Su1831 Determination of the Molecular Phenotype of Esophageal Mucosal Inflammation in Children With Eosinophilic Esophagitis Using a 1-Hour Esophageal String Test (EST)

Abstract

eosinophils in the esophageal mucosa. Aim: To characterize T cell responses in EoE in children. Methods: We enrolled 24 patients (mean age 14.1 years, range 10-17 years, 75% male), including 2with newly diagnosed EoE, 11 with treated EoE (3 with active inflammation .15 eos/hpf, 8 in remission ,5 eos/hpf), 9 with normal endoscopy, 1 patient with mild reflux esophagitis (,5 eos/hpf) and 1 patient with Barrett's esophagus. We obtained 4 esophageal pinch biopsies at the time of clinically indicated endoscopy. For T cell isolation the biopsies were enzymatically digested. Single cell suspensions were analyzed by flow cytometry using specific cell markers for TCRαβ, CD3+, CD4+, and CD8+ T lymphocytes to phenotype the cells. A subset of cells were stimulated with PMA/ionomycin to measure intracellular cytokine potential (TNF-α and IFN-γ) via flow cytometry. Results: We extracted an average of 2.7x105 cells per patient (range 0.4to 6.5 x105). The average number of CD3 TCRαβ+ cells was 57% (range 3.9-82.9%) with 41% (range 11.3-66%) CD4+ and 44% (range 15.6-81.3%) CD8+. There was a trend towards a higher percentage of CD8+ T cells in EoE patients (n = 13) compared to the other groups (n = 11) that included normal controls and reflux patients (48% vs. 41%), although, this was not significant. The CD8+/ CD4+ ratio was also slightly higher in the EoE group compared to the other groups (1.87 vs. 1.25), although these results were not significant. In 20/24 patients (11 EoE and 11 others), on average, 29% of the CD3+ cells, 32.8% of the CD4+ cells and 41% of the CD8+ cells produced TNF-α when stimulated with PMA/ionomycin (n = 20). The average IFNγ production (n = 20) was 19.5% of CD3+ cells, 15% of CD4+ cells and 37% of the CD8+ cells. There was no significant difference in TNFα and IFN-γ production between EoE patients and the other groups. Conclusion: We demonstrated the feasibility of isolating CD3+ TCRαβ+ T lymphocytes frompediatric esophageal biopsies and the ability to phenotype and functionally test these lymphocytes. We have observed higher CD8+ T cells in EoE patients compared to other groups and a higher CD8+/CD4+ ratio in EoE patients compared to the other groups. We also observed lower TNFα and IFN-γ production in EoE patients compared to the other groups. Further studies are needed to determine the role of these lymphocytes in the development of EoE.