MDM2-specific T cells from Ag^pos mice display defects in antigen-driven expansion.

<p>(A) CFSE-labelled splenocytes from Ag^neg and Ag^pos mice were stimulated with RMA-S coated with pMDM100 peptide (10 µM) or pSV9 peptide (10 µM) for 48 hrs, stained with CD8 and Vβ7 mAbs and subjected to CFSE profiling by flow cytometry of gated CD8⁺Vβ7⁺ T cells. Data are representative of at least three independent experiments. (B) To measure antigen-specific IL-2 production, splenocytes from Ag^neg mice and Ag^pos mice were stimulated with RMA-S cells coated with pMDM100 peptide (100 µM) or a control peptide, pSV9 (100 µM). After 72 hours, 50 µl of culture supernatant was harvested and murine IL-2 was measured in a CTLL bioassay. Data represent the mean±SD of triplicate values and are representative of two independent experiments. (C) CFSE-labelled splenocytes from Ag^neg mice were stimulated with RMA-S cells coated with pMDM100 or SV9 control peptides for 72 hrs. In the bottom panel, CFSE-labelled T cells were stimulated for 72h with pMDM100 in the presence of unlabelled splenocytes from Ag^pos mice. (D) CFSE-labelled splenocytes from Ag^pos mice were stimulated with RMA-S cells coated with pMDM100 or SV9 control peptides for 72 hrs. In the bottom panel, CFSE-labelled T cells were stimulated for 72h with pMDM100 in the presence of unlabelled splenocytes from Ag^neg mice.