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. 2018 Oct 25;27(21):1494–1506. doi: 10.1089/scd.2018.0124

FIG. 5.

FIG. 5.

Clonogenic assays with CB or MPB-derived HSPCs with increased TGFBI or decreased TGFBI expression. (A) The number of CFU-GM and BFU-E/CFU-GEMM colonies cultured in methylcellulose medium, and the number of megakaryocyte progenitors cultured in MegaCult produced by CB-derived HSPCs with TGFBI overexpression, normalized to controls. Shown are mean ± SEM (n = 7 for CFU-GM; n = 4 for BFU-E/CFU-GEMM; n = 4 for MegaCult). Range of absolute colonies per 500 plated cells for CFU-GM: SCR = 30–100, TGFBI = 20–70, BFU-E/CFU-GEMM: SCR = 20–40, TGFBI = 15–35. (B, C) TGFBI knockdown in CD34+ cells derived from CB (n = 6) or from MPB (n = 4) decreased CFU-GM formation compared to the control. Range of absolute colonies per 500 plated cells for CB: SCR = 25–125, SH2/SH3 = 10–75 and for MPB: SCR = 20–70, SH2/SH3 = 5–45. (B) BFU-E/CFU-GEMM formation was also decreased by TGFBI knockdown in CD34+ cells isolated from CB (n = 4). Number of absolute colonies counted per 500 plated cells for CB: SCR = 20–50, SH2/SH3 = 8–15. Colonies were enumerated 12–14 days after plating, each sample was performed in duplicate. *P < 0.05, **P < 0.01, sh2/sh3 indicates that sh2 or sh3 was used in independent experiments. SCR indicates that a scrambled shRNA control was used. CFU-GM, colony-forming unit granulocyte-macrophage; BFU-E, burst-forming unit-erythrocyte; CFU-GEMM, colony-forming unit-granulocyte, erythroid, monocyte, macrophage.