Fig. 2: In vitro exposure of CLL cells to microenvironment-like signals results in increased MSI2 levels.

A Representative flow cytometry histogram of MSI2 expression in CD19⁺CD5⁺ cells stimulated with CD40L + IL4 (green), CpGODN+IL15 (red), or not stimulated (black). The isotype control for each condition is shown as dashed lines. MSI2 protein in HD and CLL B cells (left). Black circles represent unstimulated cells, green triangles represent cells stimulated with CD40L + IL4, and red circles represent cells stimulated with CpG-ODN + IL15. B MSI2 expression in Ki67⁺ and Ki67⁻; in EdU⁺ and EdU⁻ cells and in different phases of the cell cycle. C MSI2 protein expression in undivided and divided CLL cells. Each circle/triangle/square represents one patient. D Representative signaling pathways of MSI2 expression in CLL cells. Leukemic cells were treated with CpG-ODN + IL15 for 3 days and then incubated for 24 h with PI3K inhibitor (LY294002, 50 µM) or ERK (SCH772984, 2 µM) or for 72 h with a BTK inhibitor (PCI 32765, 1 µM). MSI2 expression represented as fold change in CLL cells from 16 patients (PI3Ki, blue squares; ERKi, green triangles, and BTKi, orange circles) (middle panel). Percentage of B cells in S, G2, and M phases of the cell cycle after incubation with CpG-ODN + IL15 without and with the kinases inhibitors (left). Statistical analysis was performed using paired t-test analysis. Shown are individual values and mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.