CD4+CD25IntCD45RO+ T Cells Are Refractory to Glucocorticoid Treatment and Mediate IL-17 Production

Abstract

Purpose: : Suboptimal responses to corticosteroid therapy represent a major disease burden across medical specialities. We have previously identified a subpopulation of steroid refractory (SR) CD4⁺ T cells prevalent in patients with both SR uveitis and SR ulcerative colitis, and demonstrated that the SR phenotype is restricted to CD4⁺ T cells expressing intermediate levels of the -subunit of the IL-2 receptor (CD25^int cells). In experimental models of asthma Th17 cells have also been shown to be SR. Therefore, the purpose of this study was to further refine the SR CD4⁺CD25^int cell phenotype, and in particular to evaluate their potential for generating IL-17.

Methods: : Human PBMCs from 6 healthy volunteers were FACS Vantage sorted into CD4⁺CD25^neg/intCD45RO^+/- and cultured with CD3/CD28 beads in the presence of high-dose (10^-6M) Dex as previously described. Proliferation was then quantified at day 5 by CFSE dilution. In addition, expression of Th1, Th17 and memory cell markers in each CD4⁺CD25^neg/intCD45RO^+/- subset was determined by 9 colour flow-cytometry, both pre-culture and following stimulation with PMA/ionomycin

Results: : Inhibition of proliferation by Dex, quantified as percentage suppression relative to positive control cultures, was significantly greater in CD4⁺CD25^intCD45RO^-, CD4⁺CD25^negCD45RO⁺ and CD4⁺CD25^negCD45RO^- cells than in CD4⁺CD25^intCD45RO⁺ cells (p=0.05). Pre-culture, these SR CD4⁺CD25^intCD45RO⁺ cells are unactivated (CD62L^high), 69.7% central memory (CCR7⁺,CD27⁺) and do not express IL-17 or IFNγ. However, following culture with PMA/ionomycin 76.7% of the generated IL-17⁺/ IFNγ ^- cells arise from the CD4⁺CD25^intCD45RO⁺ predominantly CCR7⁺CD27⁺ subgroup.

Conclusions: : Further to our previous data, we have further refined the phenotype of SR resistant cells to a CD4⁺CD25^intCD45RO⁺ subset. Moreover, the propensity for these cells to express IL-17, suggests that they are a potential source of pathogenic SR Th17 cells in human autoimmune diseases such as non-infectious uveitis.

Keywords: immunomodulation/immunoregulation • corticosteroids • flow cytometry