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Tumor necrosis factor (TNF)–mediated activation of the p55 TNF receptor negatively regulates maintenance of cycling reconstituting human hematopoietic stem cells by Ingunn Dybedal, David Bryder, Anna Fossum, Leiv S. Rusten, and Sten Eirik W. Jacobsen Blood Volume 98(6):1782-1791 September 15, 2001 ©2001 by American Society of Hematology Effects of agonistic and antagonistic anti-Fas Abs on Jurkat cells.Jurkat cells (20 000) were cultured in 100 μL RPMI 1640 with 10% FCS in the absence or presence of 1 μg/mL CH11 (agonistic anti-Fas Ab) and/or ZB4 (blocking anti-Fas Ab) at different concent... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology Effect of TNF on in vitro cellular expansion of CD34+ and CD34+CD38− cells.Cells were cultured in SF medium in KFT3 in the absence or presence of TNF (20 ng/mL). Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology The effect of TNF on growth of CD34+CD38− CB cells at the single cell level.CD34+CD38− CB cells were plated at one cell/well in SF medium supplemented with KFT3 (as described in “Materials and methods”) in the absence (open bars) or presence (black bars) of... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology Effect of TNF on recruitment of single CB CD34+CD38− cells into proliferation.Single CD34+CD38− CB cells were deposited with a single cell depositor, coupled to a FACSVantage, into wells in Terasaki plates containing 20 μL SF medium supplemented with KFT3 i... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology Ex vivo expanded CD34+CD38−cells exposed to TNF have severely compromised in vivo multilineage reconstituting activity.Multilineage human engraftment of a mouse transplanted with EEs of 5000 CD34+CD38− cells cultured for 7 days in KFT3 as well as a mouse tr... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology TNF activation negatively regulates maintenance of cycling CB CD34+CD38− LTC-ICs.The 150 freshly isolated CD34+ (A) or 25 to 75 CD34+CB38− CB cells (B), or their EEs following 5 to 7 days of culture in KFT3 in the absence or presence of TNF were evaluated f... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology TNF negatively regulates maintenance of cycling BM CD34+CD38− LTC-ICs.The 25 to 75 sorted CD34+CB38− BM cells and their EEs following 5 to 8 days of culture in KFT3 in the absence or presence of TNF were evaluated for 6-week LTC-CFC activity, as described i... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology Effect of in vivo TNF treatment on short-term engraftment of human CD34+ CB cells in NODSCID mice.Freshly isolated CD34+ CB cells were transplanted into irradiated NOD-SCID mice that subsequently received 3 injections of PBS (−TNF) or 5 μg TNF (+TNF) 24, 6... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology TNFR55 mediates the negative effect of TNF on CD34+CD38− candidate stem cells.CD34+CD38− BM (3 experiments) or CB (1 experiment) cells were expanded in SF medium supplemented with KFT3 in the absence or presence of TNF (20 ng/mL), or mutants binding specifi... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology 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 ... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology TNF does not enhance apoptosis of ex vivo expanded progenitors.CD34+CB cells cultured for 6 days in KFT3 in the absence or presence of TNF (20 ng/mL) were stained with anti-CD34–APC, annexin V–PE, and 7-AAD, as described in “Materials and methods.” Annexin ... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology TNF does not affect cell-cycle distribution of ex vivo–expanded CD34+ CB cells.Cell-cycle status of freshly isolated and cultured (in KFT3 in the absence or presence of TNF for 6 days) CD34+ CB cells stained with anti–Ki-67 and 7-AAD (as described in “Mater... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology TNF down-regulates expression of CD34 and promotes differentiation of ex vivo expanded CD34+ CB cells.The upper 3 sets of panels show CD34 and lineage (CD3, CD14, CD15, CD19, CD20, CD56, CD66b, glycophorin A) expression on CD34+CB cells (purity 99%) culture... Ingunn Dybedal et al. Blood 2001;98:1782-1791 ©2001 by American Society of Hematology
