Inhibition of alloantigen presentation by cyclosporine

Cyclosporine (CsA) was examined for its ability to inhibit alloantigen presentation by spleen cells in a primary mixed lymphocyte reaction; by gamma interferon-induced P388.D1 macrophages to an alloreactive T cell clone; and by a B cell lymphoma, BID.β to an alloreactive T cell hybridoma. Alloantigen-presenting cells were treated with CsA or its inactive analogs for 2 hr, washed extensively (four times), and added to the T cells. Using this protocol, CsA maximally inhibited al-lorecognition by the T cells at 1000 ng/ml in all three systems. An HPLC assay for CsA cell failed to detect any significant CsA carryover into the T cell assays. Supernatant transfer experiments also failed to demonstrate CsA carryover in the more sensitive T cell hybridoma assay. These transfer experiments also failed to demonstrate the generation of inhibitory factors during the assay. Northern blot analysis and a cell-surface ELISA failed to observe any decreases in MHC class II induction in/on P388.D1 cells with CsA present during the induction. Due to the lack of detectable (<10 ng/ml) CsA carryover, we hypothesize that CsA has a direct effect on the formation of stimulatory MHC class II in our alloreactive systems.