Search Results (291)

Background: The study evaluates the immunomodulatory potential of secukinumab (SECU) and honokiol (HONK) in a murine model of allergic asthma complicated by acute lung injury (ALI), with an emphasis on modulating key inflammatory pathways. The rationale is driven by the necessity to attenuate Th17-mediated cytokine cascades, wherein IL-17 plays a critical role, as well as to explore the adjunctive anti-inflammatory effects of HONK on Th1 cytokine production, including IL-6, TNF-α, and Th2 cytokines. Methods: Mice were sensitized and challenged with ovalbumin (OVA) and lipopolysaccharide (LPS) was administrated to exacerbate pulmonary pathology, followed by administration of SECU, HONK (98% purity, C₁₈H₁₈O₂), or their combination. Quantitative analyses incorporated OVA-specific IgE measurements, differential cell counts in bronchoalveolar lavage fluid (BALF), and extensive cytokine profiling in both BALF and lung tissue homogenates, utilizing precise immunoassays and histopathological scoring systems. Results: Both SECU and HONK, when used alone or in combination, display significant immunomodulatory effects in a murine model of allergic asthma concomitant with ALI. The combined therapy synergistically reduced pro-inflammatory mediators, notably Th1 cytokines, such as TNF-α and IL-6, as measured in both BALF and lung tissue homogenates. Conclusions: The combined therapy showed a synergistic attenuation of pro-inflammatory mediators, a reduction in goblet cell hyperplasia, and an overall improvement in lung histoarchitecture. While the data robustly support the merit of a combinatorial approach targeting multiple inflammatory mediators, the study acknowledges limitations in cytokine diffusion and the murine model’s translational fidelity, thereby underscoring the need for further research to optimize clinical protocols for severe respiratory inflammatory disorders. Full article

Background/Objectives: The ongoing evolution of SARS-CoV-2 has highlighted the limitations of parenteral vaccines in preventing viral transmission, largely due to their failure to elicit robust mucosal immunity. Methods: Here, we evaluated an intranasal (IN) vaccine formulation consisting of recombinant receptor-binding domain (RBD) adsorbed onto human probiotic Bacillus subtilis DG101 spores. Results: In BALB/c mice, IN spore-RBD immunization induced strong systemic and mucosal humoral responses, including elevated specific IgG, IgM, and IgA levels in serum, bronchoalveolar lavage fluid (BALF), nasal-associated lymphoid tissue (NALT), and saliva. It further promoted mucosal B cell and T cell memory, along with a Th1/Tc1-skewed T cell response, characterized by increased IFN-γ-expressing CD4⁺ and CD8⁺ T cells in the lungs. Conclusions: All in all, these findings highlight the potential of intranasal vaccines adjuvanted with probiotic B. subtilis spores in inducing sterilizing immunity and limiting SARS-CoV-2 transmission. Full article

Background: The efficacy of EA575 in the treatment of respiratory diseases is described in various clinical studies, improving patients’ disease-related symptoms. However, mechanistic in vivo data proving its beneficial effects are limited. Methods: Focusing on the treatment of acute airway inflammation and accompanying cough, this study aimed to elucidate antitussive and mucoactive properties of EA575, applying two animal models. Animals were treated orally twice daily for 7 days, resulting in 43, 215.2, or 430.5 mg/kg bw/d of EA575. Antitussive effects were investigated within an acute lung inflammation model of bleomycin-treated guinea pigs after citric acid exposure. Hereby, the number of coughs, enhanced pause (penH), and bronchoalveolar lavage fluid (BALF) were investigated. Mucoactivity of EA575 was assessed within a murine model, determining phenol red concentration in BALF. Results: EA575 treatment within the acute lung inflammation model reduced cough events up to 56% while reducing inflammatory cell influx in BALF dose-dependently, e.g., reducing neutrophils in BALF up to 70.9%. This suggests a strong connection between anti-inflammatory and antitussive properties of EA575. Furthermore, penH decreased in a dose-dependent manner, suggesting an ease in respiration. Mucoactivity was shown by a dose-dependent increase in phenol red concentration in BALF up to 38.9%. Notably, EA575/salbutamol co-administration resulted in enhanced phenol red secretion compared to respective single administrations. Conclusions: These data highlight the benefits of EA575 in treating cough-related respiratory diseases, particularly when accompanied by sputum, as EA575 has been shown to obtain mucoactivity. Furthermore, the combinatory effect of EA575/salbutamol treatment provides a foundation for future research in the treatment of chronic respiratory diseases. Full article

Background: Acute cellular rejection (ACR) remains a common complication following lung transplantation and is a major risk factor for chronic lung allograft dysfunction (CLAD). Although transbronchial biopsy (TBB) is the diagnostic gold standard, it is invasive and may be contraindicated in certain patients. This study aimed to assess the diagnostic utility of combining bronchoalveolar lavage fluid (BALF) lymphocyte counts with cytokine profiling—particularly interleukin-17A (IL-17A)—in lung transplant recipients with elevated peripheral blood eosinophil (EOS) counts. Methods: We retrospectively analyzed 108 BALF and matched TBB samples from 74 lung transplant recipients with EOS counts >200 cells/μL, collected between 2014 and 2020. BALF lymphocyte percentages and levels of cytokines (IL-4, IL-6, IL-10, IL-13, IL-15, IL-17A, IFN-γ, TNF) were quantified. Associations with histologically confirmed ACR were evaluated using generalized estimating equation models. Results: ACR was diagnosed in 57% of TBB samples. BALF lymphocyte percentages were significantly higher in ACR cases (median 8% vs. 4%, p < 0.001). Each 1% increase in lymphocytes was associated with a 10% increase in the odds of ACR (OR 1.102; 95% CI 1.076–1.129). IL-17A levels were also significantly elevated in ACR (OR 1.047; 95% CI 1.003–1.092; p = 0.032), but with moderate discriminative ability (AUC = 0.629). The combination of BALF lymphocyte counts and IL-17A levels improved diagnostic performance (AUC > 0.76). Conclusions: The combined assessment of BALF lymphocyte counts and IL-17A levels in recipients with elevated EOS offers a promising non-invasive strategy to support the diagnosis of ACR. Prospective studies are needed to validate these findings and further refine personalized diagnostic approaches to ACR. Full article

Background/Objectives: Acute lung injury (ALI) is an inflammatory condition characterized by tissue barrier damage, which leads to vascular leakage, pulmonary edema, and compromised gas exchange. Lipopolysaccharides (LPS) are a component of Gram-negative bacteria, which trigger inflammation by Toll-like receptor 4 (TLR4) activation. Herein, we investigated the possibility that Pasireotide (PAS) exerts protective effects in an experimental model of ALI. Methods: C57BL/6 male mice received an intratracheal injection of saline or LPS, followed by PAS or vehicle treatment. Bronchoalveolar lavage fluid (BALF) was collected via tracheal catheterization, and Western blot analysis was used to detect protein expression variations. Results: Our results suggest that PAS treatment alleviates LPS-induced mouse lung injury and inflammation. JAK/STAT and MAPK activation levels in the inflamed lungs were suppressed due to PAS treatment, as well as BALF protein concentration. Additionally, PAS counteracted LPS-induced Grp94 protein reduction, suggesting the involvement of ATF6 in PAS-triggered barrier-protective effects. Grp94 is a downstream ATF6 target. Conclusions: Our data demonstrate that PAS protects mouse lungs against LPS in an experimental model of ALI. Full article

Aletris spicata (Thunb.) Franch. (AS), a traditional edible and medicinal plant for treating asthma, was investigated for its therapeutic mechanisms. Liquid chromatography-high-resolution tandem mass spectrometry (LC-HRMS/MS) analysis identified 33 compounds in AS. In ovalbumin (OVA)-induced asthmatic mice, AS significantly reduced inflammatory cells (neutrophils, lymphocytes, eosinophils) in bronchoalveolar lavage fluid (BALF) and decreased IL-4, IL-5, IL-13, TNF-α, and serum IgE while increasing IFN-γ. AS alleviated lung and intestinal inflammation, reduced ROS and MDA levels, and enhanced SOD activity. Immunohistochemistry and Western blot revealed AS upregulated Nrf2/HO-1 expression and inhibited NF-κB p65 nuclear translocation. Gut microbiota studies demonstrated AS restored intestinal flora homeostasis by modulating the richness, diversity, and composition. Spearman correlation analysis identified significant relationships between oxidative stress markers, inflammatory cytokines, and specific gut bacteria. These findings indicate that AS mitigates asthma through antioxidant effects (Nrf2/HO-1 pathway), anti-inflammatory actions (NF-κB pathway), and gut microbiota modulation. The study provides a scientific basis for developing AS as a natural anti-asthma treatment or functional food. The multi-target mechanism involving oxidative stress, inflammation, and gut flora highlights AS’s comprehensive therapeutic potential for asthma management. Full article

An experimental model of acute pulmonary damage was developed based on the intravenous injection of the phospholipase A₂ (PLA₂)-rich venom of Pseudechis papuanus (Papuan black snake) in mice. Venom caused pulmonary edema, with the accumulation of a protein-rich exudate, as observed histologically and by analysis of bronchoalveolar lavage fluid (BALF). In parallel, venom induced an increase in all of the pulmonary mechanical parameters evaluated, without causing major effects in terms of tracheal and bronchial reactivity. These effects were abrogated by incubating the venom with the PLA₂ inhibitor varespladib, indicating that this hydrolytic enzyme is responsible for these alterations. The venom was cytotoxic to endothelial cells in culture, hydrolyzed phospholipids of a pulmonary surfactant, and reduced the activity of angiotensin-converting enzyme in the lungs. The pretreatment of mice with the nitric oxide synthase inhibitor L-NAME reduced the protein concentration in the BALF, whereas no effect was observed when mice were pretreated with inhibitors of cyclooxygenase (COX), tumor necrosis factor-α (TNF-α), bradykinin, or neutrophils. Based on these findings, it is proposed that the rapid pathological effect of this venom in the lungs is mediated by (a) the direct cytotoxicity of venom PLA₂ on cells of the capillary–alveolar barrier, (b) the degradation of surfactant factor by PLA₂, (c) the deleterious action of nitric oxide in pulmonary tissue, and (d) the cytotoxic action of free hemoglobin that accumulates in the lungs as a consequence of venom-induced intravascular hemolysis. Our findings offer clues on the mechanisms of pathophysiological alterations induced by PLA₂s in a variety of pulmonary diseases, including acute respiratory distress syndrome (ARDS). Full article

Neutrophil extracellular traps (NETs) are associated with the extracellular release of nuclear chromatin decorated with cytoplasmic proteins. Excessive release of NETs has been reported in chronic lung diseases, including chronic obstructive pulmonary disease (COPD). However, the role of NETs in the pathogenesis of COPD remains unclear. Peptidylarginine deaminase 4 (PAD4) contributes to NET formation. Therefore, in an elastase (ELS)-induced emphysema mouse model, we examined the role of PAD4 using Padi4 gene knockout (KO) mice. First, we confirmed that ELS induced NET formation in the parenchyma of the lungs. PAD4 deficiency suppressed ELS-induced NET expression and tended to ameliorate the lung tissue injury. The cellular profile of bronchoalveolar lavage fluid (BALF) did not differ between the two groups. Additionally, PAD4 deficiency ameliorated emphysema and apoptosis in lung cells. Finally, we examined the effects of PAD4 on comprehensive gene expression signatures using RNA sequencing. Enrichment analysis of the transcriptomic data revealed that the expression of several genes associated with COPD pathogenesis was altered in the KO mice. Overall, the results suggest that PAD4 deficiency improves NET formation and emphysema in the lungs; this pathway can be a potential therapeutic target for the treatment of COPD. Full article

Ricin is a highly potent toxin that causes severe lung injury upon inhalation by initiating a complex cascade of cellular responses that ultimately leads to cell death. The low-density lipoprotein receptor-related protein 1 (LRP1) is a multifunctional receptor involved in various physiological processes, including ricin-mediated toxicity. This study explores the role of LRP1 shedding in the development of ricin-induced lung injury. Analysis of bronchoalveolar lavage fluid (BALF) from ricin-intoxicated mice and swine showed a significant increase in soluble LRP1 (sLRP1) levels, whereas serum LRP1 levels remained largely unchanged, suggesting the lungs are the primary source of sLRP1 release. In vitro assays demonstrated the formation of ricin-sLRP1 complexes, indicating that sLRP1 in BALF retained ricin-binding capability. Flow cytometric analysis of lung cells revealed a reduction in both the percentage and total number of LRP1-expressing cells following ricin exposure. Further investigation of specific lung cell populations showed that alveolar epithelial type II (AT-II) cells, despite experiencing significant injury, exhibited minimal LRP1 shedding. No shedding of LRP1 occurred in neutrophils. In contrast, fibroblasts, which were resistant to ricin-induced cell death, exhibited increased shedding of LRP1 and a corresponding decrease in membrane-bound LRP1 expression. This shedding of the LRP1 ectodomain was mediated by metalloproteinases. Immunohistochemical staining further confirmed decreased LRP1 expression in fibroblasts from ricin-exposed mice. Macrophages also showed substantial LRP1 shedding, despite undergoing significant depletion. These findings highlight the complex cell-specific nature of LRP1 shedding in response to ricin intoxication and suggests the potential role of LRP1 in modulation of cellular susceptibility and resistance to ricin-induced lung injury. Full article

Streptococcus pneumoniae (S. pneumoniae) is a leading cause of pneumonia, and its interaction with host acute-phase proteins remains underexplored. Serum amyloid A1 (SAA1), an acute-phase protein, plays a crucial role in immune modulation. This study investigates the role of SAA1 in the early stages of respiratory infection by S. pneumoniae and its potential contribution to bacterial adaptation under acidic stress. We used a murine nasal infection model to simulate the early phase of S. pneumoniae invasion into the lower respiratory tract. Levels of SAA1 and C-reactive protein (CRP) in bronchoalveolar lavage fluid (BALF) and serum were quantified using ELISA. In vitro assays examined the effect of serum and recombinant SAA1 on bacterial survival under acidic conditions. Fluorescence-labeled recombinant SAA1 and microscopy were utilized to assess SAA1 internalized by S. pneumoniae. Following nasal infection, SAA1 levels in BALF were significantly reduced, whereas CRP levels remained unchanged. In vitro, serum enhanced S. pneumoniae’s resistance to acidic byproducts including formic, lactic, and acetic acids. Specifically, formic acid promoted bacterial uptake of SAA1, and this internalization improved bacterial tolerance to acidic conditions. Fluorescence microscopy confirmed that SAA1 is internalized by S. pneumoniae. S. pneumoniae can internalize SAA1 to bolster resistance to acid stress, particularly formic acid. This study reveals a novel host–pathogen interaction mechanism wherein S. pneumoniae exploits host acute-phase proteins for environmental adaptation, offering new insights into bacterial survival strategies during infection. Full article

Ambient ozone (O₃) pollution, which has become a global problem, is associated with damage to various biological systems, as determined by many studies. However, there is limited experimental evidence regarding the systemic damage induced by O₃ exposure, and there are few associated studies on mice. In the present investigation, we constructed a subacute C57BL/6J female mouse model involving exposure to 0, 0.5, 1, or 2 ppm O₃ for 28 days (3 h/day). Body weights, pulmonary function, hematology, serum biochemistry, inflammatory factors, and injuries to various organs were assessed for O₃-exposed mice. After O₃ exposure, especially to 2 ppm O₃, mice showed a loss of body weight, abnormal glucose and lipid metabolism, respiratory and nervous system injuries, an inflammatory response, and pathological changes, which supported the data reported for epidemiology studies. In addition, the IL-6 levels in bronchoalveolar lavage fluid (BALF), the lungs, the livers, the kidneys, and the brains were increased, which indicated that IL-6 was associated with the damage to various organs induced by O₃ exposure. The present report highlights the pathological injury to various organs and provides a basis for further studies of the molecular mechanisms associated with O₃ exposure. Full article

Background: Bronchoalveolar lavage fluid (BALF) analysis is essential for the accurate diagnosis and management of interstitial lung diseases (ILDs). Despite established guidelines, variability in sample handling may affect diagnostic accuracy. This study aimed to evaluate how different storage conditions impact BALF cell counts and differentials to guide optimal sample handling practices. Methods: Forty patients who underwent BAL at Chiba University Hospital from June to December 2024 were included. BALF samples were allocated into five groups based on processing conditions: immediate analysis within 1 h, storage at either at 4 °C or room temperature (RT) for 6 h, or storage at 4 °C or RT for 24 h. Total cell counts (TCC) and differential counts were measured and compared among conditions. Results: TCC remained stable over 24 h at both 4 °C (p = 0.86) and RT (p = 0.90). Similarly, the percentages of eosinophils, lymphocytes, and macrophages did not significantly change at either temperature (all p > 0.05). Notably, neutrophil percentages showed a significant decline over time under both storage conditions—at 4 °C (p = 0.02) and at room temperature (p < 0.01). Post hoc tests revealed a notable decreasing trend at 6 h and significant reductions by 24 h at 4 °C (p = 0.09 and p = 0.02, respectively), and significant decreases at both 6 and 24 h at RT (p = 0.01, <0.01). Conclusions: Among the various cell types in BALF, neutrophil proportions are particularly susceptible to storage conditions, showing a significant decline over time—especially at room temperature—while other cell types remain stable for up to 24 h. Therefore, prompt processing or appropriate refrigeration of BALF is essential to ensure reliable cytological analysis and accurate clinical interpretation. Full article

Pulmonary exposure to carbon nanotubes (CNTs) has been linked to a series of adverse respiratory effects in animal models, including inflammation, genotoxicity, fibrosis, and granuloma formation, the degree and characteristics of which are considered dependent upon the detailed physicochemical properties of the material as inhaled. To further explore the effect of variations in physicochemical properties on pulmonary effects, two different multi-walled CNTs (MWCNTs) were tested in vivo: a pristine MWCNT (pMWCNT) (NM-401) and a surface-modified MWCNT (MWCNT-COOH). Female Sprague–Dawley rats were whole-body exposed for 28 days to MWCNT aerosols (pMWCNT (0.5 and 1.5 mg/m³) and MWCNT-COOH (1.5 and 4.5 mg/m³)) and followed up to 1 year post-exposure. The inhalation exposures resulted in relatively low estimated lung deposition. Bronchoalveolar lavage fluid (BALF) analysis indicated inflammation levels broadly consistent with deposited dose levels. Lung histopathology indicated that both MWCNTs produced very limited toxicological effects; however, global mRNA expression levels in lung tissue and BALF cytokines indicated different characteristics for the two MWCNTs. For example, pMWCNT but not MWCNT-COOH exposure induced osteopontin production, suggestive of potential pre-fibrosis/fibrosis effects linked to the higher aspect ratio aerosol particles. This is of concern as brightfield and enhanced darkfield microscopy indicated the persistence of pMWCNT fibres in lung tissue. Full article

Background: Tuberculosis (TB) is a respiratory infectious disease, and the current TB vaccine has low local lung protection. We aim to optimize immune pathways to improve the immunogenicity of vaccines. Methods: In the immunogenicity study, 50 BALB/c mice were randomly divided into the following: (1) phosphate buffered saline (PBS)+intramuscular injection combined with electroporation (EP) group (100 μL), (2) pVAX1+EP group (50 μg/100 μL), (3) ag85ab+EP group (50 μg/100 μL), (4) pVAX1+pulmonary delivery (PD) group (50 μg/50 μL), and (5) ag85ab+PD group (50 μg/50 μL). Immunization was given once every 2 weeks for a total of three times. The number of IFN-γ-secreting lung and spleen lymphocytes was determined by enzyme-linked immunospot assay (ELISPOT). The levels of Th1, Th2, and Th17 cytokines in the culture supernatants of lung and spleen lymphocytes were detected with the Luminex method. The proportion of FoxP3 regulatory T cells in splenocytes was determined by flow cytometry. The levels of IgG-, IgG1-, and IgG2a-specific antibodies in plasma and IgA antibody in bronchoalveolar lavage fluid (BALF) were determined by enzyme-linked immunosorbent assay (ELISA). Results: The PD and EP routes of Mycobacterium tuberculosis (M. tb) ag85ab DNA vaccine can effectively induce the responses of IFN-γ-secreting lung and spleen lymphocytes, and induce dominant Th1 and Th17 cell immune responses. The PD route can induce earlier, greater numbers and stronger responses of pulmonary effector T cells, with higher levels of the specific antibody IgA detected in BALF. High levels of the specific antibodies IgG, IgG1, and IgG2α were detected in the plasma of mice immunized by the EP route. Conclusions: The PD route of DNA vaccines can more effectively stimulate the body to produce strong cellular and mucosal immunity than the EP route, especially local cellular immunity in the lungs, which can provide early protection for the lungs. It can significantly improve the immunogenicity of the ag85ab DNA vaccine, suggesting a feasible and effective approach to DNA immunization. Full article

Perfluorooctane sulfonate (PFOS), a persistent organic pollutant, has raised significant public health concerns because of its widespread environmental presence and potential toxicity. Epidemiological studies have linked PFOS exposure to respiratory diseases, but the underlying molecular mechanisms remain poorly understood. Male C57 BL/6J mice were divided into a control group receiving Milli-Q water, a low-dose PFOS group (0.2 mg/kg/day), and a high-dose PFOS group (1 mg/kg/day) administered via intranasal instillation for 28 days. Lung tissue transcriptome sequencing revealed significantly enriched differentially expressed genes in the Ras and Rap signaling pathways. Key genes including Rap1b, Kras, and BRaf as well as downstream genes, such as MAPK1 and MAP2K1, exhibited dose-dependent upregulation in the high-dose PFOS exposure group. Concurrently, the downstream effector proteins MEK, ERK, ICAM-1, and VEGFa were significantly elevated in bronchoalveolar lavage fluid (BALF). These alterations are mechanistically associated with increased oxidative stress, inflammatory cytokine release, and pulmonary tissue damage. The results indicated that PFOS-induced lung injury is likely predominantly mediated through the activation of the Rap1b- and Kras-dependent BRaf-MEK-ERK axis. These findings highlight the critical role of Ras/Rap signaling pathways in PFOS-associated respiratory toxicity and underscore the need to develop therapeutic interventions targeting these pathways to mitigate associated health risks. Full article