T6 cells undergo differentiation to macrophages and DCs in response to GM-CSF and inhibit proliferation of erythroleukemic cells in vitro. (A) T6 (5 × 104) cells were cultured for 3 days in the presence of SCF: SCF + GM-CSF and SCF + GM-CSF + IL-4. The number of viable cells was calculated using the trypan blue exclusion assay. Results from triplicate experiments (mean ± SEM) are presented as the percentage of viable cells (number of cells at day 3 to number cells at day 0). **Significant differences between the no cytokine control and cytokine-treated cells (P < .005). (B) Immunohistochemical staining of adherent T6 cells, after 3 days of incubation with SCF + GM-CSF or SCF + GM-CSF + IL-4 for CD68 (macrophage marker) and 33D1 (DC marker). Brown color represents positive staining. For negative controls, the primary antibody was omitted. Original magnification ×400. T6 cells were cultured with 100 ng/mL SCF, GM-CSF, or IL-4 for 3 days. The adherent cells from this treatment (C) or culture supernatant (D) was added to 5 × 104 CB3 cells for an additional 3 days. Viable cells were determined using the trypan blue exclusion assay. Results are reported as the mean ± SEM of 3 separate experiments. **P < .005. (B) Slides were viewed with a Leica DM LB2 microscope using a 40×/0.65NA air objective. Staining was done with Carazzi hematoxylin. Images were acquired using a Leica DFC300FX camera and Leica Application Suite software (Version 3.1.0).
