Figure 7.

Pyrophosphate antigen stimulated γδ T cells differentiated with IL-15 upregulate surface CD107a and display cytotoxic and anti-proliferative activity against MOLT-4 leukemic cells. (A) Purified γδ T cells were differentiated with 25 ng/mL IL-15 for 4 days, stimulated with isopentenyl pyrophosphate (IPP), farnesyl-PP (FPP) or geranylgeranyl-PP (GGPP) (each at 30 μM) for 4 hours and immunofluorescently stained for surface expression of CD107a in the absence of monensin (filled histogram) vs. isotype control (open histogram). (B) MOLT-4 cells were labeled with 0.5 μM carboxyfluorescein diacetate succinimidyl ester (CFSE) and mixed with IL-15 differentiated γδ T cells (IL-15 γδ T cells) at a ratio of 1:5. The cell mixture was stained with fluorophore-conjugated antibodies against CD3 and TCRVδ2 and analyzed by flow cytometry. In a 96-well round-bottom plate, CFSE-labeled MOLT-4 cells (3 × 10⁴) were co-cultured with 1.5 × 10⁵ IL-15 differentiated γδ T cells (ratio 1:5) that had been pre-activated with pyrophosphate antigens (30 μM) for 4 h. After overnight incubation (in the absence of pyrophosphate antigens), cells were stained for CD3 and TCRVδ2 (as described above) and analyzed by flow cytometry (bottom panel). Data are representative of 3 independent experiments with 2 different donors.