Figure 4. The AWE vectors identify different sub-populations at different times of hematopoietic differentiations.

(A) AWE-transduced H9 cells were incubated in hematopoietic differentiation media and analyzed for CD45, CD31 and CD34 expression at days 10, 15 and 22. eGFP⁺ (right) and eGFP⁻ (left) populations were first analyzed for expression of CD45 and CD31. hESCs CD31⁺CD45⁺ and CD31⁺CD45⁻ were further analyzed for expression of CD34 (top and bottom plots respectively). (B) A similar experiment as in A) was performed using pLVTHM -transduced H9 cells. Note that using the pLVTHM constitutive vectors there are no differences in eGFP expression from day 10 to day 22. (C) AWE- marked cells are enriched in CD34^dim hemogenic progenitors and hematopoietic cells. Graph representing fold enrichment of eGFP⁺ versus eGFP⁻ population in hemogenic progenitors (CD45⁻CD31⁺), hematopoietic progenitors (CD45⁺CD34⁺) and myeloid cells (CD45⁺CD33⁺). CD34^dim and CD34^bright progenitor cells were analyzed independently. Data were obtained by dividing the percentages of the different cell types in the eGFP⁺ fraction of the AWE-transduced hESCs (from Figure 4A, right plots) with the values obtained in the eGFP⁻ population (Figure 4A, left plots) and plotted as fold enrichment. Data represent average of at least five separate experiments (+/− SEM) using H9 and AND-1cells at medium stage of hematopoietic development (day 10 for H9 and day 15 for AND-1). ** P = 0.008; * P = 0.032.