Search Results (210)

Despite the widespread accessibility of vaccines and antivirals, seasonal influenza virus epidemics continue to pose a threat to public health. In this study, we constructed a recombinant replication-deficient simian adenovirus type 25 vector carrying the full-length hemagglutinin (HA) of the H1N1 influenza virus, named rSAd25-H1. Both systemic and mucosal humoral immune responses, as well as the protective efficacy, were assessed in mice immunized via the intramuscular (IM) or intranasal (IN) route. A single-dose IM or IN administration of rSAd25-H1 elicited a robust systemic IgG antibody response, including hemagglutination inhibition antibodies. As expected, only IN immunization was able to induce IgA production in serum and respiratory mucosa. Notably, a single dose of rSAd25-H1 at the highest dose (10¹⁰ viral particles) conferred complete protection against lethal homologous H1N1 challenge in mice despite the route of administration. These findings demonstrate the potential of simian adenovirus type 25-based vectors as a promising candidate for intranasal vaccine development targeting respiratory pathogens. Full article

Respiratory viral diseases infecting poultry lead to variable lesions in the respiratory organs, including nasal sinuses, trachea, lungs, and air sacs. Additional involvement of eyelids/conjunctiva was reported. The distribution and the intensity of lesions depend on multiple factors, including virulence, the host’s immunity, and secondary or concurrent infections. It may be challenging to detect remarkable lesions during experimental infections conducted in a controlled environment because some viruses fail to produce the intense lesions seen in field cases. This creates a challenge in developing a reliable model to study pathogenicity or vaccine efficacy experimentally. The development of the proposed histologic respiratory index (HRI) aims to help monitor the least microscopic changes that can be scored, thereby creating an objective and accurate grading of lesions in experimentally infected birds. HRI scores the changes in eyelids/conjunctiva and respiratory mucosa, including hyperplasia, metaplasia, inflammatory cellular infiltration in the submucosa, including lymphocytes and heterophils, and vascular changes (vasculitis) in nasal sinuses, trachea, and lungs. The score was validated in birds infected experimentally with avian metapneumovirus (aMPV) and low pathogenic avian influenza (LPAI-H4N6). The HRI reliably graded higher scores in the respiratory organs of experimentally infected birds compared with non-infected control ones. The HRI is the first of its type with poultry viral respiratory pathogens and it was initially proven to be a reliable in pathogenicity and vaccine trials of certain poultry respiratory viral diseases. Full article

The equine respiratory and reproductive tract microbiomes are complex and subject to constant fluctuations. Among the microbial inhabitants, Streptococcus equi subsp. zooepidemicus (SEZ) is recognized as the dominant bacterium. It is an opportunistic pathogen that may occasionally lead to various types of infections. A key virulence factor of SEZ is the streptolysin S (SLS) toxin, which is responsible for the characteristic β-hemolysis on blood agar and tissue damage. Viruses and bacteria may interact and aggravate lesions and disease. This study aimed to evaluate the impact of an SLS-containing supernatant from SEZ on the nasal and vaginal mucosa and the subsequent replication of equine herpesviruses. The SLS-containing supernatant was prepared, and three 10-fold dilutions (optical density “OD” 10⁻², 10⁻³, 10⁻⁴) were applied to equine nasal and vaginal explants. Untreated and EGTA-treated explants served as controls. Epithelial integrity was assessed by measuring the thickness and intercellular spaces. Nasal explants were inoculated with EHV-1 and EHV-4, while vaginal explants received EHV-1 and EHV-3. Viral replication was estimated via immunofluorescence staining and confocal microscopy. SLS-containing supernatants 10⁻² and 10⁻³ compromised epithelial integrity. Viral replication increased in explants treated with SLS 10⁻³, demonstrating SLS’s damaging effects on the epithelium, facilitating equine herpesvirus replication. Full article

Calreticulin is a highly conserved and multifunctional molecular chaperone ubiquitously expressed in humans and animals. Beyond its well-established roles in calcium homeostasis, protein folding, and immune regulation, recent studies in aquatic species have suggested a previously unrecognized antimicrobial function of calreticulin. These findings raise the question of whether calreticulin also exerts antibacterial activity in terrestrial mammals, which has not been systematically investigated to date. To address this knowledge gap, we successfully constructed and expressed recombinant goat calreticulin using the Pichia pastoris expression system, yielding a protein of over 99% purity that predominantly exists in dimeric form. Functional assays demonstrated that both recombinant goat and human calreticulin exhibited preliminary inhibitory activity against Escherichia coli, Salmonella typhimurium, and Pasteurella multocida. Calreticulin was capable of binding to these three bacterial species as well as bacterial lipopolysaccharides (LPS). Notably, in the presence of Ca²⁺, calreticulin induced bacterial aggregation, indicating a potential mechanism for limiting bacterial dissemination and proliferation. Given the high anatomical, genetic, and physiological similarity between goats and humans—particularly in respiratory tract structure and mucosal immune function—neonatal goats were selected as a relevant model for evaluating the in vivo antimicrobial efficacy of calreticulin. Accordingly, we established an intranasal infection model using Pasteurella multocida to assess the protective role of calreticulin against respiratory bacterial challenge. Following infection, calreticulin expression was markedly upregulated in the nasal mucosa, trachea, and lung tissues. Moreover, intranasal administration of exogenous calreticulin significantly alleviated infection-induced pathological injury to the respiratory system and effectively decreased bacterial loads in infected tissues. Collectively, this study systematically elucidates the antimicrobial activity of calreticulin in a mammalian model and highlights its potential as a natural immune effector, providing novel insights for the development of host-targeted antimicrobial strategies. Full article

Respiratory syncytial virus (RSV) is a major etiological agent of lower respiratory tract infections, particularly among infants and the elderly. Activation of B cells in the mucosa and the production of specific neutralizing antibodies are essential for protective immunity against pulmonary infection. B-cell activating factor (BAFF) is a critical survival factor for B cells and has been associated with antiviral responses; however, its regulation during RSV infection remains poorly understood. This study examined BAFF regulation in BEAS-2B cells exposed to RSV or IFN-β. The treatments resulted in a progressive increase in gene expression over time, accompanied by higher protein levels. BAFF mRNA peaked at 12 h post-infection and declined by 48 h, coinciding with the release of soluble BAFF protein into the culture supernatant. Pre-treatment with anti-IFN-β antibodies prior to RSV infection reduced both BAFF mRNA and protein levels, indicating that IFN-β plays a regulatory role in BAFF production by airway epithelial cells. Western blot analysis revealed membrane-bound BAFF (~31 kDa) in non-infected cells, with elevated expression at 24 h post-infection. By 48 h, this form was cleaved into a soluble ~17 kDa form, which was detected in the supernatant. Immunostaining further demonstrated reduced surface expression of membrane-bound BAFF in RSV-infected cells compared to uninfected controls, suggesting that RSV infection promotes the cleavage and release of BAFF into the extracellular environment. Additionally, the release of BAFF was not affected by furin convertase inhibition or ER–Golgi transport blockade, indicating a potentially novel cleavage mechanism. Co-culturing BAFF produced by BEAS-2B cells with isolated B cells enhanced B cell viability. Overall, these results indicate that RSV infection stimulates BAFF production in airway epithelial cells through a pathway involving IFN-β, potentially contributing to B cell activation and promoting local antibody-mediated immunity. Understanding this mechanism may offer valuable insights for improving mucosal vaccine strategies and enhancing immunity against respiratory pathogens. Full article

Particulate matter (PM) is a major environmental pollutant implicated in various respiratory diseases. However, its impact on the upper respiratory tract, particularly the nasal and paranasal sinus mucosa, remains poorly understood. This study aimed to investigate the acute inflammatory effects of PM exposure on the sinonasal mucosa and evaluate the natural recovery process in a controlled rat model. Ten-week-old male Sprague Dawley rats were exposed to incense-derived PM in a custom-designed exposure chamber for 2 h daily for seven consecutive days. Rats were sacrificed at 3, 7, and 14 days post-exposure. Histopathologic changes were assessed using hematoxylin and eosin and Alcian blue staining, and mucosal gene expression of inflammatory cytokines including interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, interferon (IFN)-γ, and IL-5 and MUC5AC was quantified using real-time reverse transcription polymerase chain reaction. PM exposure induced significant histological alterations, including epithelial thickening, inflammatory cell infiltration, and goblet cell hyperplasia, which peaked at 7 days post-exposure. Expression levels of TNF-α and IFN-γ were significantly elevated at 7 days compared to controls. The sinonasal mucosa in the 14-day post-exposure groups exhibited a remarkable decrease in goblet cell numbers, and IL-1β and TNF-α expression. Short-term exposure to high concentrations of PM resulted in acute inflammatory changes in the sinonasal mucosa of rats, including epithelial thickening and goblet cell hyperplasia. These changes were partially resolved after exposure ended, indicating that PM-induced sinonasal inflammation may be at least partially reversible. Full article

Background/Objectives: Coronaviruses (CoVs) are a large family of respiratory viruses that cause respiratory illnesses ranging from mild colds to severe diseases such as severe acute respiratory syndrome and pandemics. The nasal cavity is a primary site for CoV entry and transmission. The study aimed to prepare a novel mucoadhesive emulgel specifically formulated with simple, safe, and cost-effective excipients to create a barrier on the nasal mucosa that impedes CoV infection. This formulation strategy was specifically designed to enable rapid and straightforward in vivo translation, addressing a critical gap in preventive measures against respiratory viruses. Methods: Three emulgels, containing macadamia oil, Carbopol and different percentages of hydroxypropyl methylcellulose (1, 1.2 and 1.5% (w/v), HPMC), were properly prepared and characterized for mucoadhesion, viscosity, and spreadability. The biological activity against SARS-CoV-2 was evaluated in vitro on infected epithelial cells. Results: The emulgel with 1.2% HPMC demonstrated optimal physicochemical properties (mucoadhesion: 342 ± 9 mN/cm²; viscosity: 1080 ± 83 cp; spreadability: 7.27 ± 0.06 cm) suitable for nasal application. Importantly, in vitro biological assays demonstrated that this emulgel significantly inhibits SARS-CoV-2 infection in epithelial cells, indicating an effective barrier to viral diffusion. Conclusions: By employing readily available, safe, and inexpensive excipients, this novel mucoadhesive emulgel offers a practical, scalable, and rapidly translatable nasal prophylactic approach to prevent early SARS-CoV-2 infection, addressing a critical unmet need in pandemic preparedness. Full article

Background and Objectives: The common cold (acute rhinopharyngitis) and acute rhinosinusitis are highly prevalent conditions that significantly impact quality of life, often leading to nasal congestion, inflammation, and discomfort. Given the growing demand for non-pharmacological treatment options, particularly for vulnerable populations such as children and pregnant women, alternative therapies are increasingly being explored. NESOSPRAY HE-C, a nasal spray formulated with a glycerol-based filmogenic solution, acts by forming a protective osmotic film on the nasal mucosa. This mechanism facilitates mechanical cleansing, enhances decongestion, and reduces inflammation while preserving mucosal integrity. Its purely topical and mechanical mode of action provides a non-systemic alternative for symptom management. Materials and Methods: This randomized, double-blind, parallel-group clinical trial evaluated the efficacy and safety of NESOSPRAY HE-C (n = 29) compared to a placebo nasal spray (n = 26) in patients aged ≥ 3 years diagnosed with the common cold or acute rhinosinusitis. Participants had a baseline Rhinosinusitis Symptom Severity Score (RSSS) of ≥25/50. Treatment consisted of administering 2–3 sprays per nostril, four times daily, every 4 to 6 h, for up to 8 days or until symptom resolution. The primary outcomes included changes in total RSSS, Wisconsin Upper Respiratory Symptom Survey (WURSS) score, and individual symptom scores (rhinorrhea, nasal congestion, cough, poor sleep, facial pain, and fever). Safety assessments included adverse event monitoring and treatment tolerability, with subgroup analyses performed for children and pregnant women. Results: Baseline demographics were comparable between the treatment groups. NESOSPRAY HE-C demonstrated a significantly greater reduction in total RSSS from Day 3 onward (p = 0.0008), with sustained superiority through Day 8 (p < 0.0001). Significant improvements in rhinorrhea and nasal congestion were observed within 2 h of administration (p = 0.0089), while reductions in cough (p = 0.0052), poor sleep (p = 0.0005), and facial pain (p = 0.0111) emerged by Day 3. Fever reduction was most pronounced on Days 6 (p = 0.0001) and 8 (p = 0.0312), indicating a delayed but significant effect. In terms of the WURSS score, NESOSPRAY HE-C showed a significant improvement from Day 1, with a greater reduction in symptom severity compared to placebo. This trend of greater improvement continued through Day 8. The treatment was well tolerated, with no reports of serious adverse events or allergic reactions. Efficacy was consistent across all subgroups, including children, pregnant women, and adults. Conclusions: NESOSPRAY HE-C provides rapid and sustained symptom relief for the common cold and acute rhinosinusitis, serving as a safe and effective non-pharmacological alternative to conventional treatments. By leveraging its osmotic action and barrier-forming properties, it facilitates mechanical cleansing, enhances decongestion, and reduces inflammation while preserving mucosal integrity. Additionally, by forming a protective film on the nasal mucosa, it protects against future irritations, further supporting its role as a valuable therapeutic option, particularly for individuals seeking non-systemic symptom management. Full article

Post-COVID-19 is a chronic infection-related syndrome, including exacerbations of pre-existing or newly diagnosed conditions that have been established after the acute phase of COVID-19 and have demonstrated a wide range of systemic effects beyond the lungs. SARS-CoV-2 attaches to its receptor, angiotensin-converting enzyme 2 (ACE-2). Transmembrane serine protease 2 (TMPRSS2) facilitates viral entry and spread. ACE-2 receptors are detectable in several tissues, including the respiratory mucosa, digestive tract, heart, kidney, and brain. Several investigations have demonstrated an increase in digestive manifestations post-acute COVID-19, likely related to an alteration in the intestinal microbiota following infection. These changes can lead to a loss of species diversity, resulting in an overgrowth of opportunistic pathogens and deprivation of commensal bacteria. In this context, post-infection irritable bowel syndrome shows an increased incidence compared to controls. Growing evidence also suggests the enduring presence of SARS-CoV-2 in the gut tissue. Studies are ongoing to investigate antiviral agents that counteract prolonged COVID-19 symptoms. Therefore, the objectives of this review were to summarize the digestive manifestations, focusing on irritable bowel syndrome and therapeutic strategies. This review gives an overview of studies published in English in the last two years on the PubMed database. Full article

Respiratory syncytial virus (RSV) is the leading cause of respiratory infections in children. Extracellular vesicles (EVs), released by airway epithelial cells, contain proteins and different families of non-coding RNAs (EV cargo) that can modulate the responses of target cells to viral infection. Nasal mucosa is a primary site of viral entry and the source of EVs present in the upper airway secretions. In this study we characterized proteins, including inflammatory mediators and cytokines, and the piwi-interacting RNA (piRNAs) cargo of EVs isolated from pediatric human nose organoids (HNO) and nasopharyngeal secretions (NPS) positive for RSV. Using Proximity Extension Assay (PEA) and Luminex multi-target arrays, we found significant enrichment in several chemokines and other mediators/biomarkers, including CCL2, CCL20, CXCL5, CX3CL1, CXCL6, MMP-1, MMP-10, uPA, Flt3L, ARNT and CD40 in EVs secreted by RSV-infected HNO compared to control mock HNO. Analysis of NPS samples from RSV infected children revealed that CCL3, CCL20, CXCL8, uPA, VEGFA, were concentrated in the NPS-EV fraction. LC-MS/MS and Gene Ontology indicated that RSV positive NPS-EVs originate from different cellular sources, with the most abundant proteins from neutrophils and epithelial cells. A total of 490 piRNAs were detected by NGS sequencing of small RNA libraries obtained from NPS-EVs, which has not been reported prior to this study. Identification of inflammatory mediators and small non-coding RNAs which are compartmentalized in EVs contributes to understanding mechanisms of virus-mediated pathogenesis in RSV infections. Full article

Aging affects all tissues in an organism, including the tracheobronchial tree, with structural and functional changes driven by mechanisms such as oxidative stress, cellular senescence, epigenetic modifications, mitochondrial dysfunction, and telomere shortening. Airway aging can be accelerated by intrinsic or extrinsic factors. This review brings together information from the literature on the molecular changes occurring in all layers of the tracheobronchial airway wall. It examines the biomolecular changes associated with aging in the mucosa, submucosa, cartilage, and smooth muscle of the airways. At the mucosal level, aging reduces ciliary function and disrupts mucin homeostasis, impairing mucociliary clearance and contributing to chronic respiratory diseases such as COPD (Chronic Obstructive Pulmonary Disease). Cellular senescence and oxidative stress drive extracellular matrix remodeling and chronic inflammation. Airway cartilage undergoes age-related changes in collagen and fibronectin composition, leading to increased stiffness, while heightened MMP (Matrix Metalloproteinases) activity exacerbates ECM (extracellular matrix) degradation. In airway smooth muscle, aging induces changes in calcium signaling, hypertrophy, and the secretion of pro-inflammatory mediators, further perpetuating airway remodeling. These changes impair respiratory function and increase susceptibility to chronic respiratory conditions in the elderly. By consolidating current knowledge, this review aims to provide a comprehensive overview of the molecular changes occurring in the respiratory tract with aging and to highlight new molecular perspectives for future research on this topic. Full article

Respiratory virus infections, such as those caused by influenza viruses, respiratory syncytial virus (RSV), and coronaviruses, pose a significant global health burden. While the immune system’s adaptive components, including memory T cells, are critical for recognizing and combating these pathogens, recurrent infections and variable disease outcomes persist. Memory T cells are a key element of long-term immunity, capable of responding swiftly upon re-exposure to pathogens. They play diverse roles, including cross-reactivity to conserved viral epitopes and modulation of inflammatory responses. However, the protective efficacy of these cells is influenced by several factors, including viral evolution, host age, and immune system dynamics. This review explores the dichotomy of memory T cells in respiratory virus infections: their potential to confer robust protection and the limitations that allow for breakthrough infections. Understanding the underlying mechanisms governing the formation, maintenance, and functional deployment of memory T cells in respiratory mucosa is critical for improving immunological interventions. We highlight recent advances in vaccine strategies aimed at bolstering T cell-mediated immunity and discuss the challenges posed by viral immune evasion. Addressing these gaps in knowledge is pivotal for designing effective therapeutics and vaccines to mitigate the global burden of respiratory viruses. Full article

Background/Objectives: Human parainfluenza virus type 1 (hPIV-1) is a major cause of serious respiratory diseases in young children. Annually, hPIV-1 results in approximately 10,000 hospitalizations in the United States due to croup, bronchiolitis, and/or pneumonia, and 10,000 deaths worldwide due to acute lower respiratory tract infections among children less than 5 years of age. Despite the burden of disease, no vaccine for hPIV-1 is currently approved. Sendai virus (SeV) is a murine PIV-1. It has structural similarities with hPIV-1 and is currently under clinical development as an hPIV-1 Jennerian vaccine. Attributes of SeV include the following: (a) needleless delivery, (b) rapid and durable serum antibody responses after a single intranasal administration, (c) durable IgG and IgA responses in the nasal mucosa, and (d) use as a platform for recombinant vaccines against multiple pediatric pathogens. Evaluation of the tolerability, safety, and immunogenicity of intranasal SeV in healthy adults and seropositive children 3 to 6 years of age was previously conducted and supported vaccine advancement to evaluation in younger children. Methods: Three seropositive children 1 to 2 years of age received a single intranasal dose of 5 × 10⁵ EID₅₀ SeV (SENDAI, Clinicaltrials.gov NCT00186927). Adverse events were collected for 28 days post-vaccine administration using diary cards and participants were followed for six months in total. Sera were collected longitudinally for clinical laboratory and virus-specific antibody tests. Nasal swabs were collected longitudinally for virus and mucosal antibody tests. Results: Intranasal SeV was well tolerated, with only mild grade 1–2 events that resolved spontaneously. No serious adverse events, medically attended adverse events, or adverse events causing protocol termination were reported. One participant had positive nasal swabs for inoculated SeV during the first week after vaccination. Although children had measurable PIV-1-specific serum antibodies at baseline, intranasal SeV vaccination resulted in significant serum antibody increases in all participants. Similarly, there were significant increases in PIV-1-specific nasal IgG and IgA levels in all participants. Elevated antibody levels persisted through the six months of follow-up. Conclusions: Intranasal SeV was well tolerated and uniformly immunogenic in seropositive children 1 to 2 years of age. Results encourage the further evaluation of SeV and SeV-based recombinants as potential intranasal vaccines for the prevention of infection by hPIV-1 and other serious respiratory pathogens. Full article

Oral Allergy Syndrome (OAS) is an allergic reaction that occurs upon contact of the mouth and throat with food, leading to symptoms primarily affecting the oral mucosa. In patients with allergic rhinitis, OAS may develop due to cross-reactivity between the pollen allergens responsible for allergic rhinitis, and specific plant-derived foods. This particular type of OAS is known as Pollen Food Allergy Syndrome (PFAS). The difference in prevalence of PFAS across different regions of the world is attributed to various factors, including environmental exposure and dietary habits. Southern Europe’s temperate climate favors the blooming of many allergenic plants, making respiratory allergies and PFAS significant public health concerns. There is a regional variation in pollen in Southern Europe, contributing to differences in the presence of panallergens—such as profilins, pathogenesis-related class 10 (PR-10) proteins and lipid transfer proteins (LTPs)—which mediate PFAS. In order to examine the epidemiology, pathogenesis, and diagnostic approaches of OAS and PFAS, focusing on their prevalence and impact in Southern European adults, a narrative review was performed. Data from Portugal, Spain, France, Italy, Albania, Greece, and Türkiye were retrieved. The main outcome of this review was that the frequency of PFAS varies across studies, not only between countries but also within the same country, due to vegetation variability across regions as well as methodological differences and the year of study. However, despite these differences, PFAS emerges as a common issue in Southern Europe, underscoring the need for effective diagnosis and management. Full article

Background/Objectives: In vitro models play a crucial role in preclinical respiratory research, enabling the testing and screening of mucosal formulations, dosage forms, and inhaled drugs. Mucociliary clearance (MCC) is an essential defense mechanism in mucosal drug delivery but is often impaired in respiratory diseases. Despite its importance, standardized in vitro MCC assays are rarely reported. Furthermore, many published methods primarily measure cilia beat frequency (CBF), which requires high-speed cameras that are not accessible to all laboratories. Therefore, this study aimed to develop a physiologically relevant, differentiated in vitro model of the respiratory epithelium that incorporates both beating cilia and functional MCC. We chose porcine airway mucosa as an alternative to human tissue due to ethical considerations and limited availability. The established model is designed to provide a reproducible and accessible method for a broad range of research laboratories. Methods: The previously published tracheal mucosal primary cell (TMPC DS) model, derived from porcine tissue, lacked the presence of beating cilia, which are crucial for effective MCC analysis. For accurate MCC assessment, beating cilia are essential as they play a key role in mucus clearance. To address this limitation, the here-described ciliated tracheal mucosal primary cell (cTMPC) model was developed. cTMPCs were isolated from porcine tissue and cultured under air–liquid interface (ALI) conditions for 21 days to promote differentiation. This model was evaluated for cell morphology, tight junction formation, ciliated and mucus-producing cells, barrier function, gene expression, and tracer/IgG transport. MCC and the model’s suitability for standardized MCC assays were assessed using an inverted microscope. In contrast to the TMPC DS model, which lacked beating cilia and thus could not support MCC analysis, the cTMPC model allows for comprehensive MCC studies. Results: The developed differentiated in vitro model demonstrated key structural and functional features of the respiratory epithelium, including well-differentiated cell morphology, tight junction integrity, ciliated and mucus-producing cells, and effective barrier function. Functional MCC was observed, confirming the model’s potential for standardized clearance assays. Conclusions: This differentiated in vitro model closely replicates the structural and functional characteristics of in vivo airways. It provides a valuable platform for studying mucociliary clearance, toxicology, drug uptake, and evaluating mucosal formulations and dosage forms in respiratory research. Full article