Figure 2

Recruitment of T-ALL cells by the spleen environment. (a) Single-cell suspension from the spleen or BM of normal mice was placed in the lower compartment of a transwell plate, and GFP⁺ cells were placed in the upper compartment. The GFP⁺ cells that migrated to the lower compartment were counted at the indicated time point using FACS analysis (n = 5). (b) The concentration (pg/ml) of cytokines/chemokines in BM, spleen and serum samples was determined by the MILLIPLEX® MAP Multiplex Immunoassay Kits. (c) Expression of MIP-3β in BM, spleen, thymus and liver cells from normal mice was measured by real-time PCR. (d) T-ALL cells were placed in the upper chamber of a transwell plate, and conditioned medium from normal spleen or BM cells, α-MEM medium or medium containing MIP-3β, MCP-5 or IL-1a was placed in the lower compartment. The GFP⁺ cells that migrated to the lower compartment in 4 hours were counted by FACS analysis (n = 5). (e) T-ALL cells were placed in the upper chambers, and α-MEM medium (Control), normal BM cells (BM) or spleen (Spleen) cells were placed in the lower chambers. Neutralizing antibodies against CCR7 and MIP-3β were in upper or lower chambers, respectively. The migrated GFP⁺ cells were counted at 4 hours by FACS (n = 5). All columns represent the means ± SD, and the statistical analysis was performed using one-way ANOVA with multiple comparison test (*, p < 0.05; **, p < 0.01; ***, p < 0.001).