Abstract :
Objective
Myelodysplastic syndromes (MDS) are characterized by peripheral cytopenia and ineffective hematopoiesis. In adult-onset MDS and in certain inherited marrow failure syndromes, apoptosis is increased and is mediated mainly through activation of the Fas pathway. It is unclear whether the various myelodysplastic disorders share the same apoptosis pathways. I investigated apoptosis pathways in a patient with refractory cytopenia with ring sideroblasts associated with congenital 4p deletion to determine the mechanism for bone marrow failure.
Methods
Marrow cells and lymphoblast cell lines generated from peripheral blood were analyzed for apoptosis and protein expression by flow cytometry, Western blot, and confocal microscopy, either directly or after γ irradiation (15 G). Cell viability after treatment with inhibitors of specific apoptosis pathways was also determined.
Results
Compared to controls, the patientʹs marrow and lymphoblastoid cells showed significantly higher apoptosis rates and activation of caspase-3. Investigation of the mitochondrial apoptosis pathway showed a consistent pro-apoptosis profile, namely, upregulation of Bax, Bax-α, cytochrome c, and Apaf1, and low bcl-2. Differences between the patientʹs and the normal cells were further accentuated after irradiation; p53 expression was strikingly higher in the patient only after irradiation. In contrast, Fas and FADD expression on the patientʹs and the controlʹs cells were comparable. Addition of caspase 3 or caspase 9 inhibitors markedly increased patient cell viablity, but blocking anti-Fas antibody did not.
Conclusion
The ineffective hematopoiesis in this case is explained by increased apoptosis and is linked to hyperactivation of the mitochondrial cell death machinery and not to the Fas pathway, which might be secondary to an intramitochondrial defect. This information is crucial because the development of anti-apoptotic agents for the treatment of MDS may not be universally efficacious and should target the specific derangement.
