Suppressor activity among CD4+,CD25++ T cells is discriminated by membrane-bound tumor necrosis factor alpha

Objective. Previous studies have shown that the suppressive capacity of CD4+,CD25++ T cells is compromised in patients with rheumatoid arthritis (RA) and restored by anti-tumor necrosis factor alpha (anti-TNF alpha) therapy. Given the lack of specific cell surface markers for human Treg cells, this study aimed to define surface markers for identifying and enriching Treg cells with enhanced regulatory ability within the CD4+,CD25++ T cell compartment and to provide additional understanding of the effects of anti-TNF alpha antibodies in humans. Methods. The expression of membrane-bound TNF alpha in human peripheral blood CD4+ T cells was analyzed by flow cytometry in healthy individuals and RA patients before and after anti-TNF alpha treatment. Membrane-bound TNF alpha-positive and TNF alpha-negative CD4+,CD25++ T cells were purified by fluorescence-activated cell sorting, and their suppressive capacity was assessed in vitro by a standard suppression assay. Results. A substantial number of CD4 +,CD25++ T cells expressed membrane-bound TNF alpha. Membrane-bound TNF alpha-positive CD4+,CD25++ T cells displayed reduced antiinflammatory cytokine production and less potent suppressor capacity, since 4 times more cells were required to achieve 50% inhibition compared with their membrane-bound TNF alpha-negative counterparts. Treatment of RA patients with TNF alpha-specific anti-bodies led to a reduction in the number of membrane-bound TNF alpha-positive CD4+,CD25++ T cells from peripheral blood. Conclusion. Our data indicate that the absence of membrane-bound TNF alpha on CD4+,CD25++ T cells can be used to characterize and enrich for Treg cells with maximal suppressor potency. Enrichment of membranebound TNF alpha-negative. CD4+,CD25+ cells in the CD4 +,CD25++ T cell compartment may contribute to restoring the compromised suppressive ability of CD4 +,CD25++ T cell populations in RA patients after anti-TNF alpha treatment.