Fig. 3: Restoring MYCT1 expression in cultured human HSPCs improves ex vivo expansion and engraftment ability.
From: MYCT1 controls environmental sensing in human haematopoietic stem cells
Experimental outline (a) for evaluating the effects of MYCT1 OE on CB HSPC expansion (b) and engraftment ability before (c–e) and after (f–h) extended culture. b, Fold expansion of control or MYCT1 OE total cells, CD34+CD38– cells, total HSPCs (CD34+CD38–CD90+CD45RA–) and immunophenotypic LT-HSCs (CD34+CD38–CD90+CD45RA–EPCR+ITGA3+). n = 6 replicates from 3 independent experiments, mean ± s.e.m., ratio-paired two-tailed t-test. c–e, A total of 500 or 2,500 control or MYCT1 OE HSPCs were sorted (CD34+CD38–CD90+GFP+) and transplanted 96 h after transduction into NBSGW mice. c, Percentage of mice with human haematopoietic (≥0.1% human CD45+), multilineage (myeloid, B lymphoid, and T lymphoid or other) or human erythroid (CD71+GlyA+) engraftment. Two-tailed paired t-test. d, Percentage of total human CD45 cells in BM. Median and all individual values are shown, two-tailed Mann–Whitney test. e, Estimation of repopulating unit (RU) frequency within transplanted cells after 96 h. f–h, The progeny of control or MYCT1 OE HSPCs sorted GFP+ 72 h after transduction transplanted after 10 additional days in culture (15 days total). f, Percentage of mice with human haematopoietic, multilineage or erythroid engraftment. Two-tailed paired t-test. g, Percentage of human CD45 cells in the BM for the different doses. Median and individual values are shown. h, Estimated RU frequency within transplanted cells at day 15. In e and h, the frequency of reconstituting units and P values were calculated using ELDA41. In c–e, n = 25 mice for 500 HSPCs and n = 17 mice for 2,500 HSPCs for each group transduced with control and MYCT1 OE vectors. See Extended Data Fig. 7a and Supplementary Table 4 for mouse numbers and sex of the mice. For f–h, a total of n = 34 mice for each group transduced with control and MYCT1 OE vectors. See Extended Data Fig. 8a and Supplementary Table 5 for distribution of mice and sex of the mice. For c–h, engraftment was assessed in BM 12 weeks after transplantation. Schematic in a was created with BioRender (https://www.biorender.com).
