Objective: The onset of effect of Thiopurines is delayed for several months. The aim of this study was to investigate immune mechanisms for this delay.
Methods: The effects of Thiopurines on human peripheral blood T-cells and on lamina propria lymphocytes were investigated for apoptosis induction by Annexin-V/PI and for cytokine secretion by intra-cellular staining and ELISA assays. To investigate Thiopurines' mechanism of effect in vivo, Balb/C mice were co-immunized with HEL/OVA antigens, and then repeatedly challenged by HEL only, while being treated by Mercaptopurine or vehicle alone for either 4 or 20 weeks. The memory response of CD4+ splenocytes towards HEL/OVA was then determined by CFSE-dilution.
Results: Thiopurines arrested the proliferation of stimulated T-cells but did not enhance the apoptosis of either resting T-cells or activated T-cells until day 5 post-stimulation. Despite the proliferation arrest, stimulated T-cells successfully differentiated into effector cells, as evidenced by their capacity for pro-inflammatory cytokine secretion, potent adhesion and cytotoxicity. Prolonged Mercaptopurine treatment of mice for 20 weeks selectively reduced the CD4+-memory response to a repeatedly encountered HEL antigen, but did not affect the T-cell memory pool to the previously exposed OVA antigen. A shorter, 4 weeks, treatment with Mercpatopurine did not inhibit the memory response to either antigen.
Conclusions: These data propose a model whereby despite Thiopurine treatment, cycle-arrested T-cells can still differentiate into potent effector cells capable of propagating tissue inflammation. However, recurrent antigen encounters result in incremental depletion of specific T-cell memory that may herald the eventual anti-inflammatory activity of Thiopurines
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