Serum- and cytokine-free EC cocultures supplemented with low doses of SCF, Flt3L, and TPO expand phenotypically marked HPCs/HSPCs and differentiated hematopoietic progenitors. Human umbilical cord blood (hCB) was enriched for CD34⁺ cells, and 5 × 10⁴ CD34⁺ cells were cocultured with or without E4⁺ECs. (A-C) Cocultured hematopoietic cells were cumulatively expanded. Day 12 was chosen for analyses because of rapid attrition of hematopoietic cells, which were cultured without feeder cells. (A) Total hematopoietic cell expansion. (B) Total CD45⁺CD34⁺ HPC expansion. (C) Total phenotypically marked HSPC expansion defined by CD45⁺ Lin⁻CD34^hi+CD45RA⁻CD49f⁺. Note that human hematopoietic cells that were cocultured with E4⁺ECs expanded significantly more compared with cultures without vascular feeder cells. (D) Representative contour plots of day 12 cumulative expansion of CD45⁺Lin⁻CD34^hi+CD45RA⁻CD49f⁺ on E4⁺ECs. (E) Representative contour plots of day 12 cumulative expansion of CD33⁺CD14⁺ and CD33⁺CD15⁺ myeloid progenitors on E4⁺ECs. (F) Representative contour plots of day 12 cumulative expansion of CD51/61⁺CD41a⁺CD42b⁺ megakaryocytic progenitor cells on E4⁺ECs. Ranges of fold expansion are listed in supplemental Table 1. Results were analyzed using Student t test, and P < .05 was considered significant. Data are mean ± SD.