The negative effect of TNF on self-renewal of candidate human HSCs is not dependent on the Fas pathway.

CD34⁺CB38⁻ CB (one experiment) or BM (2 experiments) cells were cultured for 5 days in SF medium supplemented with KFT3 in the absence or presence of TNF (20 ng/mL) with or without ZB4 (1 μg/mL). Data show the mean number of CFCs generated (total of 8-10 replicates in 3 experiments) from 50 freshly isolated (open bar) or the EEs of 50 cultured (black bars) cells. ZB4 was also added to the long-term culture by the weekly medium changes to obtain the indicated concentration throughout the 6 weeks of coculture. The overall CFC values of cells cultured in the absence of TNF varied from 8 to 345 (5 of 8 replicates > 100). For cells cultured in the presence of TNF with or without ZB4, LTC-CFC varied from 0 (14 replicates) to 30 (1 replicate). ZB4 added to long-term cultures of fresh cells seeded on day 0 did not affect CFC formation (I.D. and S.E.W.J., unpublished observation, 2000).