation and injury, regardless of LPS challenge. miR-192 expression in lung tissues and alveolar macrophages was diminished during obesity and further decreased with LPS stimulation. Obesity-induced overexpression of FTO decreased the m6A modification of pri-miR-192, inhibiting the generation of miR-192. In vitro, inhibition of miR-192 enhanced LPS-induced polarization of M1 macrophages and activation of the AKT/ NF-κB inflammatory pathway, while overexpression of miR-192 suppressed these reactions. BIG1 was confirmed as a target gene of miR-192, and its overexpression offset the protective effects of miR-192. In vivo, when miR-192 was overexpressed in obese mice, the activation of pulmonary macrophages and the extent of lung injury were significantly improved upon LPS challenge.ConclusionsOur study indicates that obesity-induced downregulation of miR-192 expression exacerbates LPS-induced ALI by promoting macrophage activation. Targeting macrophages and miR-192 may provide new therapeutic avenues for obesity-associated ALI.Graphical

Effect of TAK242 on MCP-1 and TGF-β in COPD Rats

Objective: To investigate the mechanism of MCP-1 and TGF-β regulation by TAK242 in COPD rats. Methods: Thirty-six SD rats were randomly divided into normal, COPD control, and TAK242 groups. The normal group was freely fed, and the other groups used the method of fumigation plus lipopolysaccharide tracheal drip to establish an experimental animal model of COPD. After successful modeling, each experimental group received 0.9% NaCl solution and corresponding drugs by intraperitoneal injection for 7 d. After drug administration, lung function was examined; pathological changes in lung tissue were observed by light microscopy with hematoxylin-eosin staining; mRNA expression of MCP-1 and TGF-β was detected by q-PCR; and protein expression of MCP-1 and TGF-β in lung tissue was detected by Western blot and IHC, TGF-β protein expression in rat lung tissue. Results: Compared with the normal group, rats in the COPD control group showed signs and symptoms of COPD, decreased lung function, and increased expression of MCP-1 and TGF-β. The TAK242 group showed decreased expression of MCP-1 and TGF-β compared to the COPD control group. Conclusion: MCP-1, and TGF-β played a crucial role in the early stage of COPD fibrosis. TAK242 could ameliorate airway inflammation and inhibit the progression of COPD lung fibrosis in pre-existing rats in COPD model rats.

Identification of HK3 as a promising immunomodulatory and prognostic target in sepsis-induced acute lung injury

BackgroundSepsis is a life-threatening global disease with a significant impact on human health. Acute lung injury (ALI) has been identified as one of the primary causes of mortality in septic patients. This study aimed to identify candidate genes involved in sepsis-induced ALI through a comprehensive approach combining bioinformatics analysis and experimental validation. MethodsThe datasets GSE65682 and GSE32707 obtained from the Gene Expression Omnibus database were merged to screen for sepsis-induced ALI related differentially expressed genes (DEGs). Functional enrichment and immune infiltration analyses were conducted on DGEs, with the construction of protein-protein interaction (PPI) networks to identify hub genes. In vitro and in vivo models of sepsis-induced ALI were used to study the expression and function of hexokinase 3 (HK3) using various techniques including Western blot, real-time PCR, immunohistochemistry, immunofluorescence, Cell Counting Kit-8, Enzyme-linked immunosorbent assay, and flow cytometry. ResultsThe results of bioinformatics analysis have identified HK3, MMP9, and S100A8 as hub genes with diagnostic and prognostic significance for sepsis-induced ALI. The HK3 has profound effects on sepsis-induced ALI and exhibits a correlation with immune regulation. Experimental results showed increased HK3 expression in lung tissue of septic mice, particularly in bronchial and alveolar epithelial cells. In vitro studies demonstrated upregulation of HK3 in lipopolysaccharide (LPS)-stimulated lung epithelial cells, with cytoplasmic localization around the nucleus. Interestingly, following the knockdown of HK3 expression, lung epithelial cells exhibited a significant decrease in proliferation activity and glycolytic flux, accompanied by an increase in cellular inflammatory response, oxidative stress, and cell apoptosis. ConclusionsIt was observed for the first time that HK3 plays a crucial role in the progression of sepsis-induced ALI and may be a valuable target for immunomodulation and therapy.Bioinformatics analysis identified HK3, MMP9, and S100A8 as hub genes with diagnostic and prognostic relevance in sepsis-induced ALI. Experimental findings showed increased HK3 expression in the lung tissue of septic mice, particularly in bronchial and alveolar epithelial cells. In vitro experiments demonstrated increased HK3 levels in lung epithelial cells stimulated with LPS, with cytoplasmic localization near the nucleus. Knockdown of HK3 expression resulted in decreased proliferation activity and glycolytic flux, increased inflammatory response, oxidative stress, and cell apoptosis in lung epithelial cells.

Exploring the Mechanisms of the Huangqin Qingfei Decoction in Pneumonia Treatment for Lipopolysaccharide-Induced Mouse Model: A Network Pharmacology Analysis and Experimental Validation

Objective: In this study, a network pharmacological approach, and in vivo validation experiments were used to screen the active components of the Huangqin Qingfei decoction (HQFD) for their therapeutic efficacy in pneumonia. Methods: The active ingredients and targets of the HQFD were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform, while pneumonia-related targets were obtained from GeneCards, Online Mendelian Inheritance in Man, and the Therapeutic Target Database. A protein–protein interaction network was constructed to identify core targets, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses using Metascape. A lipopolysaccharide-induced pneumonia mouse model was established, and verification of the main targets predicted by network pharmacology and therapeutic efficacy assessment was conducted using pathological staining, Western blot, and polymerase chain reaction. Results: We identified 51 active components in the HQFD that primarily affected pneumonia-related targets AKT1, PTGS2, MAPK14, and MMP9. The outcomes of the pathway enrichment analyses conducted by the KEGG and the GO demonstrated that the HQFD regulates genes associated with pathways in cancer, the lipid and atherosclerosis, as well as the AGE-RAGE signaling pathway in diabetic complications. Experiments on animals demonstrated that the AKT1, PTGS2, MAPK14, and MMP9 mRNA expression and p-AKT, p-P38, PTGS2, and MMP9 protein expression in lung tissues were significantly upregulated in the model group when compared to the control group. AKT1, PTGS2, MAPK14, MMP9 mRNA expression, and p-AKT, p-P38, PTGS2, and MMP9 protein expression were all lower in the HQFD groups in contrast to the model group. Conclusions: The effects of HQFD on pneumonia can be attributed to its active ingredients, including beta-sitosterol, wogonin, and quercetin. The mechanism involves multiple targets and pathways, including the PI3K-AKT signaling pathway, MAPK signaling pathway, and NF-kappa B signaling pathway, and modulation of AKT, PTGS2, MAPK14, and MMP9 expression. The HQFD reduces inflammatory damage and pneumonia symptoms via these mechanisms.

Promotion of transcription factor EB-dependent autophagic process by curcumin alleviates arsenic-caused lung oxidative stress and inflammation in mice

Arsenic is a human carcinogen widely distributed in the environment, and arsenic exposure from drinking water has received widespread attention as a global public health problem. Curcumin is a natural bioactive substance with high efficiency and low toxicity extracted from turmeric, which has a variety of biological properties such as anti-oxidation, anti-inflammation, anti-cancer, and immuno-modulatory activities. Curcumin is widely used in daily life as a food additive and dietary supplement. However, its protective effects in lung injuries by chronic arsenic exposure orally remain unexplored. In this study, curcumin treatment not only significantly accelerated arsenic elimination and improved lung tissue morphology, but also decreased arsenic-generated ROS by activating Nrf2 and its down-stream antioxidants. Further, curcumin alleviated inflammatory changes in mice exposed to arsenic for 6 and 12 weeks, as manifested by lung MPO levels, total protein and cellular levels in bronchoalveolar lavage fluid (BALF), serum IL-4 levels, and MAPK/NF-κB expression in lung tissue. Notably, our study also confirmed that curcumin could promote the expression and nuclear translocation of the transcription factor EB (TFEB), as well as activate TFEB-regulated autophagy in lung tissue of arsenic-treated mice, accompanied by inhibition of the AKT-mTOR signaling pathway. Overall, our study here suggests that natural bioactive compound curcumin could alleviate arsenic-induced pulmonary oxidative stress and inflammation in vivo, which is closely related to enhanced TFEB activity and induction of the autophagic process.

Chronic intermittent hypoxia leads to disruption of clock genes expression in mouse lung tissues: Potential consequences on lung cell senescence

Chronic intermittent hypoxia leads to disruption of clock genes expression in mouse lung tissues: Potential consequences on lung cell senescence

Ubiquitin Ligase Parkin Regulates the Stability of SARS-CoV-2 Main Protease and Suppresses Viral Replication.

The highly infectious coronavirus SARS-CoV-2 relies on the viral main protease (Mpro, also known as 3CLpro or Nsp5) to proteolytically process the polyproteins encoded by the viral genome for the release of functional units in the host cells to initiate viral replication. Mpro also interacts with host proteins of the innate immune pathways, such as IRF3 and STAT1, to suppress their activities and facilitate virus survival and proliferation. To identify the host mechanism for regulating Mpro, we screened various classes of E3 ubiquitin ligases and found that Parkin of the RING-between-RING family can induce the ubiquitination and degradation of Mpro in the cell. Furthermore, when the cells undergo mitophagy, the PINK1 kinase activates Parkin and enhances the ubiquitination of Mpro. We also found that elevated expression of Parkin in the cells significantly decreased the replication of SARS-CoV-2 virus. Interestingly, SARS-CoV-2 infection downregulates Parkin expression in the mouse lung tissues compared to healthy controls. These results suggest an antiviral role of Parkin as a ubiquitin ligase targeting Mpro and the potential for exploiting the virus-host interaction mediated by Parkin to treat SARS-CoV-2 infection.

ANXA3 interference inactivates ERK/ELK1 pathway to mitigate inflammation and apoptosis in sepsis-associated acute lung injury

Acute lung injury (ALI) is a prevailing and deadly complication of sepsis coupled with increasing incidence and fatality rate. Annexin A3 (ANXA3) has been unraveled to be upregulated during sepsis. This study purposed to assess the role and the mechanism of ANXA3 in sepsis-induced ALI. After the construction of mouse model of sepsis, the pathological changes of mice lung tissues were estimated by H&E staining. ANXA3 expression in mice lung tissues and serum was examined. The degree of pulmonary edema and the levels of inflammatory factors in bronchoalveolar lavage fluid (BALF) were analyzed. In lipopolysaccharide (LPS)-induced mouse ALI model in vitro, CCK-8 assay measured cell viability and flow cytometry analysis detected cell apoptosis. Besides, ELISA assay detected the release of inflammatory cytokines. Western blot analyzed the expression of proteins associated with inflammation, apoptosis and extracellular-signal-regulated kinase (ERK)/ETS-like gene 1 (ELK1) signaling. Results revealed that ANXA3 was overexpressed in the lung tissues and serum of septic mice. Following the knockdown of ANXA3, sepsis-induced lung injury was alleviated, manifested as reduced lung edema, decreased inflammatory cell infiltration and inhibited cell apoptosis. Additionally, ANXA3 silence blocked ERK/ELK1 signaling both in sepsis mouse models and in vitro model of ALI induced by lipopolysaccharide (LPS). Moreover, the inhibitory effects of ANXA3 silencing on ERK/ELK1 signaling activation, the viability damage, inflammation and apoptosis in LPS-induced mouse ALI model in vitro were partially reversed by ERK activator. Collectively, depletion of ANXA3 exerted suppressive effects on the inflammation and apoptosis in sepsis-induced ALI through blocking ERK/ELK1 signaling.

Localized Administration of Bcar3 siRNA via Nano-Self-Assembly to Treat Idiopathic Pulmonary Fibrosis by Disrupting Macrophage-Fibroblast Crosstalk.

Idiopathic pulmonary fibrosis (IPF) is a severe interstitial lung disease characterized by chronic lung injury leading to macrophage infiltration and fibroblast activation. However, there is no effective therapeutic strategy targeting the crucial crosstalk between macrophages and fibroblasts to halt IPF progression. Studies were conducted in IPF patients and fibrotic mice models to elucidate the role of Bcar3 in the pathogenesis of pulmonary fibrosis. The effect of Bcar3 on macrophage polarization, fibroblast activation, and related signaling pathways were next investigated to unravel the underlying mechanisms. Our study elucidates a marked increase in Bcar3 expression in lung tissues from IPF patients and fibrotic mice, recording 1.7 and 7.8-fold increases compared to control subjects, respectively. Additionally, Bcar3 was found to significantly enhance macrophage activation and fibroblast differentiation, observable in both in vivo and in vitro settings. Mechanistically, the upregulation of Bcar3 in macrophages was reliant on Stat6, while in fibroblasts, it depended on TGFβR1/Smad3. Furthermore, Bcar3 augmented IL-4/Stat6 pathway in macrophages and TGF-β/Smad3 pathway in fibroblasts, supporting a synergistic activation loop that expedited lung fibrogenesis. Notably, intratracheal injection of liposomes containing Bcar3 siRNA precisely delivered gene therapeutics to lung macrophages and fibroblasts, effectively reducing Bcar3 expression to 59% of baseline levels. Importantly, this intervention protected mice from lung fibrosis induced by either FITC or bleomycin, as well as human precision-cut lung slices against TGF-β1 stimulation. Our study underscores the pivotal role of Bcar3 in orchestrating the macrophage-fibroblast crosstalk during pulmonary fibrosis progression. Targeting Bcar3 emerges as a novel therapeutic avenue to halt IPF progression and enhance patient prognosis.

Pirfenidone and nintedanib attenuate pulmonary fibrosis in mice by inhibiting the expression of JAK2.

Pirfenidone and nintedanib were approved by the Food and Drug Administration (FDA) for the treatment of idiopathic pulmonary fibrosis (IPF). These two drugs can slow the progression of the disease, but the specific mechanisms are not fully understood. In the current study, bleomycin (BLM) induced pulmonary fibrosis in mice was accompanied by high p-JAK2 expression in lung tissue, mainly in the nucleus. The expression of p-JAK2 significantly decreased after intragastric administration of pirfenidone and nintedanib. p-JAK2 is reportedly expressed mainly in the cytoplasm and exerts its effect by activating downstream p-STAT3 in the nucleus. In vivo experiments, pulmonary fibrosis was induced in mice with BLM and then treated with pirfenidone and nintedanib. The levels of transforming growth factor-β (TGF-β1), SP-A, SP-D and KL-6 in serum were measured by enzyme-linked immunosorbent assay (ELISA). Pathological staining was performed to assess lung fibrosis in mice, Western blot was performed to detect the expression levels of relevant proteins, and immunofluorescence was performed to observe the fluorescence expression of p-JAK2. In cellular experiments, MLE12 was stimulated with TGF-β1 and intervened with TGF-β1 receptor inhibitor and si-JAK2, pirfenidone and nintedanib, respectively, and the related protein expression levels were detected by Western blot. In both in vivo and in vitro experiments, pirfenidone and nintedanib were found to attenuate the expression of lung fibrosis markers by inhibiting the expression of JAK2, which may reduce the entry of p-JAK2 into the nucleus by downregulating JAK2 phosphorylation through inhibition of the TGF-β receptor. In contrast, inhibition of JAK2 expression greatly reduced the expression of TGF-β receptor and α-smooth muscles actin (a myofibroblast activation marker). In both in vivo and in vitro experiments, the present study demonstrated that TGF-β1 promotes JAK2 phosphorylation through a non-classical pathway, and conversely, inhibition of JAK2 expression affects the TGF-β1 signalling pathway. Therefore, we speculate that TGF-β1 and JAK2 signaling pathways interact with each other and participate in fibrosis.

Therapeutic potential of dihydroartemisinin in mitigating radiation-induced lung injury: Inhibition of ferroptosis through Nrf2/HO-1 pathways in mice.

Radiation-induced lung injury (RILI) is a common consequence of thoracic radiation therapy that lacks effective preventative and treatment strategies. Dihydroartemisinin (DHA), a derivative of artemisinin, affects oxidative stress, immunomodulation, and inflammation. It is uncertain whether DHA reduces RILI. In this work, we investigated the specific mechanisms of action of DHA in RILI. Twenty-four C57BL/6J mice were randomly divided into four groups of six mice each: Control group, irradiation (IR) group, IR + DHA group, and IR + DHA + Brusatol group. The IR group received no interventions along with radiation treatment. Mice were killed 30 days after the irradiation. Morphologic and pathologic changes in lung tissue were observed with hematoxylin and eosinstaining. Detection of hydroxyproline levels for assessing the extent of pulmonary fibrosis. Tumor necrosis factor α (TNF-α), transforming growth factor-β (TGF-β), glutathione peroxidase (GPX4), Nuclear factor erythroid 2-related factor 2 (Nrf2), and heme oxygenase-1 (HO-1) expression in lung tissues were detected. In addition, mitochondrial ultrastructural changes in lung tissues were also observed, and the glutathione (GSH) content in lung tissues was assessed. DHA attenuated radiation-induced pathological lung injury and hydroxyproline levels. Additionally, it decreased TNF-α and TGF-β after irradiation. DHA may additionally stimulate the Nrf2/HO-1 pathway. DHA upregulated GPX4 and GSH levels and inhibited cellular ferroptosis. Brusatol reversed the inhibitory effect of DHA on ferroptosis and its protective effect on RILI. DHA modulated the Nrf2/HO-1 pathway to prevent cellular ferroptosis, which reduced RILI. Therefore, DHA could be a potential drug for the treatment of RILI.

Hydroxytyrosol Alleviates Methotrexate-Induced Pulmonary Fibrosis in Rats: Involvement of TGF-β1, Tissue Factor, and VEGF.

Methotrexate (MTX) is an indispensable drug used for the treatment of many autoimmune and cancerous diseases. However, its clinical use is associated with serious side effects, such as lung fibrosis. The main objective of this study is to test the hypothesis that hydroxytyrosol (HT) can mitigate MTX-induced lung fibrosis in rats while synergizing MTX anticancer effects. Pulmonary fibrosis was induced in the rats using MTX (14 mg/kg/week, per os (p.o.)). The rats were treated with or without HT (10, 20, and 40 mg/kg/d p.o.) or dexamethasone (DEX; 0.5 mg/kg/d, intraperitoneally (i.p.)) for two weeks concomitantly with MTX. Transforming growth factor beta 1 (TGF-β1), interleukin-4 (IL-4), thromboxane A2 (TXA2), vascular endothelial growth factor (VEGF), 8-hydroxy-2-deoxy-guanosine (8-OHdG), tissue factor (TF) and fibrin were assessed using enzyme-linked immunosorbent assay (ELISA), immunofluorescence, and RT-PCR. Pulmonary fibrosis was manifested by an excessive extracellular matrix (ECM) deposition and a marked increase in TGF-β1 and IL-4 in lung tissues. Furthermore, cotreatment with HT or dexamethasone (DEX) significantly attenuated MTX-induced ECM deposition, TGF-β1, and IL-4 expression. Similarly, HT or DEX notably reduced hydroxyproline contents, TXA2, fibrin, and TF expression in lung tissues. Moreover, using HT or DEX downregulated the gene expression of TF. A significant decrease in lung contents of VEGF, IL-8, and 8-OHdG was also observed in HT + MTX- or DEX + MTX -treated animals in a dose-dependent manner. Collectively, the results of our study suggest that HT might represent a potential protective agent against MTX-induced pulmonary fibrosis.

Pro-Fibrotic Effects of CCL18 on Human Lung Fibroblasts Are Mediated via CCR6.

Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease of unknown origin, with a median patient survival time of ~3 years after diagnosis without anti-fibrotic therapy. It is characterized by progressive fibrosis indicated by increased collagen deposition and high numbers of fibroblasts in the lung. It has been demonstrated that CCL18 induces collagen and αSMA synthesis in fibroblasts. We aimed to identify the CCL18 receptor responsible for its pro-fibrotic activities. We used a random phage display library to screen for potential CCL18-binding peptides, demonstrated its expression in human lungs and fibroblast lines by PCR and immunostaining and verified its function in cell lines. We identified CCR6 (CD196) as a CCL18 receptor and found its expression in fibrotic lung tissue and lung fibroblast lines derived from fibrotic lungs, but it was almost absent in control lines and tissue. CCL18 induced receptor internalization in a CCR6-overexpressing cell line. CCR6 blockade in primary human lung fibroblasts reduced CCL18-induced FGF2 release as well as collagen-1 and αSMA expression. Knockdown of CCR6 in a mouse fibroblast cell line abolished the induction of collagen and α-smooth muscle actin expression. Our data indicate that CCL18 triggers pro-fibrotic processes via CCR6, highlighting its role in fibrogenesis.

A novel flavonol-polysaccharide from Tamarix chinensis alleviates influenza A virus-induced acute lung injury. Evidences for its mechanism of action

BackgroundTamarix chinensis Lour. is a Chinese medicine used for treating inflammation-related diseases and its crude polysaccharides (MBAP90) exhibited significant anticomplement activities in vitro. PurposeTo obtain anticomplement homogenous polysaccharides from MBAP90 and explore its therapeutic effects and potential mechanism on influenza A virus (IAV)-induced acute lung injury (ALI). MethodsAnticomplement activity-guided fractionation of the water-soluble crude polysaccharides from the leaves and twigs of T. chinensis were performed by diethylaminoethyl-52 (DEAE-52) cellulose and gel permeation columns to yield a homogeneous polysaccharide MBAP-5, which was further characterized using ultra-high-performance liquid chromatography-ion trap tandem mass spectrometry (UPLC-IT-MS) and nuclear magnetic resonance (NMR) analysis. In vitro, the anticomplement activity of MBAP-5 through classical pathway was measured using a hemolytic test. The therapeutic effects of MBAP-5 on ALI were evaluated in H1N1-infected mice. H&E staining, enzyme linked immunosorbent assay (ELISA), immunohistochemistry, and western blot were used to systematically access lung histomorphology, inflammatory cytokines, degree of complement component 3c, 5aR, and 5b-9 (C3c, C5aR, and C5b-9) deposition, and inflammasome signaling pathway protein expressions in lung tissues. ResultsMBAP-5 was a novel flavonol-polysaccharide with the molecular weight (Mw) of 153.6 kDa. Its structure was characterized to process a backbone of →4)-α-D-GlcpA-(1→, →6)-α-D-Glcp-(1→, →3,4)-α-D-Glcp-(1→, →3,4,6)-α-D-Glcp-(1→, and →4,6)-β-D-Glcp-(1→, as well as branches of α-L-Araf-(1→ and β-D-Galp-(1→. Particularly, O-3 of →3,4,6)-α-D-Glcp-(1→ was substituted by quercetin. In vitro assay showed that MBAP-5 had a potent anticomplement activity with a CH50 value of 102 ± 4 µg/ml. Oral administration of MBAP-5 (50 and 100 mg/kg) effectively attenuated the H1N1-induced pulmonary injury in vivo by reducing pulmonary edema, virus replication, and inflammatory responses. Mechanistically, MBAP-5 inhibited the striking deposition and contents of complement activation products (C3c, C5aR, and C5b-9) in the lung. Toll-like receptor 4 (TLR4) /transcription factor nuclear factor κB (NF-κB) signaling pathway was constrained by MBAP-5 treatment. In addition, MBAP-5 could suppress activation of the inflammasome pathways, including Nod‐like receptor pyrin domain 3 (NLRP3), cysteinyl aspartate specific proteinase-1/12 (caspase-1/12), apoptosis‑associated speck‑like protein (ASC), gasdermin D (GSDMD), interleukin (IL)-1β, and IL-18 expressions. ConclusionsA novel flavonol-polysaccharide MBAP-5 isolated from T. chinensis demonstrated a therapeutic effect against ALI induced by IAV attack. The mechanism might be associated with inhibition of complement system and inflammasome pathways activation.

Adenosine diphosphate ribosylation factor 6 inhibition protects burn sepsis induced lung injury through preserving vascular integrity and suppressing ASC inflammasome transmission

BackgroundSevere burns are devastating injuries with significant immune dysfunction and result in substantial mortality and morbidity due to sepsis induced organ failure. Acute lung injury is the most common type of organ injury in sepsis, however, the mechanisms of which are poorly understood and effective therapeutic measures are limited. This study is aimed to investigate the effect of a small Guanosine triphosphatase (GTPase), Adenosine diphosphate ribosylation factor 6 (ARF6), on burn sepsis induced lung injury, and discuss the possible mechanisms. MethodsBurn sepsis was established in male C57BL/6 mice. Mice were anesthetised by intramuscular injection of ketamine and xylazine hydrochloride, then 30% TBSA full thickness burn followed by sub-eschar injection of lipopolysaccharide. Animals were treated with intraperitoneal injection of a small molecule inhibitor of ARF6: NAV-2729, or vehicle, right after the burn and sepsis stimuli were inflicted. Lung tissues were harvested for histopathological observation and the acute lung injury scores were calculated. Organ permeability, Vascular Endothelial Cadherin (VE-cadherin) expression, inflammatory cytokine levels and myeloperoxidase activity in lung tissues were detected. Rat pulmonary microvascular endothelial cells (PMVECs) were stimulated by burn sepsis serum with or without 10 μM NAV-2729. The ARF6 activation, VE-cadherin expression, inflammasome activity, adapter protein apoptosis speck-like protein containing a caspase recruiting domain (ASC) specks and cytokines secretion were determined. Student’s t test was used for comparison between two groups. Multiple comparisons among groups were performed by using analysis of variance, with Tukey’s test for the post hoc test. ResultsNAV-2729 treatment attenuated burn sepsis induced lung injury and promoted survival of burn septic mice by preserving VE-cadherin expression in endothelial cell adherent junction and limited vascular hyperpermeability in lung tissues. Moreover, inflammatory cytokine expression and inflammatory injury in lung tissues were alleviated. Mechanistically, NAV-2729 enhanced vascular integrity by inhibiting ARF6 activation and restoring VE-cadherin expression in PMVECs. In addition, NAV-2729 inhibited ARF6-dependent phagocytosis of ASC specks, thus preventing inflammation propagation mediated by cell-to-cell transmission of ASC specks. ConclusionsARF6 inhibition preserved vascular integrity by restoring expression of VE-cadherin and suppressed the spread of inflammation by affecting phagocytosis of ASC specks, thus protected against sepsis induced lung injury and improve survival of burn septic animals. The findings of this study implied potential therapeutics by which ARF6 inhibition can protect lung function from septic induced lung injury and improve outcomes in burn sepsis.

Proteomic analysis of pulmonary arteries and lung tissues from dogs affected with pulmonary hypertension secondary to degenerative mitral valve disease.

In dogs with degenerative mitral valve disease (DMVD), pulmonary hypertension (PH) is a common complication characterized by abnormally elevated pulmonary arterial pressure (PAP). Pulmonary arterial remodeling is the histopathological changes of pulmonary artery that has been recognized in PH. The underlying mechanisms that cause this arterial remodeling are poorly understood. This study aimed to perform shotgun proteomics to investigate changes in protein expression in pulmonary arteries and lung tissues of DMVD dogs with PH compared to normal control dogs and DMVD dogs without PH. Tissue samples were collected from the carcasses of 22 small-sized breed dogs and divided into three groups: control (n = 7), DMVD (n = 7) and DMVD+PH groups (n = 8). Differentially expressed proteins were identified, and top three upregulated and downregulated proteins in the pulmonary arteries of DMVD dogs with PH including SIK family kinase 3 (SIK3), Collagen type I alpha 1 chain (COL1A1), Transforming growth factor alpha (TGF-α), Apoptosis associated tyrosine kinase (AATYK), Hepatocyte growth factor activator (HGFA) and Tyrosine-protein phosphatase non-receptor type 13 (PTPN13) were chosen. Results showed that some of the identified proteins may play a role in the pathogenesis of pulmonary arterial remodeling. This study concluded shotgun proteomics has potential as a tool for exploring candidate proteins associated with the pathogenesis of PH secondary to DMVD in dogs.

Identification and prognostic biomarkers among ZDHHC4/12/18/24, and APT2 in lung adenocarcinoma

S-palmitoylases and S-depalmitoylases are differentially expressed in various cancers and several malignant tumors and show a strong prognostic ability. Notwithstanding, the potential clinical impact of S-palmitoylases and S-depalmitoylases, particularly in the prognosis and progression of lung adenocarcinoma (LUAD), has not been clarified. Expression levels of S-palmitoylases and S-depalmitoylases in LUAD were investigated using TCGA. GEPIA was used to evaluate the mRNA levels of S-palmitoylases and S-depalmitoylases at different pathological stages. Metascape was used to investigate the biological significance of S-palmitoylases and S-depalmitoylases. The Kaplan–Meier plotter was used to analyze the prognostic value of S-palmitoylases and S-depalmitoylases. CBioportal was used to analyze gene alterations in S-palmitoylases and S-depalmitoylases. UALCAN was used to examine DNA promoter methylation levels of S-palmitoylases and S-depalmitoylases. Finally, we investigated the relationship between S-palmitoylases, S-depalmitoylases, and tumor-infiltrating immune cells using TIMER. Correlations with immune checkpoint-related genes were determined using the R packages reshape2, ggpubr, ggplot2, and corrplot. PCR was also performed to assess the degree of ZDHHC4/12/18/24 and APT2 transcript expression in lung adenocarcinoma and adjacent normal lung tissues. HPA was utilized to investigate protein levels of S-palmitoylases and S-depalmitoylases in LUAD and normal lung tissue. Our study found that ZDHHC2/3/4/5/6/7/9/12/13/16/18/20/21/23/24, APT1/2, PPT1, LYPLAL1, ABHD4/10/11/12/13 and ABHD17C mRNA expression was significantly upregulated in LUAD, whereas ZDHHC1/8/11/11B/14/15/17/19/22, ABHD6/16A and ABHD17A mRNA expression was significantly downregulated. The functions of the differentially expressed S-palmitoylases and S-depalmitoylases were mainly associated with protein-cysteine S-palmitoyltransferase and protein-cysteine S-acyltransferase activities. Patients with high expression of ZDHHC4/12/18/24, APT2, ABHD4, ABHD11 and ABHD12 had a shorter overall survival. Infiltration of six immune cells (B cells, CD8+ T cells, CD4+ T cells, macrophages, neutrophils, and dendritic cells) was closely associated with the expression of ZDHHC4/12/18/24 and APT2. ZDHHC4/12/18/24 and APT2 positively correlated with the immune checkpoint-related gene CD276. We assessed the mRNA levels of ZDHHC4/12/18/24 and APT2 using qRT-PCR and found increased expression of ZDHHC4/12/18/24 in LUAD compared with healty control lung tissues. ZDHHC4/12/18/24, and APT2 are potential prognostic biomarkers of LUAD. Their expression levels could be related to the tumor microenvironment in LUAD.

Quercetin alleviates PM2.5-induced chronic lung injury in mice by targeting ferroptosis.

PM2.5 is a well-known harmful air pollutant that can lead to acute exacerbation and aggravation of respiratory diseases. Although ferroptosis is involves in the pathological process of pulmonary disease, the potential mechanism of ferroptosis in PM2.5-caused lung inflammation and fibrosis need to be further clarified. Quercetin is a phenolic compound that can inhibit ferroptosis in various diseases. Hence, this study explores the role of ferroptosis in lung injury induced by PM2.5 in order to further elucidate the beneficial effect of quercetin and its underlying mechanism. C57BL/6J mice were treated with either saline or PM2.5 by intratracheal instillation 20 times (once every two days). Additionally, PM2.5-treated mice were supplemented with two doses of quercetin. Lung injury, lipid peroxidation, iron content and ferroptosis marker protein expression and the Nrf2 signaling pathway were evaluated. In vitro, cell experiments were applied to verify the mechanisms underlying the links between Nrf2 signaling pathway activation and ferroptosis as well as between ferroptosis and inflammation. In vivo, PM2.5 increased lung inflammation and caused lung fibrosis and increased lipid peroxidation contents, iron contents and ferroptosis markers in lung tissues; these effects were significantly reversed by quercetin. Additionally, quercetin upregulated the nuclear Nrf2 expression and downregulated Keap1 expression in lung tissues of PM2.5-exposed mice. Quercetin decreased lipid peroxidation products, iron contents and ferroptosis levels and increased the nuclear translocation of Nrf2 and the degradation of Keap1 in PM2.5-exposed BEAS-2B cells. Moreover, we found that quercetin and dimethyl fumarate markedly decreased lipid peroxidation production and ferroptosis by activating the Nrf2-Keap1 pathway in PM2.5-exposed cells. Furthermore, quercetin reduced inflammatory cytokines and TGF-β1 in PM2.5-exposed cells. Our data suggested that Nrf2 is involved in ferroptosis in PM2.5-induced lung injury, and quercetin can alleviate these adverse effects via activating Nrf2-Keap1 signaling pathway.

Effects of alamandine on monocrotaline-induced pulmonary hypertension in rats.

Pulmonary arterial hypertension (PAH) is a severe and often fatal disease that is associated with oxidative stress and inflammation. Alamandine, a component of the renin-angiotensin system, known for its antioxidative, anti-inflammatory, and antifibrotic effects, has been investigated in this study to determine if it has protective effects against PAH induced by monocrotaline (MCT) and if these effects are associated with oxidative stress, inflammatory factors, and inducible nitric oxide synthase (iNOS). Rats were administered MCT (40 mg/kg) on day 0 and then received alamandine (50 mg/kg/day) via mini-osmotic pumps for 21 days starting one day later. Hemodynamic parameters, electrocardiograms, superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA), inflammatory cytokines (TNF-α, IL-1β, and NF-κB), iNOS, and MrgD receptor expression in lung tissue were evaluated at the end of the 21-day period. The MrgD receptor was quantified through immunofluorescent staining, and the histopathology of lung tissues was evaluated using hematoxylin and eosin staining. The results showed that alamandine treatment significantly improved hemodynamic parameters, oxidative stress markers, inflammatory factors, and electrocardiographic data. Furthermore, treatment with alamandine decreased the levels of iNOS. Additionally, alamandine treatment decreased the expression levels of MrgD receptors in the lung tissue of MCT-induced PAH. In summary, this study indicates that alamandine has protective effects against monocrotaline-induced PAH, and these effects may be attributed to the inhibition of oxidative stress, inflammatory parameters, and iNOS.

Remimazolam attenuates inflammation in bronchopneumonia through the inhibition of NLRP3 activity by PDPK1 ubiquitination.

Bronchopneumonia is the most common pneumonia in childhood. Therefore, we tested the effects of Remimazolam presented Bronchopneumonia and its possible mechanisms. Phillygenin increased survival rate, reduced W/D ratio, and lung injury score, and inhibited IL-1β, IL-6, TNF-α, and INF-γ levels in mice model of bronchopneumonia. Remimazolam induced PDPK1 and p-AKT protein expressions, and suppressed NLRP3 protein expression in lung tissue of mice model. Invitro model, Remimazolam also induced PDPK1 and p-AKT protein expressions, and suppressed NLRP3 protein expression. Remimazolam also inhibited inflammation levels invitro model. PDPK1 inhibitor, PHT-427 (100 mg/kg) reduced survival rate, increased W/D ratio and lung injury score, and promoted inflammation levels in mice model of bronchopneumonia by treated with Remimazolam. PHT-427 suppressed PDPK1 and p-AKT protein expressions and induced NLRP3 protein expression in mice model of bronchopneumonia by treated with Remimazolam. Remimazolam interlinked PDPK1 protein. Remimazolam increased the expressions of PDPK1 and p-AKT invitro model. Remimazolam reduced PDPK1 ubiquitination invitro model.