Figure 4.
Figure 4. Overexpression of FADDwt and decreased ES-MDS erythroid precursor growth. (A) Lentivirus constructs. The FADDwt coding sequence was inserted into the TRIP-ΔU3-EF1α IRES EGFP vector. cPPT-CTS indicates the central polypurine tract-central termination sequence. The TRIP-ΔU3-EF1α/EGFP vector was used as control. (B) Transduction efficiency measured as the percentage of EGFP+ cells ± SEM (**P < .001) in EGFP-transduced (□) or FADDwt-transduced (▪) erythroid precursors from ES-MDS (n = 10), AS-MDS (n = 5), or control (n = 5) subjects at day 7 and 14. (C) Membrane Fas in EGFP-transduced (□) or FADDwt-transduced (▪) cells on day 7 and day 14. Results are expressed as means ± SEM of the ratio of fluorescence intensity (RFI) between anti-Fas monoclonal antibody and isotypic control (*P < .05). (D) Death of erythroid precursors transduced with control EGFP (□) versus FADDwt (▪) containing vectors. Results are expressed as mean percentages of annexin V+ cells in day-7 and day-14 cultures (*P = .01). (E) Growth curves of ES-MDS (n = 10), AS-MDS (n = 5), or control (n = 5) erythroid precursors transduced with EGFP (□) or FADDwt (▴) vectors from day 0 to day 15. Results are expressed as mean cumulative cell numbers (× 104) ± SEM.

Overexpression of FADDwt and decreased ES-MDS erythroid precursor growth. (A) Lentivirus constructs. The FADDwt coding sequence was inserted into the TRIP-ΔU3-EF1α IRES EGFP vector. cPPT-CTS indicates the central polypurine tract-central termination sequence. The TRIP-ΔU3-EF1α/EGFP vector was used as control. (B) Transduction efficiency measured as the percentage of EGFP+ cells ± SEM (**P < .001) in EGFP-transduced (□) or FADDwt-transduced (▪) erythroid precursors from ES-MDS (n = 10), AS-MDS (n = 5), or control (n = 5) subjects at day 7 and 14. (C) Membrane Fas in EGFP-transduced (□) or FADDwt-transduced (▪) cells on day 7 and day 14. Results are expressed as means ± SEM of the ratio of fluorescence intensity (RFI) between anti-Fas monoclonal antibody and isotypic control (*P < .05). (D) Death of erythroid precursors transduced with control EGFP (□) versus FADDwt (▪) containing vectors. Results are expressed as mean percentages of annexin V+ cells in day-7 and day-14 cultures (*P = .01). (E) Growth curves of ES-MDS (n = 10), AS-MDS (n = 5), or control (n = 5) erythroid precursors transduced with EGFP (□) or FADDwt (▴) vectors from day 0 to day 15. Results are expressed as mean cumulative cell numbers (× 104) ± SEM.

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