Figure 1.

Human ECs support robust expansion of hematopoietic stem/progenitor cells. (A): Kinetics of expansion of SS and granulocyte colony‐stimulating factor/stem cell factor‐primed BM‐derived CD34⁺CD38⁻ cells after coculture with cytokines with or without human ECs (n = 3 naïve donors per condition). (B): Analysis of hematopoietic colony‐forming cell (CFC) potential of BM‐derived CD34⁺ cells after 7‐day expansion with cytokines with or without EC. Right: Images of CFU‐M (top) and BFU‐E (bottom) colonies. (C): Phase contrast (left) and fluorescent (middle) images of P140K‐MGMT‐GFP transduced macaque SS BM CD34⁺ cells and EC after 7 days of coculture. Right: Flow cytometry analysis of GFP and CD34 coexpression in gene‐modified CD34⁺ cells after EC expansion. (D): Summary of P140K‐MGMT‐GFP lentivirus‐transduced SS BM CD34⁺ cell expansion by flow cytometry analysis for detection of CD34⁺CD38⁻ and LT‐HSPC phenotype (CD34⁺CD49f⁺Thy1⁺CD38⁻CD45RA⁻; n = 3 naïve donors). (E): Microscopy showing morphology of Wright‐stained cytospin samples of gene‐modified CD34⁺ hematopoietic cells after coculture with cytokines with or without EC. (F): CFC analysis showing frequency and morphology of CFUs generated from gene‐modified CD34⁺ cells after expansion with cytokines with or without EC. Data are shown as the mean from the three experiments (donors) ± SD. CFC assays were conducted with three macaque donors and three biologic replicates per donor. Total CFUs are expressed per 10⁵ cells plated in MethoCult (StemCell Technologies). The colony types included BFU‐E, CFU‐M, CFU‐GM, and CFU‐GEMM. Statistical analysis used the Student t test: ∗, p < .05; ∗∗, p < .005. Abbreviations: BM, bone marrow; BFU‐E, burst‐forming unit‐erythroid; CFU, colony‐forming unit; CFU‐E, colony‐forming unit‐erythroid burst; CFU‐GEMM, colony‐forming unit‐granulocyte‐erythroid‐monocyte‐macrophage; CFU‐GM, colony‐forming unit‐granulocyte‐macrophage; CFU‐M, colony‐forming unit‐macrophage; EC, endothelial cell; GFP = green fluorescent protein; LT‐HSPC, long‐term hematopoietic and progenitor stem cell; SS, steady‐state.