The Endogenous Th17 Response in NO<inf>2</inf>-Promoted Allergic Airway Disease Is Dispensable for Airway Hyperresponsiveness and Distinct from Th17 Adoptive Transfer

Severe, glucocorticoid-resistant asthma comprises 5-7% of patients with asthma. IL-17 is a biomarker of severe asthma, and the adoptive transfer of Th17 cells in mice is sufficient to induce glucocorticoid-resistant allergic airway disease. Nitrogen dioxide (NO2) is an environmental toxin that correlates with asthma severity, exacerbation, and risk of adverse outcomes. Mice that are allergically sensitized to the antigen ovalbumin by exposure to NO2 exhibit a mixed Th2/Th17 adaptive immune response and eosinophil and neutrophil recruitment to the airway following antigen challenge, a phenotype reminiscent of severe clinical asthma. Because IL-1 receptor (IL-1R) signaling is critical in the generation of the Th17 response in vivo, we hypothesized that the IL-1R/Th17 axis contributes to pulmonary inflammation and airway hyperresponsiveness (AHR) in NO2-promoted allergic airway disease and manifests in glucocorticoid-resistant cytokine production. IL-17A neutralization at the time of antigen challenge or genetic deficiency in IL-1R resulted in decreased neutrophil recruitment to the airway following antigen challenge but did not protect against the development of AHR. Instead, IL-1R-/- mice developed exacerbated AHR compared to WT mice. Lung cells from NO2-allergically inflamed mice that were treated in vitro with dexamethasone (Dex) during antigen restimulation exhibited reduced Th17 cytokine production, whereas Th17 cytokine production by lung cells from recipient mice of in vitro Th17-polarized OTII T-cells was resistant to Dex. These results demonstrate that the IL-1R/Th17 axis does not contribute to AHR development in NO2-promoted allergic airway disease, that Th17 adoptive transfer does not necessarily reflect an endogenously-generated Th17 response, and that functions of Th17 responses are contingent on the experimental conditions in which they are generated. © 2013 Martin et al

Visualization of Murine Intranasal Dosing Efficiency Using Luminescent Francisella tularensis: Effect of Instillation Volume and Form of Anesthesia

Intranasal instillation is a widely used procedure for pneumonic delivery of drugs, vaccine candidates, or infectious agents into the respiratory tract of research mice. However, there is a paucity of published literature describing the efficiency of this delivery technique. In this report we have used the murine model of tularemia, with Francisella tularensis live vaccine strain (FTLVS) infection, to evaluate the efficiency of pneumonic delivery via intranasal dosing performed either with differing instillation volumes or different types of anesthesia. FTLVS was rendered luminescent via transformation with a reporter plasmid that constitutively expressed the Photorhabdus luminescens lux operon from a Francisella promoter. We then used an IVIS Spectrum whole animal imaging system to visualize FT dissemination at various time points following intranasal instillation. We found that instillation of FT in a dose volume of 10 µl routinely resulted in infection of the upper airways but failed to initiate infection of the pulmonary compartment. Efficient delivery of FT into the lungs via intranasal instillation required a dose volume of 50 µl or more. These studies also demonstrated that intranasal instillation was significantly more efficient for pneumonic delivery of FTLVS in mice that had been anesthetized with inhaled (isoflurane) vs. parenteral (ketamine/xylazine) anesthesia. The collective results underscore the need for researchers to consider both the dose volume and the anesthesia type when either performing pneumonic delivery via intranasal instillation, or when comparing studies that employed this technique

Semaphorin 4A Stabilizes Human Regulatory T Cell Phenotype via Plexin B1

We previously reported that neuroimmune semaphorin (Sema) 4A regulates the severity of experimental allergic asthma and increases regulatory T (Treg) cell numbers in vivo; however, the mechanisms of Sema4A action remain unknown. It was also reported that Sema4A controls murine Treg cell function and survival acting through neuropilin 1 (NRP-1) receptor. To clarify Sema4A action on human T cells, we employed T cell lines (HuT78 and HuT102), human PBMCs, and CD4+ T cells in phenotypic and functional assays. We found that HuT78 demonstrated a T effector-like phenotype (CD4+CD25lowFoxp3-), whereas HuT102 expressed a Treg-like phenotype (CD4+CD25hi Foxp3+). Neither cell line expressed NRP-1. HuT102 cells expressed Sema4A counter receptor Plexin B1, whereas HuT78 cells were Sema4A+. All human peripheral blood CD4+ T cells, including Treg cells, expressed PlexinB1 and lacked both NRP-1 and -2. However, NRP-1 and Sema4A were detected on CD3negativeCD4intermediate human monocytes. Culture of HuT cells with soluble Sema4A led to an upregulation of CD25 and Foxp3 markers on HuT102 cells. Addition of Sema4A increased the relative numbers of CD4+CD25+Foxp3+ cells in PBMCs and CD4+ T cells, which were NRP-1negative but PlexinB1+, suggesting the role of this receptor in Treg cell stability. The inclusion of anti-PlexinB1 blocking Ab in cultures before recombinant Sema4A addition significantly decreased Treg cell numbers as compared with cultures with recombinant Sema4A alone. Sema4A was as effective as TGF-\u3b2 in inducible Treg cell induction from CD4+CD25depleted cells but did not enhance Treg cell suppressive activity in vitro. These results suggest strategies for the development of new Sema4A-based therapeutic measures to combat allergic inflammatory diseases. ImmunoHorizons, 2019, 3: 71-87