GM-CSF-receptor signaling mediates BCR/ABL-independent NI resistance through activation of STAT-5. (A) Intracellular staining of p-STAT-5 (ic-p-STAT-5) and ic-p-CrkL in LAMA-cells before (dark histogram) and after treatment (light histogram) with NI or GM-CSF as indicated. (B) Analysis of ic-p-STAT-5 according to GM-CSFR α (CD116) expression levels; (Bi) Blue denotes high CD116 expressers; and yellow, low CD116 expressers. (Bii) ic-p-STAT-5 regulation according to high and low CD116 expression levels (blue and yellow) before (full lines) and after (dotted lines) treatment with NI or NI plus GM-CSF. (Biii) Viability of CD116high and CD116low populations of LAMA cells after exposure for 24 hours to NI with or without GM-CSF. The circles indicate viable cells according to the typical scatter characteristics of viable LAMA cells. (C) GM-CSF maintains growth and survival of LAMA-cells in the presence of high doses of IM and NI. LAMA cells (4 × 105 cells / well) were cultured in 96 well plates and exposed to a final concentration of 4μM IM or NI in the presence or absence of GM-CSF as indicated. Colonies formed after 14 to 30 days and are depicted as resistant colonies per 106 input cells at the beginning of the culture. One of 3 experiments is shown, respectively, showing very similar results.
