Figure 1

CD41 and CD144 during ES cell differentiation. (A) Flow cytometric analysis of stem cell differentiation. CCE murine ES cells differentiated into cystic embryoid bodies after removal of LIF (see methods). Flow cytometry was used to determine the expression of VEGF-R2, CD144, and CD41. VEGF-R2 positve cells were first detected at 2.5 days after the removal of LIF and represent approximately 11% of the cells. CD144 and CD41 are first detected at day 3.5. Both markers, CD144 and CD41, represent a subset of VEGF-R2 positive cells (4.4 and 5.5%, respectively). Only a small percentage of CD144 positive cells (1.7%) co-express CD41 at day 3.5 (bottom right panel). Isotype controls are shown in the upper right panel. (B) CD144 and CD41-expressing cells remain a subset of VEGF-R2 positive cells (5.2 and 3.8%, respectively) at day 6.5, however, at this time point, the two cell markers are expressed on different cell populations (lower right panel). (C) A schematic of ES cell differentiation demonstrating the populations of cells isolated for microarray analysis. VEGF-R2 is a marker of the hemangioblast (HB). CD41 is one of the earliest markers for the hematopoietic progenitor (HP) cells and can be detected as early as day 3.5, concurrently with the endothelial-specific marker CD144 (VE-cadherin) expressed on the angioblast (AB) at day 3.5, and also on differentiated endothelial cells (ET) at day 6.5. (D) Protein expression of CD144 (VE-cadherin) and CD41 in embryonic stem cell differentiation. Isolated proteins from developing stem cells on days 2.5 to 8.5 were separated on a 10% SDS-PAGE. Western blot analysis was performed with antibodies directed against CD144, CD41, and β-actin.