The Potential Role of CD4+CD25int T-Cells in Inflammatory Diseases Refractory to Treatment With Glucocorticoids

Abstract

Purpose:: 30% of patients with posterior segment intraocular inflammation (uveitis) are refractory to treatment with moderate doses of steroids. Murine studies have shown that CD4⁺CD25⁺cells are resistant to glucocorticoid (GC) induced apoptosis and we hypothesise that this T-cell subset is responsible for perpetuating disease in steroid-resistant patients. As clinical steroid resistance is easily defined in ulcerative colitis, we wished to define with this paradigm whether steroid-resistant patients have a subpopulation of CD4⁺ T-cells which continue to proliferate despite exposure to GCs, and to compare the effect of GCs on human CD4⁺CD25^- and CD4⁺CD25^int T-cells.

Methods:: PBMCs from 16 healthy human volunteers and 7 patients with ‘steroid-resistant’ ulcerative colitis were cultured with CD3 / CD28 in the presence of supratherapeutic concentrations of dexamethasone (Dex). CD3⁺CD25^- and CD3⁺CD25^int T-cells, flow cytometrically sorted from PBMCs, were also stimulated and exposed to increasing concentrations of Dex. The proliferative response of CD4⁺ T cell subpopulations after 5 days culture was quantified by CFSE dilution.

Results:: (i.) CD3⁺ CD4⁺ cells from patients with steroid-resistant disease have a characteristic CFSE profile, with a clearly defined proliferating subpopulation (57.1%) despite high concentrations of Dex. The proportion of proliferating cells is much reduced in normal volunteers (19.0%; p=0.0002). (ii.) CD4⁺ lymphocytes from CD3⁺CD25^int cell cultures proliferate well in the presence of supra-therapeutic concentrations of Dex (10^-6M), while CD25^- cell proliferation is fully suppressed at moderate Dex concentrations (10^-7M)

Conclusions:: Human CD4⁺CD25^int cells are resistant to the anti-proliferative effects of GCs, and patients with clinically defined steroid-resistant disease have a distinct population of steroid-resistant CD3⁺CD4⁺ T-cells. This suggests that CD4⁺CD25^int cells may be responsible for perpetuating steroid-resistant inflammatory diseases and represent a potential target for therapeutic intervention. Our flow-cytometric assay also has potential clinical application as a predictive test of treatment response for individual patients.

Keywords: immunomodulation/immunoregulation • corticosteroids • flow cytometry