Mucosal TLR2-activating protein-based vaccination induces potent pulmonary immunity and protection against SARS-CoV-2 in mice

Current vaccines against SARS-CoV-2 substantially reduce mortality, but protection against infection is less effective. Enhancing immunity in the respiratory tract, via mucosal vaccination, may provide protection against infection and minimise viral spread. We tested a novel subunit vaccine in mice, consisting of SARS-CoV-2 Spike protein with a TLR2-stimulating adjuvant, delivered to mice parenterally or mucosally. Both routes of vaccination induced substantial neutralising antibody (nAb) titres, however, mucosal vaccination uniquely generated anti-Spike IgA, increased nAb in the serum and airways, and increased lung CD4+ T-cell responses. TLR2 is expressed by respiratory epithelia and immune cells. Using TLR2 deficient chimeric mice, we determined that TLR2 expression in either compartment facilitated early innate responses to mucosal vaccination. By contrast, TLR2 on hematopoietic cells was essential for optimal lung-localised, antigen-specific responses. In a K18-hACE2 mice, vaccination provided complete protection against disease and sterilising lung immunity against SARS-CoV-2. These data support mucosal vaccination as a strategy to improve protection in the respiratory tract against SARS-CoV-2 and other respiratory viruses.

The Betacoronavirus PHEV Replicates and Disrupts the Respiratory Epithelia and Upregulates Key Pattern Recognition Receptor Genes and Downstream Mediators, Including IL-8 and IFN-λ

The neurotropic betacoronavirus porcine hemagglutinating encephalomyelitis virus (PHEV) primarily infects and replicates in the swine upper respiratory tract, causing vomiting and wasting disease and/or encephalomyelitis in suckling pigs. This study investigated the modulation of key early innate immune genes at the respiratory epithelia in vivo, on tracheal tissue sections from experimentally infected pigs, and in vitro , on air-liquid interface porcine respiratory cell cultures.

Squamous and Respiratory Metaplasia After Olfactory Mucosal Resection

Resection of the olfactory mucosa (OM) is sometimes unavoidable during surgery; however, it is not known whether the OM can completely recover thereafter. The aim of this study was to uncover whether the OM fully recovers after mucosal resection and describe the process of OM regeneration. 8-week-old male Sprague–Dawley rats (n = 18) were subjected to OM resection at the nasal septum; six rats were euthanized for histological examination 0, 30, and 90 days after surgery. Immunohistochemistry was performed to identify olfactory receptor neuron (ORN) lineage cells [mature and immature ORNs and ORN progenitors, and olfactory ensheathing cells (OECs)], as well as dividing and apoptotic cells. Squamous and respiratory metaplasia and inflammatory cell infiltration were also assessed. On day 30 after resection, the mucosa had regenerated, and mainly contained thin nerve bundles, basal cells, and immature ORNs, with a few mature ORNs and OECs. On day 90, the repaired nasal mucosa had degenerated into stratified squamous or ciliated pseudostratified columnar epithelia, with reducing ORNs. The lamina propria contained numerous macrophages. Partial regeneration was observed within 1 month after OM resection, whereas subsequent degeneration into squamous and respiratory epithelia occurred within 3 months. Given the poor persistence of ORNs and OECs, OM resection is likely to result in olfactory impairment. Overall, surgeons should be cautious not to injure the OM during surgery.

Case Report: Esophageal Bronchus in a Neonate, With Image, Histological, and Molecular Analysis

In this case report, we describe the clinical course of a neonate who presented initially with respiratory distress and later with choking during feeding. He was subsequently found to have an esophageal bronchus to the right upper lung lobe, a rare communicating bronchopulmonary foregut malformation. Histological and molecular analysis of the fistula and distal tissues revealed that the proximal epithelium from the esophageal bronchus has characteristics of both esophageal and respiratory epithelia. Using whole exome sequencing of the patient's and parent's DNA, we identified gene variants that are predicted to impact protein function and thus could potentially contribute to the phenotype. These will be the subject of future functional analysis.

Chr15q25 Genetic Variant rs16969968 Alters Cell Differentiation in Respiratory Epithelia

The gene cluster region, CHRNA3/CHRNA5/CHRNB4, encoding for nicotinic acetylcholine receptor (nAChR) subunits, contains several genetic variants linked to nicotine addiction and brain disorders. The CHRNA5 single-nucleotide polymorphism (SNP) rs16969968 is strongly associated with nicotine dependence and lung diseases. Using immunostaining studies on tissue sections and air-liquid interface airway epithelial cell cultures, in situ hybridisation, transcriptomic and cytokines detection, we analysed rs16969968 contribution to respiratory airway epithelial remodelling and modulation of inflammation. We provide cellular and molecular analyses which support the genetic association of this polymorphism with impaired ciliogenesis and the altered production of inflammatory mediators. This suggests its role in lung disease development.

At the Intersection Between SARS-CoV-2, Macrophages and the Adaptive Immune Response: A Key Role for Antibody-Dependent Pathogenesis But Not Enhancement of Infection in COVID-19

Since entering the world stage in December of 2019, SARS-CoV-2 has impacted every corner of the globe with over 1.48 million deaths and caused untold economic damage. Infections in humans range from asymptomatic to severe disease associated with dysregulation of the immune system leading to the development of acute respiratory distress syndrome (ARDs).The distinct shift in peripheral monocyte activation and infiltration of these cells into the respiratory tract in ARDs patients suggests severe COVID-19 may largely result from damage to the respiratory epithelia by improperly activated macrophages. Here, we present evidence that dysregulation of the immune response in COVID-19 begins with activation of macrophages by non-neutralizing antibodies and induction of ACE2 expression, rendering these cells susceptible to killing by SARS-CoV-2. Death of macrophages occurs independently of viral replication and leads to the release of inflammatory mediators and modulation of the susceptibility of downstream epithelial cells to SARS-CoV-2.

Haemophilin-Producing Strains of Haemophilus haemolyticus Protect Respiratory Epithelia from NTHi Colonisation and Internalisation

Nontypeable Haemophilus influenzae (NTHi) is a significant respiratory tract pathogen responsible for infections that collectively pose a substantial health and socioeconomic burden. The clinical course of these infections is largely dictated by NTHi interactions with host respiratory epithelia, and thus, approaches that disrupt colonisation and invasion may have significant therapeutic potential. Survival, successful host–cell interactions, and pathogenesis are reliant on NTHi’s ability to sequester host-derived haem. Previously, we demonstrated the therapeutic potential of exploiting this haem-dependence using a closely related competitor bacterium, Haemophilus haemolyticus (Hh). Hh strains capable of producing the novel haem-binding protein haemophilin (Hpl) possessed potent inhibitory activity by restricting NTHi access to haem in a broth co-culture environment. Here, we extend this work to cell culture models that more closely represent the human respiratory epithelium and show that Hh strains with high levels of hpl expression protect epithelial cell line monolayers against adhesion and invasion by NTHi. Inhibitory activity was dependent on the level of Hpl production, which was stimulated by NTHi challenge and nasopharyngeal cell exposure. Provided these protective benefits translate to in vivo applications, Hpl-producing Hh may have probiotic utility against NTHi infections by inhibiting requisite nasopharyngeal colonisation.

Rational drug design for sore throat—An aspirin-based treatment that addresses bradykinin-induced inflammation

Often thought of as a minor health concern, sore throat or pharyngitis is an important public health issue. It is one of the most common symptoms of upper respiratory diseases including COVID-19 and is a leading cause of physician visits and antibiotic prescriptions. Despite being on the market for decades, few over the counter sore throat medications are proven to heal sore throat. In studying pharyngitis using organotypic human respiratory tissue stimulated with bradykinin, we saw an increase in prostaglandin E2 (PGE2) and interleukin-8 (IL-8) in response to bradykinin. Bradykinin is one of the first inflammatory signals for pharyngitis and it increases PGE2 in human subjects. If left unregulated, PGE2 may further increase inflammation via the COX pathway and via IL-8, a proinflammatory chemokine responsible for neutrophil infiltration and possibly thus, a cytokine storm. Acetyl salicylic acid (ASA), a non-specific COX inhibitor, was able to mitigate a bradykinin-induced increase in PGE2 in our studies. However, ASA was inflammatory above its small therapeutic window, greatly increasing levels of PGE2 and IL-8 above those seen with bradykinin stimulation alone. Similar to other systems, the respiratory epithelia maintains a delicate balance of pro-inflammatory and anti-inflammatory signals in order to keep the respiratory barrier intact. To our knowledge, this is the first study to try and elucidate the complex mechanisms involved in healing pharyngitis, an inflammatory condition of the respiratory epithelia. Biovanta™, a formula containing ASA mitigated bradykinin-induced inflammation more strongly than ASA alone in organotypic human respiratory tissues. Surprisingly, we found that many of the most common over the counter sore throat therapies exacerbate inflammation and IL-8 in organotypic human respiratory tissues, suggesting these common treatments may possibly increase the likelihood of further respiratory complications in people.Competing interest statementThis study was funded entirely by Applied Biological Laboratories, a private company that owns the Biovanta™ product. Unless otherwise indicated all experiments were performed at Applied Biological Laboratories research facility located at the SUNY Downstate Biotechnology Incubator, a part of StartUP NY. All of the authors were employees of Applied Biological Laboratories at the time the experiments were performed.

Endogenous zinc nanoparticles in the rat olfactory epithelium are functionally significant

The role of zinc in neurobiology is rapidly expanding. Zinc is especially essential in olfactory neurobiology. Naturally occurring zinc nanoparticles were detected in olfactory and nasal respiratory epithelia and cilia in animals. The addition of these nanoparticles to a mixture of odorants, including ethyl butyrate, eugenol, and carvone, considerably increased the electrical responses of the olfactory sensory receptors. Studies of these nanoparticles by ransmission electron microscopy (TEM) and selected area electron diffraction revealed metal elemental crystalline zinc nanoparticles 2–4 nm in diameter. These particles did not contain oxidized zinc. The enhancement of the odorant responses induced by the endogenous zinc nanoparticles appears to be similar to the amplification produced by engineered zinc nanoparticles. Zinc nanoparticles produce no odor response but increase odor response if mixed with an odorant. These effects are dose-dependent and reversible. Some other metal nanoparticles, such as copper, silver, gold, and platinum, do not have the effects observed in the case of zinc nanoparticles. The olfactory enhancement was observed in young and mature mouse olfactory epithelium cultures, in the dissected olfactory epithelium of rodents, and in live conscious dogs. The physiological significance of the detected endogenous metal nanoparticles in an animal tissue has been demonstrated for the first time. Overall, our results may advance the understanding of the initial events in olfaction.