Two subsets of eosinophils have been described: resident eosinophils with homeostatic functions (rEOS) in healthy subjects and in patients with nonallergic eosinophilic asthma, and inflammatory eosinophils (iEOS) in blood and lung samples from patients with allergic asthma. We explored if it would be possible to identify different subsets of eosinophils using flow cytometry and the gating strategy applied to induced sputum. We conducted an observational cross-sectional single-center study of 62 patients with persistent allergic asthma. Inflammatory cells from induced sputum samples were counted by light microscopy and flow cytometry, and cytokine levels in the supernatant were determined. Two subsets of eosinophils were defined that we call E1 (CD66b-high and CD15-high) and E2 (CD66b-low and CD15-low). Of the 62 patients, 24 were eosinophilic, 18 mixed, 10 paucigranulocytic, and 10 neutrophilic. E1 predominated over E2 in the eosinophilic and mixed patients (20.86% vs. 6.27% and 14.42% vs. 4.31%, respectively), while E1 and E2 were similar for neutrophilic and paucigranulocytic patients. E1 correlated with IL-5, fractional exhaled nitric oxide, and blood eosinophils. While eosinophil subsets have been identified for asthma in blood, we have shown that they can also be identified in induced sputum.

Background: The co-culture of rat airway myocytes with activated, antigen-specific CD4⁺ T cells induces myocyte proliferation and serves as a model of hyperplasic, T-cell-driven airway smooth muscle remodeling (Ramos-Barbon D. et al. J Clin Invest 2005;115:1580-9). Aims and objectives: We aimed to translate this model to human cells for drug testing. Methods: We activated blood memory T cells from house dust mite (HDM)-allergic subjects using D. Pteronyssinus extract on Ficoll-separated mononuclear cells. We co-cultured cell cycle-arrested human airway myocytes (Lonza, France) with the antigen-activated CD4⁺ T cells for 48 hours. The negative growth control was 0.3% fetal bovine serum (FBS) medium. We quantified myocyte proliferation by flow cytometric detection of 5-ethynyl-2 deoxyuridine (EdU) uptake by smooth muscle alpha-actin (α-SMA)⁺ cells. Results: We present preliminary data from three subjects and independent experiments. D. pteronyssinus stimulation led to CD4⁺ T-cell enrichment from baseline 52.18% (50.51%-60.11%) (median, range) to 84.95% (82.37%-86.01%) CD4⁺ cells (P=0.017). Activated CD4⁺ T-cell co-culture induced EdU uptake by 21.89% (18.15%-29.00%) α-SMA⁺ cells versus 5.83% (5.11%-6.17%) control (P=0.006). Conclusions: In HDM-allergic subjects, the activation of blood CD4⁺ memory T cells, and their induction of co-cultured human airway myocyte proliferation, show analogy with the rat. The data support the feasibility of a rat-to-human translation of the antigen-specific, CD4⁺ T-cell-induced myocyte hyperplasia model as a potential drug-testing tool. Funded by the Instituto de Salud Carlos III (FIS), Spain, & FEDER