Stroma-dependent apoptosis in clonal hematopoietic precursors correlates with expression of PYCARD.

Publication Type:

Journal Article


Blood, Volume 113, Issue 3, p.649-58 (2009)


2009, APOPTOSIS, Blotting, Western, Caspase 3, Cell Communication, Cell Line, Center-Authored Paper, Coculture Techniques, Cytoskeletal Proteins, Flow Cytometry Core Facility, gene expression, Hematopoietic Stem Cells, Humans, Myelodysplastic Syndromes, Receptors, Tumor Necrosis Factor, Reverse Transcriptase Polymerase Chain Reaction, RNA Interference, Stromal Cells, TNF-Related Apoptosis-Inducing Ligand, Tumor Necrosis Factor-alpha


The role of the marrow microenvironment in the pathophysiology of myelodysplastic syndromes (MDSs) remains controversial. Using stromal/hematopoietic cell cocultures, we investigated the effects of stroma-derived signals on apoptosis sensitivity in hematopoietic precursors. The leukemia-derived cell line KG1a is resistant to proapoptotic ligands. However, when cocultured with the human stromal cell line HS5 (derived from normal marrow) and exposed to tumor necrosis factor-alpha (TNF-alpha), KG1a cells showed caspase-3 activation and induction of apoptosis. Apoptosis was contact dependent. Identical results were obtained in coculture with primary stroma. Gene-expression profiling of KG1a cells identified coculture-induced up-regulation of various genes involved in apoptosis, including PYCARD. Suppression of PYCARD expression in KG1a by miRNA interfered with apoptosis. Knockdown of the TNF receptor 1 (TNFR1) or TNFR2 in HS5 cells had no effect. However, knockdown of R1 in KG1a cells prevented TNF-alpha-induced apoptosis, while apoptosis was still induced by TNF-alpha-related apoptosis-inducing ligand. Primary CD34(+) cells from MDS marrow, when cocultured with HS5 and TNF-alpha, also underwent apoptosis. In contrast, no apoptosis was observed in CD34(+) cells from the marrow of healthy donors. These data indicate that stroma may convey not only protective effects on hematopoietic cells, but, dependent upon the milieu, may also facilitate apoptosis.