Evaluation of the apoptosis activation in bone marrow fresh erythroblasts.

(A) The cytofluorimetric analysis of the GpA⁺ population revealed a consistent presence of the proenzyme constitutive CPP32 and FLICE (caspases 3 and 8) in fresh erythroblasts from both MM patients with aggressive disease (no. 16 and 17) as compared with control MGUS patient no. 5, whereas ICE was only slightly represented. The measurement of these native proteases in Jurkat cells used as the control Fas⁺ cellular model confirmed their considerable expression in these cells. (B) Differential cytofluorimetric detection of CPP32-cleaved subunits in fresh erythroblasts from patients no. 16 and 5. The increment of both 12-kd and 89-kd (PARP) products in cells from patient no. 16 supported the previous activation of erythroblast apoptosis in vivo. A similar effect was detectable in Jurkat cells after their overnight incubation with the conditioned SN from myeloma cells of patient no. 16 containing high levels of Fas-L, as compared with untreated control cells (red). (C) Fluorescent pattern of erythroid apoptosis in bone marrow smears from patient no. 16. An erythroblast (top: PE positivity of glycophorin A) is juxtaposed (bottom) to a myeloma cell FITC⁺ for κ chains and is positive to the TUNEL assay (bottom) for the fluorescent condensation of the chromatin.