Airways Of Mice Research Articles

High-fat diet and palmitic acid amplify airway type 2 inflammation.

Metabolic dysfunction such as elevated levels of saturated fatty acids (SFA) may play a role in obese asthma, but its contribution to airway inflammation remains unclear. We sought to determine the role of high-fat diet (HFD) and palmitic acid (PA), a major form of SFA, in regulating type 2 inflammation. Airway samples from asthma patients with or without obesity, mouse models and human airway epithelial cell culture were utilized to test if SFA amplify type 2 inflammation. Asthma patients with obesity had higher levels of airway PA than asthma patients without obesity. HFD increased the levels of PA in mice, and subsequently enhanced IL-13-induced airway eosinophilic inflammation. PA treatment amplified airway eosinophilic inflammation in mice that were previously exposed to IL-13 or house dust mite. IL-13 alone or in combination with PA increased dipeptidyl peptidase 4 (DPP4) release (soluble DPP4) and/or activity in mouse airways and human airway epithelial cells. Inhibition of DPP4 activity by linagliptin in mice pre-exposed to IL-13 or both IL-13 and PA increased airway eosinophilic and neutrophilic inflammation. Our results demonstrated the exaggerating effect of obesity or PA on airway type 2 inflammation. Up-regulation of soluble DPP4 by IL-13 and/or PA may serve as a mechanism to prevent excessive type 2 inflammation. Soluble DPP4 may have the therapeutic potential in asthma patients with obesity who have an endotype with mixed airway eosinophilic and neutrophilic inflammation.

Direct activation of Airway Sensory C-fibers by SARS-CoV-2 Spike protein

Activation of sensory nerves, in particular nociceptive C-fibers, is a feature of most respiratory viruses. Evidence of such activation is found in the classical consequences of C-fiber activation including sneezing, sore throat, coughing, and reflex secretions. As well as causing the troubling symptoms of viral infection, the activation of these nerves allows viruses to escape the body and be transmitted to other hosts, i.e. nociceptor activation amplifies spread in a community. In addition, activation of airway vagal C-fibers can lead to strong reflex bronchoconstriction and excessive secretions that likely contribute to the exacerbation of asthma particularly in children. In theory, viral infection leads to C-fiber activation by two general mechanisms. The first is that viral infection of epithelial cells leads to the production of a mediator(s) that stimulates the C-fiber terminals. The second is that the virus itself directly activates the nerves. We used three orthogonal approaches to address our hypothesis that the coronavirus spike protein directly activates (evokes action potential discharge) vagal C-fibers in mouse airways. We used male C57/BL6 mice to evaluate whether coronavirus spike protein interacts; directly with C-fiber terminals in an isolated vagus-trachea/lung preparation. Action potentials were recorded using extracellular electrodes positioned near the relevant cell bodies in the vagal sensory nodose/jugular ganglion complex. Spike protein S1 (4ug/ml) was perfused through the mouse trachea into the receptive field of an identified C-fiber (c.v. <1 m/s). 40% of C-fibers (12 out of 30) responded acutely to the S1 protein with action potential discharge. In the same type of ex-vivo experimental set-up, two-photon imaging technique was also used to evaluate the % of fibers activated by spike protein. Consistent with our electrophysiological studies we found that 45% of the C-fibers (241 fibers out of 538 fibers in 4 experiments) were stimulated by the S1protein. To determine whether spike protein was indirectly activating C-fibers in the lung, i.e. requiring other cells within the airway tissue, we evaluated its effect on neurons dissociated from the vagal ganglia with Fura-2 calcium imaging. Again, ~ 40% of the isolated neurons (65 out of 163 neurons) responded to spike protein. These data support our hypothesis that the spike protein can directly activate neurons independently of airway tissue. This supports the hypothesis that some of the nerve-related effects of SARS CoV-2 infection may involve the direct activation of sensory C-fibers. R35 HL144887 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Chemokine CCL19 promotes type 2 helper T cell differentiation and allergic airway inflammation

Abstract Allergic asthma is largely driven by the actions of allergen-specific CD4 +type 2 T helper (Th2) cells. Following allergen inhalation, these cells are induced by the interaction of naïve CD4 +T cells with antigen-bearing dendritic cells (DCs) in lung-draining lymph nodes (LNs). The molecular mechanisms underlying the induction of T helper cells have been well studied and include the migration of allergen-bearing DCs from the lung to regional LNs in a manner dependent on the chemokine receptor, CCR7. Previous studies have demonstrated that CCL21, a CCR7 ligand, is required for the accumulation of naïve T cells and activated DCs in the LNs, but the function of another ligand, CCL19, has been unknown. In the present study, we investigated the role of CCL19 in allergic airway inflammation using CCL19-deficient mice. In a mouse model of asthma, eosinophils and lymphocytes were significantly reduced in airways of CCL19-deficient mice compared with those of wildtype (WT) animals, and production of IL-4 and IL-13 in the lung was significantly diminished, suggesting that CCL19 is required for type 2 immune responses. Co-cultures of naïve WT CD4 +T cells with CCL19-deficient DCs produced less type 2 cytokines than did T cells co-cultured with WT DCs. Further, fibroblastic reticular cells (FRCs) from CCL19-deficient mouse LNs produced less Th2 cytokine production compared with WT FRCs. Addition of recombinant CCL19 to the culture of naïve CD4 +T cells increased phosphorylation of STAT5 and gene expression of Il2, Il2ra, Il2rb, Il2rg, Sta5a, Stat5b and Stat6. Together, our findings suggest that CCL19 produced by DCs and FRCs promotes Th2 differentiation through the activation of STAT5-signaling, and that this promotes allergic airway inflammation. This work was supported by Intramural Research Program of NIEHS, NIH (ZIA ES102025-09).

Allergen-stimulated secretion of DNA fragments by the airway epithelium enhances type 2 immune responses

Exposure of human bronchial epithelial cells (hBECs) and mouse airways to allergens from the fungus Alternaria alternata triggers the release of alarmin signaling molecules including ATP, IL-33 and DNA fragments. The goal of this study was to determine the mechanisms that regulate release of genomic DNA fragments and how extracellular DNA (eDNA) contributes to type 2 immunity. Treatment of hBECs with Alternaria extract induces oxidative stress, which leads to ATP release and uptake of Ca 2+ following activation of P2X receptors. DNA release was dependent on a sustained increase in intracellular [Ca 2+ ], which stimulates cleavage of caspase 3 by the proprotein convertase enzyme furin, resulting in activation of caspase 3 and ultimately nuclear DNA fragmentation as shown in comet assays. Some of the fragmented DNA was released into the media under conditions where barrier function of the epithelium was maintained, and nuclear and plasma membranes remained intact. Alternaria-induced DNA release was detected within 30 minutes following exposure, reached its peak at 4 hours and ceased after 12 hours. Pretreatment of hBECs with a calpain inhibitor resulted in continued DNA release and loss of cell viability by 24 hours. This result suggested that calpain activity contributed to termination of Alternaria-evoked DNA release. In mice, intranasal administration of Alternaria extract produced release of DNA fragments into bronchoalveolar lavage (BAL) fluid over a similar time course as observed in hBECs. In caspase 3 deficient mice however, DNA release into BAL fluid was significantly reduced. Furthermore, Alternaria-induced type 2 immune responses were suppressed following pretreatment with a DNA scavenger. Conversely, intranasal administration of mouse genomic DNA with Alternaria amplified IL-5 and IL-13 secretion into BAL fluid while DNA alone had no effect. These findings indicate that DNA release from the airway epithelium is a regulated process involving noncanonical activation of caspase 3 to produce DNA fragmentation and calpain activity, which appears to be essential for the termination of DNA release. Moreover in airways, eDNA appears to function as a secondary alarmin that acts on immune cells to enhance type 2 immunity. This work was supported by grants from the NIH (AI128729 and AI169530) to HK and SMO. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Targeting the RhoA-GEF-H1 pathway of mast cells attenuates experimental airway allergy

Mast cells are the major effector cells in allergic diseases. RhoA and its downstream pathway is associated with the pathogenesis of airway allergy. The objective of this study is to test a hypothesis that modulating the RhoA-GEF-H1 axis in mast cells can attenuate airway allergy. An airway allergic disorder (AAD) mouse model was employed. Mast cells were isolated from AAD mouse airway tissues to be analyzed by RNA sequencing. We observed that mast cells isolated from the respiratory tract of AAD mice were resistant to apoptosis. Mast cell mediator levels in nasal lavage fluid were correlated with apoptosis resistance in AAD mice. Activation of RhoA in AAD mast cells was related to resistance to apoptosis. Mast cells isolated from the airway tissues in AAD mouse exhibited strong RhoA-GEF-H1 expression. The RhoA-GEF-H1 axis was associated with the lower FasL expression in AAD mast cells. Activation of the RhoA-GEF-H1 axis promoted the production of mediators in mast cells. Inhibition of GEF-H1 facilitated the SIT-induced mast cell apoptosis and enhanced the therapeutic efficacy of AAD. In conclusion, RhoA-GEF-H1 activities are associated with resistance to apoptosis in mast cells isolated from sites of allergic lesions. The state of apoptosis resistance in mast cells is associated with the state of AAD disease. Inhibition of GEF-H1 restores the sensitivity of mast cells to apoptosis inducers, and alleviates experimental AAD in mice.

Whole-Cell MALDI-ToF MS Coupled with Untargeted Metabolomics Facilitates Investigations of Microbial Chemical Interactions.

The emergence of drug-resistant pathogens necessitates the development of new countermeasures. In this regard, the introduction of probiotics to directly attack or competitively exclude pathogens presents a useful strategy. Application of this approach requires an understanding of how a probiotic and its target pathogen interact. A key means of probiotic-pathogen interaction involves the production of small molecules called natural products (NPs). Here, we report the use of whole-cell matrix-assisted laser desorption/ionization time-of-flight (MALDI-ToF) mass spectrometry to characterize NP production by candidate probiotics (mouse airway microbiome isolates) when co-cultured with the respiratory pathogen Burkholderia. We found that a Bacillus velezensis strain inhibits growth of and elicits NP production by Burkholderia thailandensis. Dereplication of known NPs detected in the metabolome of this B. velezensis strain suggests that a previously unannotated bioactive compound is involved. Thus, we present the use of whole-cell MALDI as a broadly applicable method for screening the NP composition of microbial co-cultures; this can be combined with other -omics methods to characterize probiotic-pathogen and other microbe-microbe interactions.

Mouse guanylate-binding protein 1 does not mediate antiviral activity against influenza virus in vitro or in vivo.

Many interferon (IFN)-stimulated genes are upregulated within host cells following infection with influenza and other viruses. While the antiviral activity of some IFN-stimulated genes, such as the IFN-inducible GTPase myxoma resistance (Mx)1protein 1, has been well defined, less is known regarding the antiviral activities of related IFN-inducible GTPases of the guanylate-binding protein (GBP) family, particularly mouse GBPs, where mouse models can be used to assess their antiviral properties in vivo. Herein, we demonstrate that mouse GBP1 (mGBP1) was upregulated in a mouse airway epithelial cell line (LA-4 cells) following pretreatment with mouse IFNα or infection by influenza A virus (IAV). Whereas doxycycline-inducible expression of mouse Mx1 (mMx1) in LA-4 cells resulted in reduced susceptibility to IAV infection and reduced viral growth, inducible mGBP1 did not. Moreover, primary cells isolated from mGBP1-deficient mice (mGBP1-/- ) showed no difference in susceptibility to IAV and mGBP1-/- macrophages showed no defect in IAV-induced NLRP3 (NLR family pyrin domain containing 3) inflammasome activation. After intranasal IAV infection, mGBP1-/- mice also showed no differences in virus replication or induction of inflammatory responses in the airways during infection. Thus, using complementary approaches such as mGBP1 overexpression, cells from mGBP1-/- mice and intranasal infection of mGBP1-/- we demonstrate that mGBP1 does not play a major role in modulating IAV infection in vitro or in vivo.

Poly(I:C) Exacerbates Airway Goblet Cell Hyperplasia and Lung Inflammation in HDM-Exposed Balb/C Mice by YAP/FOXM1 Pathway

Introduction: Respiratory viral infection in childhood is closely associated with asthmatic attacks. Of all predisposing factors, viral infection is the primary contributor to acute childhood asthma exacerbations. However, the mechanisms involved in viral asthma are unclear. This study attempted to provide insights into molecular mechanisms in respiratory virus-induced acute asthma exacerbations. Methods: House dust mite (HDM) was given by intranasal administration to induce asthma in mice. Poly(I:C) was used to mimic the viral infection. A selective YAP inhibitor, verteporfin (VP), was used to investigate the role of the YAP/FOXM1 pathway. The expression of YAP, FOXM1, cytokines, and inflammatory cells in lung tissue, and bronchoalveolar lavage fluid (BALF) was determined using RT-PCR, immunohistochemical, ELISA, and flow cytometry studies. The methacholine challenge assesses airway hyperresponsiveness. In 16HBE cell experiments, we selectively inhibited YAP and FOXM1 by VP and RCM1, respectively, and detected the expression of YAP and FOXM1. Results: The experimental studies have confirmed the YAP/FOXM1 pathway plays a vital role in the differentiation and proliferation of airway club cells into goblet cells and lung inflammation. Poly(I:C) upregulated the expression of FOXM1 by activating transcription factor YAP in mice airway epithelial cells and then promoted the expression of downstream transcription factors SPDEF/MUC5AC, resulting in airway mucus hypersecretion and hyperresponsiveness. In addition, Poly(I:C) facilitates the expression of inflammatory factors in lung tissue. All of these events induce asthma exacerbations. The in vitro studies have confirmed that YAP positively regulates FOXM1 in airway epithelial cells. Conclusion: Poly(I:C) promotes airway epithelial goblet cell hyperplasia, mucus hypersecretion, and airway hyperresponsiveness. It also upregulates the expression of inflammatory factors in lung tissue and BALF in asthmatic mice by the YAP/FOXM1 pathway, resulting in asthma attacks.

Role of NaV 1.9 in inflammatory mediator-induced activation of mouse airway vagal C-fibres.

The influence of NaV 1.9 on inflammatory mediator-induced activation of airway vagal nodose C-fibres was evaluated by comparing responses in wild-type versus NaV 1.9-/- mice. A single-cell RT-PCR analysis indicated that virtually all nodose C-fibre neurons expressed NaV 1.9 (SCN11A) mRNA. Using extracellular electrophysiological recordings in an isolated vagally innervated mouse trachea-lung preparation, it was noted that mediators acting via G protein-coupled receptors (PAR2), or ionotropic receptors (P2×3) were 70-85% less effective in evoking action potential discharge in the absence of NaV 1.9. However, there was no difference in action potential discharge between wild-type and NaV 1.9-/- when the stimulus was a rapid punctate mechanical stimulus. An analysis of the passive and active properties of isolated nodose neurons revealed no difference between neurons from wild-type and NaV 1.9-/- mice, with the exception of a modest difference in the duration of the afterhyperpolarization. There was also no difference in the amount of current required to evoke action potentials (rheobase) or the action potential voltage threshold. The inward current evoked by the chemical mediator by a P2×3 agonist was the same in wild-type versus NaV 1.9-/- neurons. However, the current was sufficient to evoke action potential only in the wild-type neurons. The data support the speculation that NaV 1.9 could be an attractive therapeutic target for inflammatory airway disease by selectively inhibiting inflammatory mediator-associated vagal C-fibre activation. KEY POINTS: Inflammatory mediators were much less effective in activating the terminals of vagal airway C-fibres in mice lacking NaV 1.9. The active and passive properties of nodose neurons were the same between wild-type neurons and NaV 1.9-/- neurons. Nerves lacking NaV 1.9 responded, normally, with action potential discharge to rapid punctate mechanical stimulation of the terminals or the rapid stimulation of the cell bodies with inward current injections. NaV 1.9 channels could be an attractive target to selectively inhibit vagal nociceptive C-fibre activation evoked by inflammatory mediators without blocking the nerves' responses to the potentially hazardous stimuli associated with aspiration.

Dickkopf-1 inhibits the secretion of MUC5AC induced by Mycoplasma pneumoniae P1-C in mouse lung epithelial cells

Mycoplasma pneumoniae is the most common pathogen of respiratory tract infection in children and adults. Clinical observation shows that M. pneumoniae infection can cause massive mucus secretion in the respiratory tract, which makes the breathing of patients difficult. Studies have shown that M. pneumoniae infection can cause massive secretion of mucin 5AC (MUC5AC). Adhesin P1 plays an important role in the pathogenesis of M. pneumoniae infection by mediating the adhesion of pathogens to host cells, and the C-terminal residues of P1 (P1-C) are immunogenic. This study investigated the molecular mechanism of Wnt/β-catenin signaling pathway inhibitor Dickkopf-1 (DKK1) in the secretion of MUC5AC in mouse airway epithelial cells (MAECs) induced by P1-C. Scanning electron microscope and hematoxylin-eosin staining were used to observe the effect of P1-C on mucus secretion of MAECs. Protein chip was used to detect the secretion of cytokines and analyse the enrichment of related signaling pathways induced by P1-C in MAECs. Periodic acid schiff stain (PAS) staining, Tunel staining and Masson staining were used to detect the damage of the lungs of mouse exposed to P1-C. Immunohistochemistry was used to detect the secretion of MUC5AC expression, and Western blotting was used to reveal the molecular mechanism of DKK1-regulated secretion of MUC5AC induced by P1-C protein in MACES. The results showed that P1-C induced the massive secretion of mucus and inflammatory factors in MAECs. During P1-C infection, DKK1 down-regulated janus kinase 2 (JAK2), phosphorylation signaling and transcription activator 1 (p-STAT1) and phosphorylation signaling and activator of transcription 3 (p-STAT3) expression. Overexpression of DKK1 significantly up-regulated the expression of MUC5AC repressor transcription factor fork-head box protein A2 (FOXA2). At the same time, the expression of MUC5AC induced by P1-C was inhibited significantly. It is speculated that DKK1 can effectively reduce the secretion of MUC5AC in MAECs induced by P1-C by inhibiting the JAK/STAT1-STAT3 signaling pathway and up-regulating the expression of FOXA2.

Ambroxol-enhanced ciliary beating via voltage-gated Ca2+ channels in mouse airway ciliated cells

Ambroxol (ABX) facilitates the mucociliary clearance (MC) by enhancing ciliary beating in airways. In this study, we focused on airway ciliary beating enhanced by ABX. However, little is known about the ABX-stimulated Ca2+ signalling activating airway ciliary beating. Airway ciliated cells isolated from mice lungs were observed by a high-speed video microscope, and the activities of beating cilia were assessed by CBF (ciliary beat frequency) and CBD (ciliary bend distance, an index of amplitude). ABX (10 μM) enhanced the CBF and CBD by 30%, and the enhancement was inhibited by nifedipine (20 μM, a L-type voltage-gated Ca2+ channel (CaV) inhibitor), or a Ca2+-free solution (approximately 50%). Pre-treatment with BAPTA-AM (10 μM, a chelator of intracellular Ca2+) abolished ABX-stimulated increases in CBF, CBD and [Ca2+]i. Thus, ABX increases [Ca2+]i (intracellular Ca2+ concentration) by stimulating Ca2+ release from the internal stores and nifedipine-sensitive Ca2+ entry. A previous study demonstrated the expression of CaV1.2 in airway cilia. ABX enhanced CBF, CBD and [Ca2+]i even in a high extracellular K+ concentration (155.5 mM), suggesting that it activates CaV1.2 except by depolarization. These enhancements were inhibited by nifedipine. In conclusion, ABX, which increases [Ca2+]i by stimulating Ca2+ release from internal stores and Ca2+ entry through CaV1.2s, enhanced CBF and CBD in airway ciliated cells. ABX is a novel agonist that modulates CaV1.2 of airway beating cilia to enhance CBF and CBD.

Effect of Staphylococcus aureus in experimental pneumonia mouse model on promotion of mBD-3 expression through activation of the ERK1/2 pathway

Background/Aim. Staphylococcus aureus (S. aureus) is a gram-positive pathogen that causes various human diseases. S. aureus causes pneumonia, which is characterized by localized tissue necrosis. The aim of the study was to explore the expression of mouse ?-defensin 3 (mBD-3) induced by S. aureus in mouse lungs and the effect of mBD-3 expression on the mitogen-activated protein kinase (MAPK) pathway. Methods. An experimental model of S. aureus pneumonia in mice was developed, and the expression of mBD-3 and activation of the MAPK pathway were investigated using the methods of immunofluorescence and western blot. Results. The experimental model was created successfully. The number of white blood cells was elevated 48 and 72 hrs after the introduction of bacteria through mouse airways, and bronchiolar mucosal hyperemia was observed, along with a large number of white blood cells and mucus in the bronchioles. The mBD-3 expression levels 48 and 72 hrs after the induction of infection were greater than the levels in the control group and 24 hrs after the induction. The amount of phosphorylated extracellular signal-regulated kinase (ERK1/2) was increased 48 and 72 hrs after infection induction, compared with the levels in the control group and 24 hrs after induction. The expression of mBD-3 was lower when ERK1/2 phosphorylation was inhibited by the U0126 inhibitor. Conclusion. S. aureus in experimental pneumonia mouse model accelerates mBD-3 expression in the mouse lung mainly through an ERK1/2-dependent signaling pathway.

Scorpio and centipede ameliorate asthma by inhibiting the crosstalk between ferroptosis and inflammation in airway epithelial cells.

We previously revealed that scorpio and centipede (SC) improve the inflammatory response in asthma, whereas it is unclear whether ferroptosis is involved in this process. The asthmatic mouse model was established and lung tissues were collected for histopathological examination. The levels of tumor necrosis factor-α (TNF-α), interleukin- (IL-)1β, Fe2+, malondialdehyde (MDA), glutathione peroxidase 4 (GPX4), ferritin heavy chain 1(FTH1), and reactive oxygen species (ROS) were assessed in asthmatic mice and mouse airway epithelial cells. Our results showed that ferroptosis was induced in asthmatic mice, as evidenced by the reduction of FTH1 and GPX4 expression and the increase of MDA and Fe2+ levels (all P<0.05). Ferrostatin-1 repressed inflammation and ferroptosis of asthmatic mice. Additionally, SC significantly inhibited the levels of TNF-α, IL-1β, MDA, and Fe2+, while enhancing FTH1 and GPX4 expression. However, SC plus erastin showed the reverse results. Moreover, ferroptosis remarkably increased in asthmatic airway epithelial cells, while SC suppressed ferroptosis of the cells (all P<0.05). SC ameliorated asthma by inhibiting the crosstalk between ferroptosis and inflammation in airway epithelial cells.

Effect of icariin on the H2O2-induced proliferation of mouse airway smooth muscle cells through miR-138-5p regulating SIRT1/AMPK/PGC-1α axis.

Icariin exerts antioxidative and anti-inflammatory effects and is used in the treatment of bronchial asthma. However, the specific modes of action are uncertain. In this study, we investigated whether icariin could modulate the silencing information regulator 2-related enzyme 1 (SIRT1)/adenosine monophosphate-activated protein kinase (AMPK)/peroxisome proliferator-activated receptor gamma co-activator 1α (PGC-1α) axis by regulating miR-138-5p during H2O2-induced proliferation of mouse airway smooth muscle cells (ASMCs). Primary BALB/c mouse ASMCs were cultured using the tissue block adherence method and were induced with hydrogen peroxide (H2O2; 200μmol/L) to establish a bronchial asthma ASMC proliferation model. With the aid of Western Blot and quantitative real-time polymerase chain reaction (qRT-PCR) in H2O2-induced ASMCs, the expression of miR-138-5p, SIRT1, AMPK, PGC-1α, α-smooth muscle actin (α-SMA), transforming growth factor-β1 (TGF-β1), collagen I, and collagen III protein and mRNA were investigated. The proliferation rate and activities of superoxide dismutase1 (SOD1), reduced glutathione (GSH), malonaldehyde (MDA), and reactive oxygen species (ROS) in ASMCs were determined. The results suggest Compared with the H2O2-induced group, icariin inhibited the miR-138-5p expression; enhanced SIRT1, p-AMPK, and PGC-1α expression; attenuated MDA activity and ROS level; lowered TGF-β1, collagen I, and collagen III expression levels; and decreased the proliferation of ASMCs induced by H2O2. The dual-luciferase reporter gene assay results showed that SIRT1 is a regulatory target of miR-138-5p.The results suggest that Icariin could improve the H2O2-induced proliferation of ASMCs. The mechanism may be related to the increase of activation of SIRT1/AMPK/PGC-1α axis by suppressing the expression of miR-138-5p. Thus, SIRT1 is the regulatory target of miR-138-5p.

Proanthocyanidins attenuates ferroptosis against influenza-induced acute lung injury in mice by reducing IFN-γ

BackgroundAcute lung injury (ALI) is associated with high morbidity and mortality and is partly driven promoted by ferroptosis. Proanthocyanidins (PAs) is a natural bioactive flavonoid with anti-inflammatory and antioxidant activities. PAs can also significantly protect against acute lung inflammation and ferroptosis in alveolar epithelial cells. However, it is unclear whether PAs can alleviate ALI by reducing ferroptosis. This study aimed to evaluate the protective effects of PAs and the potential mechanisms against Influenza A virus (IAV)-induced ALI. MethodsMice were inoculated nasally with IAV to induce ALI. IAV-induced pulmonary inflammation and ferroptosis was tested by measuring the levels of malondialdehyde (MDA), glutathione (GSH), glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11 (SLC7A11) and acyl-CoA synthetase long-chain family member (ACSL4) in lung tissue. The potential targets that PAs protect against IAV-induced ALI were determined via a systemic pharmacological analysis. The molecular mechanism of PAs in ALI treatment was investigated by assessing the level of inflammation and ferroptosis markers using Western Blot and quantitative real-time PCR. ResultsSystemic pharmacological analysis suggested that PAs protect against IAV-induced pneumonia thorough TGF-β1 and its relative signaling pathway. PAs effectively alleviated histopathological lung injury, reduced inflammatory cytokines and chemokines secretion, which were increased in IAV-infected mice. Meanwhile, PAs further prevented mouse airway inflammation in ALI, concomitant with the decreased expression TGF-β1, smad2/3, p-Smad2, p-Smad3 and ferroptosis mediator IFN-γ. Furthermore，IFN-γ promotes cell lipid peroxidation and ferroptosis，PAs significantly reduced MDA and ACSL4 levels and upregulated GSH, GPX4, and SLC7A11. ConclusionOverall, PAs can attenuate ferroptosis against IAV-induced ALI via the TGF-β1/Smad2/3 pathway and is a promising novel therapeutic candidate for ALI.

Chronic allergic asthma alters m6A epitranscriptomic tagging of mRNAs and lncRNAs in the lung.

To evaluate the role of m6A methylation of mRNAs and long non-coding RNAs (lncRNAs) in chronic allergic asthma. Transcriptome-wide N6-methyladenosine (m6A) changes in BALB/c mice were profiled using immunoprecipitated methylated RNAs with microarrays in lung with chronic allergic asthma. Gene ontology (GO) and KEGG analyses were conducted. Target genes were verified by methylated RNA immunoprecipitation and real-time polymerase chain reaction (PCR). Specifically, the mRNA levels of m6A writers (METTL3, METTL14, and WTAP), and readers and erasers (FTO and ALKBH5) were estimated by real-time PCR analysis, using the SYBR-green method. IL17RB mRNA was also evaluated by PCR. Hematoxylin and eosin (H&E) staining showed that the airway and lung tissues in mice in the asthma group had extensive infiltration of inflammatory cells around the bronchioles, blood vessels, and alveoli. The lungs of those allergic asthma mice showed altered m6A epitranscriptome, whereby 1369 mRNAs and 176 lncRNAs were hypermethylated, and 197 mRNAs and 30 lncRNAs were hypomethylated (>1.5-fold vs control). Also, compared with the control group, IL17RB mRNA in lung of the asthmatic group was significantly hypermethylated (P<0.01). In the asthma group, the mRNA and the protein level of METTL14 (the key methyltransferase) and ALKBH5 (the major demethyltransferase) were significantly decreased compared with the control group (P<0.01). Chronic allergic asthma alters the lung m6A epitranscriptome, suggesting functional implications in the pathophysiology of refractory asthma. Data support methylated IL17RB mRNA possibly becoming a new therapeutic target for chronic allergic asthma.

HIF1-α upregulation induces proinflammatory factors to boost host killing capacity after Aspergillus fumigatus exposure.

Aims: HIF1-α is an important transcription factor in the regulation of the immune response. The protective function of HIF1-α in the host epithelial immune response to Aspergillus fumigatus requires further clarification. Methods: In this study we demonstrated the effect of upregulation of HIF1-α expression in A549 cells and mouse airway cells exposed to A. fumigatus in vivo. Results: The killing capacity was enhanced by boosting proinflammatory factors both in vitro and in vivo. Moreover, airway inflammation was reduced in the HIF1-α-upregulated mice. Conclusion: We identified a protective role for HIF1-α in anti-A. fumigatus immunity. Modulation of HIF1-α might be a target for the development of aspergillosis therapy.

Targeting epithelial cell-derived TWIST1 alleviates allergic asthma

It is well known that the T Helper (Th)2 bias plays a critical role in allergic asthma. Whereas the Th2 bias is maintained in the local tissues is uncertain. IL-33 is vital for the development of the Th2 polarization. TWIST-1 has an effect on regulating cellular functions. The aberrant activation of RAS sustains certain cellular activities. The aim of this study is to study the role of the interaction between activation of TWIST1 and RAS in inducing and maintaining Th2 polarization in allergic asthma. The epithelial cells of the airways (AEC) were isolated from the broncho-alveolar lavage fluids in patients with asthma. The mediators involved in the over-expression of IL-33 were determined by RNA sequencing. A mouse model was established to test the role of TWIST1 and RAS in developing allergic asthma. We observed a strong expression of TWIST1 in patients with allergic asthma that showed a positive correlation with asthmatic responses. TWIST1 favored the expression of the IL-33 in the AEC. Twist1-deficient AEC-carrying mice did not induce Th2 polarization in the airways. The expression TWIST1 in AECs was positively associated with RAS activation in AECs in patients with allergic asthma. The interaction between RAS and TWIST1 in AECs sustained airway allergic inflammation. Inhibition of TWIST1 or RAS prevented asthma-like inflammation in the mouse airways. In summary, the interaction between TWIST1 and RAS induces and maintains IL-33 expression in AECs to facilitate allergic inflammation in the respiratory tract. Inhibition of TWIST1 or RAS can prevent experimental allergic asthma.

Chronic exposure to IQOS results in impaired pulmonary function and lung tissue damage in mice

The use of IQOS brand heated tobacco products (HTPs) is increasing worldwide; however, little is known about the long-term effects of HTPs aerosol exposure on the lungs. Herein, we exposed C57BL/6 J mice for 24 weeks to clean air, IQOS aerosol, or cigarette smoke, and determined pulmonary function, lung tissue pathology, inflammation, and oxidative stress. Compared with the control group mice, IQOS group mice showed substantially decreased weight and lung function. Levels of IL-6 and TNF-a, as well as oxidative stress markers, were comparable to those found in the cigarette group. In addition, hematoxylin and eosin staining showed that the alveolar space was enlarged and that emphysema had formed in the IQOS group. Masson staining showed that collagen deposition areas were substantially increased in the airway walls in the IQOS group than in the control group. Immunohistochemical staining showed epithelial-mesenchymal transition in the airways of mice in the IQOS group. In conclusion, chronic exposure to IQOS aerosol results in impaired pulmonary function and lung tissue damage; hence, concern should be raised regarding the long-term safety of this product.

Reserve of Wnt/β-catenin Signaling Alleviates Mycoplasma pneumoniae P1-C-induced Inflammation in airway epithelial cells and lungs of mice

Mycoplasma pneumoniae (M. pneumoniae) is the most common pathogen of respiratory tract infections in both children and adults. M. pneumoniae P1 adhesin plays an important role in the pathogenesis of M. pneumoniae infection by mediating the attachment of pathogen to host cells. The inoculation of C-terminal residuals of P1 (P1-C) showed a protective role from M. pneumoniae infection. Accumulated evidence suggests that the Wnt/β-Catenin signaling is implicated in regulation of inflammatory responses to bacterial infections. However, mechanisms underlying the regulatory roles of Wnt signaling in host cells in response to M. pneumoniae infections are incompletely understood. In the present study, the impact and molecular mechanism of Wnt/β-catenin signaling in immune responses induced by M. pneumoniae P1-C were investigated. The results demonstrated that the P1-C could activate Wnt/β-catenin and Toll-like receptor (TLR) signaling in primary mouse airway epithelial cells cultured in an air-liquid interface (ALI) state. Interestingly, the inhibition of Wnt/β-catenin signaling by an adenovirus-mediated Wnt inhibitor Dickkopf-1 (Dkk1) gene transduction alleviated the P1-C induced inflammation fibrosis in mouse lung, accompanied by the reduced expression of epithelial mesenchymal transition (EMT) markers. Mechanistical analysis further demonstrated that the Dkk1 could suppress the expression of JAK2/STAT1-STAT3 and Caspase3, 8/Bax signaling in mouse lung tissues. In vitro study further revealed that XAV939, a small molecule of Wnt/β-catenin inhibitor, inhibited the P1-C-activated TLR4/MyD88 signaling and cytokine productions in primary mouse airway ALI epithelial cells. This study thus provides an insight into the function of Wnt/β-catenin signaling in regulation of the pathogenesis of M. pneumoniae infection, suggesting that targeting Wnt/β-catenin signaling by gene transduction of Dkk1, or pharmacological molecules of inhibitor may be a promised approach that worthy of further investigation in the treatment of M. pneumoniae pneumonia.