Down-regulation of the nonspecific and antigen-specific T cell cytokine response in ankylosing spondylitis during treatment with infliximab

Abstract

Objective: Treatment of active ankylosing spondylitis (AS) with the monoclonal tumor necrosis factor alpha (TNF alpha) antibody infliximab is highly clinically effective. This study was undertaken to investigate the precise mechanism of action of anti-TNF alpha treatment in AS.

Methods: Cytokine expression of CD4+ and CD8+ T cells was investigated before and 6 and 12 weeks after the start of treatment in 10 patients treated with infliximab, and before and after 6 weeks of treatment and 6 weeks after placebo was switched to infliximab in 10 patients treated initially with placebo. Peripheral blood mononuclear cells (PBMCs) were stimulated for 6 hours either nonspecifically with phorbol myristate acetate (PMA)/ionomycin or antigen specifically with a pool of 46 overlapping 18-mer peptides derived from the G1 domain of aggrecan. Cells were stained for T cell surface markers CD4 and CD8 and for the intracellular cytokines interferon-gamma (IFN gamma), TNF alpha, interleukin-4 (IL-4), and IL-10. Positive cells were quantified by flow cytometry. For monocyte-derived cytokines, PBMCs were stimulated with lipopolysaccharide (LPS) for 18 hours and TNF alpha and IL-10 in the supernatant were measured by enzyme-linked immunosorbent assay.

Results: Compared with baseline, infliximab treatment induced a significant decrease at 12 weeks in the number of CD4+ and CD8+ T cells that were positive for IFN gamma and TNF alpha upon PMA/ionomycin stimulation (P = 0.005). A significant reduction had already begun to occur at 6 weeks. No change in the percent IFN gamma or TNF alpha positivity among CD4+ and CD8+ subpopulations was observed after 6 weeks in patients treated with placebo. However, when these patients began infliximab treatment after 6 weeks of receiving placebo, there was a similar significant decrease in IFN gamma and TNF alpha production by CD4+ and CD8+ T cells (P < 0.05). Furthermore, infliximab treatment induced a significant reduction in the number of IFN gamma+ and TNF alpha+ CD8+ T cells (P = 0.005 at week 6 and week 12) after antigen-specific in vitro stimulation with G1-derived peptides. Between-group analysis showed that the change in the expression of IFN gamma and TNF alpha in both CD4+ and CD8+ T cells was significantly different between the infliximab and placebo groups (P = 0.001 for all variables). There was no change in the number of IL-10+ or IL-4+ T cells during treatment. No significant change in the production of TNFalpha and IL-10 upon in vitro stimulation of PBMCs with LPS was detectable during infliximab treatment.

Conclusion: Infliximab down-regulates both IFN gamma and TNF alpha secreted by T cells but does not induce a change in cytokines produced by monocytes during 3 months of treatment. This is likely to be a relevant mechanism for the clinical efficacy of this therapy.