Eosinophils have broad and extensive immunomodulatory capacity; recent studies have focused on the roles of distinct eosinophil subsets in specific tissue microenvironments. Ly6G is a GPI‐linked leukocyte surface Ag understood primarily as a marker of mouse neutrophils, although its full function is not known. Here, we show that Ly6G/Gr1, detected by mAbs 1A8 (anti‐Ly6G) and RB6‐8C5 (anti‐Gr1), is detected prominently on a significant fraction of eosinophils from mouse bone marrow and bone marrow‐derived culture, with fractions expressing this Ag increasing in IL‐5‐enriched microenvironments. Among our findings, we identified SiglecF+Gr1+ eosinophils in bone marrow from naïve, allergen‐challenged and IL‐5 transgenic mice; SiglecF+Gr1+ eosinophils were also prominent ex vivo in bone marrow‐derived eosinophils (bmEos) in IL‐5‐enriched culture. Reducing the IL‐5 concentration 20‐fold had no impact on the rate of generation of SiglecF+ bmEos but did result in a marked increase in the Gr1− fraction (from 17.4 ± 2% to 30 ± 2.3%, ***P < 0.005). Reducing the IL‐5 concentration also enhanced chemotaxis; SiglecF+Gr1− bmEos were considerably more responsive to eotaxin‐1 than were their SiglecF+Gr1+ counterparts. These results suggest that (i) IL‐5 regulates the expression of Ly6G/Gr1, either directly or indirectly, in cells of the eosinophil lineage, (ii) eosinophils generated in response to high concentrations of IL‐5 can be distinguished from those generated under homeostatic conditions by expression of the Ly6G/Gr1 cell surface Ag, and (iii) expression of Ly6G/Gr1 may have an impact on function, directly or indirectly, including the potential to undergo chemotaxis in response to eotaxin‐1.
Virus-induced inflammation plays a critical role in determining the clinical outcome of an acute respiratory virus infection. We have shown previously that the administration of immunobiotic Lactobacillus plantarum (Lp) directly to the respiratory tract prevents lethal inflammatory responses to subsequent infection with a mouse respiratory virus pathogen. While Lp-mediated protective responses involve non-redundant contributions of both Toll-like receptor 2 (TLR2) and NOD2, the cellular basis of these findings remains unclear. Here, we address the impact of Lp and its capacity to suppress inflammation in virus-infected respiratory epithelial cells in two cell culture models. We found that both MLE-12 cells and polarized mouse tracheal epithelial cells (mTECs) were susceptible to infection with Influenza A and released proinflammatory cytokines, including CCL2, CCL5, CXCL1, and CXCL10, in response to replicating virus. MLE-12 cells express NOD2 (81 ± 6.3%) and TLR2 (19 ± 4%), respond to Lp, and are TLR2-specific, but not NOD2-specific, biochemical agonists. By contrast, we found that mTECs express NOD2 (81 ± 17%) but minimal TLR2 (0.93 ± 0.58%); nonetheless, mTECs respond to Lp and the TLR2 agonist, Pam2CSK4, but not NOD2 agonists or the bifunctional TLR2-NOD2 agonist, CL-429. Although MLE-12 cells and mTECS were both activated by Lp, little to no cytokine suppression was observed in response to Lp followed by virus infection via a protocol that replicated experimental conditions that were effective in vivo. Further study and a more complex approach may be required to reveal critical factors that suppress virus-induced inflammatory responses.
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