Expression of activation markers and CTLA-4 on anergic cells.

Anergic T cells were found to down-regulate the expression of activation markers and display an increased expression of CTLA-4. Fluorescence-activated cell sorter (FACS) analysis was performed 48 hours after initiation of a restimulation experiment (panel A). Alloantigen-specific CD4⁺ anergic T cells were generated by stimulation with allogeneic IL-10–treated DCs, and activated CD4⁺ T cells were stimulated with allogeneic mature DCs. Subsequently, the T cells were restimulated with mature DCs and stained as described in “Materials and methods.” Kinetic studies of the surface and intracellular staining of the CTLA-4 molecule were performed during primary culture and after restimulation with mature DCs as described in “Materials and methods.” CD4⁺ T cells were analyzed during primary culture (day 5) and at 24 and 48 hours after restimulation with mature DCs (panel B).