A common polymorphism in the Intelectin-1 gene influences mucus plugging in severe asthma.

By incompletely understood mechanisms, type 2 (T2) inflammation present in the airways of severe asthmatics drives the formation of pathologic mucus which leads to airway mucus plugging. Here we investigate the molecular role and clinical significance of intelectin-1 (ITLN-1) in the development of pathologic airway mucus in asthma. Through analyses of human airway epithelial cells we find that ITLN1 gene expression is highly induced by interleukin-13 (IL-13) in a subset of metaplastic MUC5AC+ mucus secretory cells, and that ITLN-1 protein is a secreted component of IL-13-induced mucus. Additionally, we find ITLN-1 protein binds the C-terminus of the MUC5AC mucin and that its deletion in airway epithelial cells partially reverses IL-13-induced mucostasis. Through analysis of nasal airway epithelial brushings, we find that ITLN1 is highly expressed in T2-high asthmatics, when compared to T2-low children. Furthermore, we demonstrate that both ITLN-1 gene expression and protein levels are significantly reduced by a common genetic variant that is associated with protection from the formation of mucus plugs in T2-high asthma. This work identifies an important biomarker and targetable pathways for the treatment of mucus obstruction in asthma.

In situ pulmonary mucus hydration assay using rotational and translational diffusion of gold nanorods with polarization-sensitive optical coherence tomography.

Significance: Assessing the nanostructure of polymer solutions and biofluids is broadly useful for understanding drug delivery and disease progression and for monitoring therapy. Aim: Our objective is to quantify bronchial mucus solids concentration (wt. %) during hypertonic saline (HTS) treatment in vitro via nanostructurally constrained diffusion of gold nanorods (GNRs) monitored by polarization-sensitive optical coherence tomography (PS-OCT). Approach: Using PS-OCT, we quantified GNR translational (DT) and rotational (DR) diffusion coefficients within polyethylene oxide solutions (0 to 3 wt. %) and human bronchial epithelial cell (hBEC) mucus (0 to 6.4 wt. %). Interpolation of DT and DR data is used to develop an assay to quantify mucus concentration. The assay is demonstrated on the mucus layer of an air-liquid interface hBEC culture during HTS treatment. Results: In polymer solutions and mucus, DT and DR monotonically decrease with increasing concentration. DR is more sensitive than DT to changes above 1.5 wt. % of mucus and exhibits less intrasample variability. Mucus on HTS-treated hBEC cultures exhibits dynamic mixing from cilia. A region of hard-packed mucus is revealed by DR measurements. Conclusions: The extended dynamic range afforded by simultaneous measurement of DT and DR of GNRs using PS-OCT enables resolving concentration of the bronchial mucus layer over a range from healthy to disease in depth and time during HTS treatment in vitro.

Improvement of skin wound healing by giant salamander skin mucus gel wrapped with bone marrow mesenchymal stem cells via affecting integrin family molecules.

BACKGROUND: Traditional bandages, gauze, and cotton balls are increasingly insufficient for addressing complex war injuries characterized by severe bleeding and diverse wound conditions. The giant salamander, a species of high medical value, secretes a unique mucus when stimulated, which has potential applications in wound care. MATERIALS: Giant salamander skin mucus gel dressing wrapped with bone marrow mesenchymal stem cells (BMSCs-GSSM-gel) was prepared and validated. Skin wound injury of rabbit and mouse models were established. Hematoxylin and Eosin, Masson's trichrome, and Sirius red staining were performed. The platelet aggregation rate and coagulation items were measured. Transcriptome sequencing was performed to find potential differential expression genes. RESULTS: Preparation and characterization of BMSCs-GSSM-gel were performed, and BMSCs-GSSM-gel particles with a diameter of about 200 nm were obtained. BMSCs-GSSM-gel accelerated wound healing in both rabbit and mouse models. BMSCs-GSSM-gel significantly promoted hemostasis via increasing platelet aggregation rate and fibrinogen, but decreasing activated partial thromboplastin time, thrombin time, and prothrombin time. BMSCs-GSSM-gel treatment significantly impacted several genes associated with cell adhesion, inflammatory response, collagen-containing extracellular matrix, and the positive regulation of cell migration based on Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Integrin Subunit Beta 4 (ITGB4), Integrin Subunit Alpha 3 (ITGA3), and Laminin Subunit Beta 3 (LAMB3) might be involved in the wound healing process by BMSCs-GSSM-gel. CONCLUSIONS: We proved the BMSCs-GSSM-gel greatly improved the skin wound healing, and it might play a crucial role in the application fields of skin damage repair.

Hindering the biofilm of microbial pathogens and cancer cell lines development using silver nanoparticles synthesized by epidermal mucus proteins from Clarias gariepinus.

Scientists know very little about the mechanisms underlying fish skin mucus, despite the fact that it is a component of the immune system. Fish skin mucus is an important component of defence against invasive infections. Recently, Fish skin and its mucus are gaining interest among immunologists. Characterization was done on the obtained silver nanoparticles Ag combined with Clarias gariepinus catfish epidermal mucus proteins (EMP-Ag-NPs) through UV-vis, FTIR, XRD, TEM, and SEM. Ag-NPs ranged in size from 4 to 20 nm, spherical in form and the angles were 38.10°, 44.20°, 64.40°, and 77.20°, Where wavelength change after formation of EMP-Ag-NPs as indicate of dark brown, the broad band recorded at wavelength at 391 nm. Additionally, the antimicrobial, antibiofilm and anticancer activities of EMP-Ag-NPs was assessed. The present results demonstrate high activity against unicellular fungi C. albicans, followed by E. faecalis. Antibiofilm results showed strong activity against both S. aureus and P. aeruginosa pathogens in a dose-dependent manner, without affecting planktonic cell growth. Also, cytotoxicity effect was investigated against normal cells (Vero), breast cancer cells (Mcf7) and hepatic carcinoma (HepG2) cell lines at concentrations (200-6.25 µg/mL) and current results showed highly anticancer effect of Ag-NPs at concentrations 100, 5 and 25 µg/mL exhibited rounding, shrinkage, deformation and granulation of Mcf7 and HepG2 with IC50 19.34 and 31.16 µg/mL respectively while Vero cells appeared rounded at concentration 50 µg/mL and normal shape at concentration 25, 12.5 and 6.25 µg/ml with IC50 35.85 µg/mL. This study evidence the potential efficacy of biologically generated Ag-NPs as a substitute medicinal agent against harmful microorganisms. Furthermore, it highlights their inhibitory effect on cancer cell lines.

Seihaito, a Kampo medicine, attenuates IL-13-induced mucus production and goblet cell metaplasia.

Goblet cell hyperplasia and increased mucus production are features of airway diseases, including asthma, and excess airway mucus often worsens these conditions. Even steroids are not uniformly effective in mucus production in severe asthma, and new therapeutic options are needed. Seihaito is a Japanese traditional medicine that is used clinically as an antitussive and expectorant. In the present study, we examined the effect of Seihaito on goblet cell differentiation and mucus production. In in vitro studies, using air-liquid interface culture of guinea-pig tracheal epithelial cells, Seihaito inhibited IL-13-induced proliferation of goblet cells and MUC5AC, a major component of mucus production. Seihaito suppressed goblet cell-specific gene expression, without changing ciliary cell-specific genes, suggesting that it inhibits goblet cell differentiation. In addition, Seihaito suppressed MUC5AC expression in cells transfected with SPDEF, a transcription factor activated by IL-13. Furthermore, Seihaito attenuated in vivo goblet cell proliferation and MUC5AC mRNA expression in IL-13-treated mouse lungs. Collectively, these findings demonstrated that Seihaito has an inhibitory effect on goblet cell differentiation and mucus production, which is at least partly due to the inhibition of SPDEF.

Coarse-grained modeling and dynamics tracking of nanoparticles diffusion in human gut mucus.

The gastrointestinal (GI) tract's mucus layer serves as a critical barrier and a mediator in drug nanoparticle delivery. The mucus layer's diverse molecular structures and spatial complexity complicates the mechanistic study of the diffusion dynamics of particulate materials. In response, we developed a bi-component coarse-grained mucus model, specifically tailored for the colorectal cancer environment, that contained the two most abundant glycoproteins in GI mucus: Muc2 and Muc5AC. This model demonstrated the effects of molecular composition and concentration on mucus pore size, a key determinant in the permeability of nanoparticles. Using this computational model, we investigated the diffusion rate of polyethylene glycol (PEG) coated nanoparticles, a widely used muco-penetrating nanoparticle. We validated our model with experimentally characterized mucus pore sizes and the diffusional coefficients of PEG-coated nanoparticles in the mucus collected from cultured human colorectal goblet cells. Machine learning fingerprints were then employed to provide a mechanistic understanding of nanoparticle diffusional behavior. We found that larger nanoparticles tended to be trapped in mucus over longer durations but exhibited more ballistic diffusion over shorter time spans. Through these discoveries, our model provides a promising platform to study pharmacokinetics in the GI mucus layer.

AFANet: Adaptive feature aggregation for polyp segmentation.

In terms of speed and accuracy, the deep learning-based polyp segmentation method is superior. It is essential for the early detection and treatment of colorectal cancer and has the potential to greatly reduce the disease's overall prevalence. Due to the various forms and sizes of polyps, as well as the blurring of the boundaries between the polyp region and the surrounding mucus, most existing algorithms are unable to provide highly accurate colorectal polyp segmentation. Therefore, to overcome these obstacles, we propose an adaptive feature aggregation network (AFANet). It contains two main modules: the Multi-modal Balancing Attention Module (MMBA) and the Global Context Module (GCM). The MMBA extracts improved local characteristics for inference by integrating local contextual information while paying attention to them in three regions: foreground, background, and border. The GCM takes global information from the top of the encoder and sends it to the decoder layer in order to further investigate global contextual feature information in the pathologic picture. Dice of 92.11 % and 94.76 % and MIoU of 91.07 % and 94.54 %, respectively, are achieved by comprehensive experimental validation of our proposed technique on two benchmark datasets, Kvasir-SEG and CVCClinicDB. The experimental results demonstrate that the strategy outperforms other cutting-edge approaches.

Wogonoside Ameliorates Airway Inflammation and Mucus Hypersecretion via NF-κB/STAT6 Signaling in Ovalbumin-Induced Murine Acute Asthma.

Asthma is recognized as a chronic respiratory illness characterized by airway inflammation and airway hyperresponsiveness. Wogonoside, a flavonoid glycoside, is reported to significantly alleviate the inflammation response and oxidative stress. Herein, this study aimed to investigate the therapeutic effect and underlying mechanism of wogonoside on airway inflammation and mucus hypersecretion in a murine asthma model and in human bronchial epithelial cells (16HBE). BALB/c mice were sensitized and challenged with ovalbumin (OVA). Pulmonary function and the number of cells in the bronchoalveolar lavage fluid (BALF) were examined. Pathological changes in lung tissue in each group were evaluated via hematoxylin and eosin and periodic acid-Schiff staining, and changes in levels of cytokines in BALF and of immunoglobulin E in serum were determined via an enzyme-linked immunosorbent assay. The expression of relevant genes in lung tissue was analyzed via real-time PCR. Western blotting and immunofluorescence were employed to detect the expression of relevant proteins in lung tissue and 16HBE cells. Treatment with 10 and 20 mg/kg wogonoside significantly attenuated the OVA-induced increase of inflammatory cell infiltration, mucus secretion, and goblet cell percentage and improved pulmonary function. Wogonoside treatment reduced the level of T-helper 2 cytokines including interleukin (IL)-4, IL-5, and IL-13 in BALF and of IgE in serum and decreased the mRNA levels of cytokines (IL-4, IL-5, IL-6, IL-13, and IL-1ß and tumor necrosis factor-α), chemokines (CCL-2, CCL-11, and CCL-24), and mucoproteins (MUC5AC, MUC5B, and GOB5) in lung tissues. The expression of MUC5AC and the phosphorylation of STAT6 and NF-κB p65 in lung tissues and 16HBE cells were significantly downregulated after wogonoside treatment. Thus, wogonoside treatment may effectively decrease airway inflammation, airway remodeling, and mucus hypersecretion via blocking NF-κB/STAT6 activation.

Validation of Salamander Dermal Mucus Swabs as a Novel, Nonlethal Approach for Amphibian Metabolomics and Glutathione Analysis Following Pesticide Exposure.

Evaluating biomarkers of stress in amphibians is critical to conservation, yet current techniques are often destructive and/or time-consuming, which limits ease of use. In the present study, we validate the use of dermal swabs in spotted salamanders (Ambystoma maculatum) for biochemical profiling, as well as glutathione (GSH) stress response following pesticide exposure. Thirty-three purchased spotted salamanders were acclimated to laboratory conditions at Washington College (Chestertown, MD, USA) for 4 weeks. Following acclimation, salamanders were randomly sorted into three groups for an 8-h pesticide exposure on soil: control with no pesticide, 2,4-dichlorophenoxyacetic acid (2,4-D), or chlorpyrifos. Before and after exposure, mucus samples were obtained by gently rubbing a polyester-tipped swab 50 times across the ventral and dorsal surfaces. Salamanders were humanely euthanized and dissected to remove the brain for acetylcholinesterase and liver for GSH and hepatic metabolome analyses, and a whole-body tissue homogenate was used for pesticide quantification. Levels of GSH were present in lower quantities on dermal swabs relative to liver tissues for chlorpyrifos, 2,4-D, and control treatments. However, 2,4-D exposures demonstrated a large effect size increase for GSH levels in livers (Cohen's d = 0.925, p = 0.036). Other GSH increases were statistically insignificant, and effect sizes were characterized as small for 2,4-D mucosal swabs (d = 0.36), medium for chlorpyrifos mucosal swabs (d = 0.713), and negligible for chlorpyrifos liver levels (d = 0.012). The metabolomics analyses indicated that the urea cycle, alanine, and glutamate metabolism biological pathways were perturbed by both sets of pesticide exposures. Obtaining mucus samples through dermal swabbing in amphibians is a viable technique for evaluating health in these imperiled taxa. Environ Toxicol Chem 2024;43:1126-1137. © 2024 SETAC.

Mucus Structure, Viscoelastic Properties, and Composition in Chronic Respiratory Diseases.

The respiratory mucus, a viscoelastic gel, effectuates a primary line of the airway defense when operated by the mucociliary clearance. In chronic respiratory diseases (CRDs), such as asthma, chronic obstructive pulmonary disease (COPD), and cystic fibrosis (CF), the mucus is overproduced and its solid content augments, changing its structure and viscoelastic properties and determining a derangement of essential defense mechanisms against opportunistic microbial (virus and bacteria) pathogens. This ensues in damaging of the airways, leading to a vicious cycle of obstruction and infection responsible for the harsh clinical evolution of these CRDs. Here, we review the essential features of normal and pathological mucus (i.e., sputum in CF, COPD, and asthma), i.e., mucin content, structure (mesh size), micro/macro-rheology, pH, and osmotic pressure, ending with the awareness that sputum biomarkers (mucins, inflammatory proteins and peptides, and metabolites) might serve to indicate acute exacerbation and response to therapies. There are some indications that old and novel treatments may change the structure, viscoelastic properties, and biomarker content of sputum; however, a wealth of work is still needed to embrace these measures as correlates of disease severity in association with (or even as substitutes of) pulmonary functional tests.

Water Sorption and Structural Properties of Human Airway Mucus in Health and Muco-Obstructive Diseases.

Muco-obstructive diseases change airway mucus properties, impairing mucociliary transport and increasing the likelihood of infections. To investigate the sorption properties and nanostructures of mucus in health and disease, we investigated mucus samples from patients and cell cultures (cc) from healthy, chronic obstructive pulmonary disease (COPD), and cystic fibrosis (CF) airways. Atomic force microscopy (AFM) revealed mucin monomers with typical barbell structures, where the globule to spacer volume ratio was the highest for CF mucin. Accordingly, synchrotron small-angle X-ray scattering (SAXS) revealed more pronounced scattering from CF mucin globules and suggested shorter carbohydrate side chains in CF mucin and longer side chains in COPD mucin. Quartz crystal microbalance with dissipation (QCM-D) analysis presented water sorption isotherms of the three types of human airway mucus, where, at high relative humidity, COPD mucus had the highest water content compared to cc-CF and healthy airway mucus (HAM). The higher hydration of the COPD mucus is consistent with the observation of longer side chains of the COPD mucins. At low humidity, no dehydration-induced glass transition was observed in healthy and diseased mucus, suggesting mucus remained in a rubbery state. However, in dialyzed cc-HAM, a sorption-desorption hysteresis (typically observed in the glassy state) appeared, suggesting that small molecules present in mucus suppress the glass transition.

Proteomic analysis of the mucus of the photosynthetic sea slug Elysia crispata.

Elysia crispata is a tropical sea slug that can retain intracellular functional chloroplasts from its algae prey, a mechanism termed kleptoplasty. This sea slug, like other gastropods, secretes mucus, a viscous secretion with multiple functions, including lubrication, protection, and locomotion. This study presents the first comprehensive analysis of the mucus proteome of the sea slug E. crispata using gel electrophoresis and HPLC-MS/MS. We identified 306 proteins in the mucus secretions of this animal, despite the limited entries for E. crispata in the Uniprot database. The functional annotation of the mucus proteome using Gene Ontology identified proteins involved in different functions such as hydrolase activity (molecular function), carbohydrate-derived metabolic processes (biological processes) and cytoskeletal organization (cell component). Moreover, a high proportion of proteins with enzymatic activity in the mucus of E. crispata suggests potential biotechnological applications including antimicrobial and antitumor activities. Putative antimicrobial properties are reinforced by the high abundance of hydrolases. This study also identified proteins common in mucus samples from various species, supporting a common mechanism of mucus in protecting cells and tissues while facilitating animal movement. SIGNIFICANCE: Marine species are increasingly drawing the interest of researchers for their role in discovering new bioactive compounds. The study "Proteomic Analysis of the Mucus of the Photosynthetic Sea Slug Elysia crispata" is a pioneering effort that uncovers the complex protein content in this fascinating sea slug's mucus. This detailed proteomic study has revealed proteins with potential use in biotechnology, particularly for antimicrobial and antitumor purposes. This research is a first step in exploring the possibilities within the mucus of Elysia crispata, suggesting the potential for new drug discoveries. These findings could be crucial in developing treatments for severe diseases, especially those caused by multidrug-resistant bacteria, and may lead to significant advances in medical research.

Inhibition of mucus secretion by niclosamide and benzbromarone in airways and intestine.

The Ca2+ activated Cl- channel TMEM16A (anoctamin 1; ANO1) is expressed in secretory epithelial cells of airways and intestine. Previous studies provided evidence for a role of ANO1 in mucus secretion. In the present study we investigated the effects of the two ANO1-inhibitors niclosamide (Niclo) and benzbromarone (Benz) in vitro and in vivo in mouse models for cystic fibrosis (CF) and asthma. In human CF airway epithelial cells (CFBE), Ca2+ increase and activation of ANO1 by adenosine triphosphate (ATP) or ionomycin was strongly inhibited by 200 nM Niclo and 1 µM Benz. In asthmatic mice airway mucus secretion was inhibited by intratracheal instillation of Niclo or Benz. In homozygous F508del-cftr mice, intestinal mucus secretion and infiltration by CD45-positive cells was inhibited by intraperitoneal injection of Niclo (13 mg/kg/day for 7 days). In homozygous F508del-cftr rats intestinal mucus secretion was inhibited by oral application of Benz (5 mg/kg/day for 60 days). Taken together, well tolerated therapeutic concentrations of niclosamide and benzbromarone corresponding to plasma levels of treated patients, inhibit ANO1 and intracellular Ca2+ signals and may therefore be useful in inhibiting mucus hypersecretion and mucus obstruction in airways and intestine of patients suffering from asthma and CF, respectively.

Mucus plugs in the airways of asthmatic subjects and smoking status.

BACKGROUND: Mucus plugs have been described in the airways of asthmatic subjects, particularly those with associated with type 2 inflammation and sputum eosinophilia. In the current study we addressed the question of whether smoking, neutrophilic inflammation and airway dimensions affected the prevalence of mucus plugs. METHODS: In a cohort of moderate to severe asthmatics (n = 50), including a group of ex-smokers and current smokers, the prevalence of mucus plugs was quantified using a semi-quantitative score based on thoracic computerized tomography. The relationships between mucus score, sputum inflammatory profile and airway architecture were tested according to patient's smoking status. RESULTS: Among the asthmatics (37% former or active smokers), 74% had at least one mucus plug. The median score was 3 and was unrelated to smoking status. A significant but weak correlation was found between mucus score, FEV1 and FEV1/FVC. Mucus score was significantly correlated with sputum eosinophils. Among former and active smokers, mucus score was correlated with sputum neutrophils. Mucus score was positively associated with FeNO in non-smoking subjects. The lumen dimensions of the main and lobar bronchi were significantly inversely correlated with mucus score. CONCLUSION: Airway mucus plugs could define an asthma phenotype with altered airway architecture and can occur in asthmatic subjects with either neutrophilic or eosinophilic sputum according to their smoking status.

The efficacy of N-acetylcysteine in decreasing airway inflammation and mucus accumulation in horses with 18 hours of head confinement.

BACKGROUND: During transportation many horses develop post-transportation infection, which can be life-threatening and end their sport career. Preventing mucus accumulation and inflammation during transportation is vital, emphasizing the need for effective strategies to enhance overall horse health welfare. OBJECTIVES: Assess the impact of N-acetylcysteine (NAC) on mucus accumulation and inflammation in horses subjected to 18 hours of head confinement. ANIMALS: Six healthy crossbred horses, 5.3 ± 2.1 years of age and weighing 387 ± 30 kg. METHODS: Prospective placebo-controlled cross-over design study. The horses' heads were restrained in their stalls for a period of 18 hours. They were studied under 4 conditions: Not confined (NC): before head confinement, placebo (P), and confined head (CH): 18 hours of head confinement without treatment, and N-Acetylcysteine (NAC): 18 hours of head confinement treated with NAC before confinement (15 mg/kg/day NAC PO for 3 days). Bronchoalveolar lavage (BAL) was performed in each condition. Mucus accumulation along the trachea was evaluated by endoscopy. RESULTS: Endoscopic scores were significantly different between CH and other conditions, whereas no significant differences were found among NC, P, and NAC. The BAL cell count (34 291 ± 2624 cells/µL), neutrophil and lymphocyte count (18 601 ± 3193 cells/µL and 3337.4 ± 593 cells/µL, respectively) in CH were significantly higher compared to NAC. Neutrophil percentage was significantly higher in CH (53.8 ± 8%) compared to horses that received NAC (20.08 ± 8%). Conversely, in comparison to NAC (66.33 ± 9%), the percentage of macrophages was significantly lower in CH (35.7 ± 10%). CONCLUSIONS: N-acetylcysteine was found to significantly decrease mucus accumulation and inflammatory cell counts in horses with head confinement.

Tadpole-Like Anisotropic Polymer/Lipid Janus Nanoparticles for Nose-to-Brain Drug Delivery: Importance of Geometry, Elasticity on Mucus-Penetration Ability.

Asymmetric geometry (aspect ratio >1), moderate stiffness (i.e., semielasticity), large surface area, and low mucoadhesion of nanoparticles are the main features to reach the brain by penetrating across the nasal mucosa. Herein, a new application has been presented for the use of multifunctional Janus nanoparticles (JNPs) with controllable geometry and size as a nose-to-brain (N2B) delivery system by changing proportions of Precirol ATO 5 and polycaprolactone compartments and other operating conditions. To bring to light the N2B application of JNPs, the results are presented in comparison with polymer and solid lipid nanoparticles, which are frequently used in the literature regarding their biopharmaceutical aspects: mucoadhesion and permeability through the nasal mucosa. The morphology and geometry of JPs were observed via cryogenic-temperature transmission electron microscopy images, and their particle sizes were verified by dynamic light scattering, atomic force microscopy, and scanning electron microscopy. Although all NPs showed penetration across the mucus barrier, the best increase in penetration was observed with asymmetric and semielastic JNPs, which have low interaction ability with the mucus layer. This study presents a new and promising field of application for a multifunctional system suitable for N2B delivery, potentially benefiting the treatment of brain tumors and other central nervous system diseases.

The combination of propylene glycol and vegetable glycerin e-cigarette aerosols induces airway inflammation and mucus hyperconcentration.

Despite concerns over their safety, e-cigarettes (e-cigs) remain a popular tobacco product. Although nicotine and flavors found in e-cig liquids (e-liquids) can cause harm in the airways, whether the delivery vehicles propylene glycol (PG) and vegetable glycerin (VG) are innocuous when inhaled remains unclear. Here, we investigated the effects of e-cig aerosols generated from e-liquid containing only PG/VG on airway inflammation and mucociliary function in primary human bronchial epithelial cells (HBEC) and sheep. Primary HBEC were cultured at the air-liquid interface (ALI) and exposed to e-cig aerosols of 50%/50% v/v PG/VG. Ion channel conductance, ciliary beat frequency, and the expression of inflammatory markers, cell type-specific markers, and the major mucins MUC5AC and MUC5B were evaluated after seven days of exposure. Sheep were exposed to e-cig aerosols of PG/VG for five days and mucus concentration and matrix metalloproteinase-9 (MMP-9) activity were measured from airway secretions. Seven-day exposure of HBEC to e-cig aerosols of PG/VG caused a significant reduction in the activities of apical ion channels important for mucus hydration, including the cystic fibrosis transmembrane conductance regulator (CFTR) and large conductance, Ca2+-activated, and voltage-dependent K+ (BK) channels. PG/VG aerosols significantly increased the mRNA expression of the inflammatory markers interleukin-6 (IL6), IL8, and MMP9, as well as MUC5AC. The increase in MUC5AC mRNA expression correlated with increased immunostaining of MUC5AC protein in PG/VG-exposed HBEC. On the other hand, PG/VG aerosols reduced MUC5B expression leading overall to higher MUC5AC/MUC5B ratios in exposed HBEC. Other cell type-specific markers, including forkhead box protein J1 (FOXJ1), keratin 5 (KRT5), and secretoglobin family 1A member 1 (SCGB1A1) mRNAs, as well as overall ciliation, were significantly reduced by PG/VG exposure. Finally, PG/VG aerosols increased MMP-9 activity and caused mucus hyperconcentration in sheep in vivo. E-cig aerosols of PG/VG induce airway inflammation, increase MUC5AC expression, and cause dysfunction of ion channels important for mucus hydration in HBEC in vitro. Furthermore, PG/VG aerosols increase MMP-9 activity and mucus concentration in sheep in vivo. Collectively, these data show that e-cig aerosols containing PG/VG are likely to be harmful in the airways.

Uncovering the antimicrobial activity of G-type lysozyme 2 derived from Cyprinus carpio mucus against bacterial and fungal pathogens.

This study aimed to explore the antimicrobic potential of mucus samples collected from Cyprinus carpio and identify the specific antimicrobial peptides responsible for its activity. The crude extract was tested against various bacterial and fungal pathogens, and its protein content and profile were analyzed. Purification steps, including gel filtration chromatography, were employed to isolate the most active fraction (peak IV), which was further identified via liquid chromatography and mass spectroscopy. The results revealed varying degrees of antimicrobial activity of the crude extract against different bacterial and fungal strains, with Leclercia adecarboxylata, Candida glabrata, and Candida parapsilosis showing the highest susceptibility. SDS-PAGE analysis demonstrated the existence of multiple low molecular weight protein bands in the crude extract, while fraction IV obtained from gel filtration chromatography exhibited the strongest antimicrobial activity. Peak IV displayed a range of minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and minimum fungicidal concentration (MFC) values against the tested pathogens, spanning from 0.038 to 4.960 mg/mL. Further investigation identified the purified peptide derived from peak IV as G-type lysozyme 2, characterized by a molecular weight of 21 kDa. These findings shed light on the existence of a highly effective antimicrobial peptide, G-type lysozyme 2, within the mucus of Cyprinus carpio. This peptide demonstrates notable activity against diverse bacterial and fungal pathogens. The insights from this study enhance our understanding of the fish's antimicrobial defense mechanisms and hold promise for developing novel antimicrobial agents.