Angiogenic phenotype and function of hESC- and fetal liver–derived CD14⁺CD16⁺ monocytic cells. (A) Tie2 expression analysis. hESCs and fetal liver–derived CD14⁺CD16⁺ cells were costained with human Tie2 antibody, and expression was studied by cytometric analysis. Plots from one representative experience are shown. The percentage of Tie2-expressing cells is the mean value of 3 independent experiments. Tie-2 was expressed by the majority of embryonic and by virtually all fetal monocytes/macrophages. (B) Human glioma U87 cells were injected subcutaneously into nude mice alone (control) or with hESC- and fetal liver–derived CD14⁺CD16⁺ cells (n = 6 for each group). Developing CD34⁺ blood vessels were studied by immunochemistry at days 5-7. The vascular area was calculated by digital image analysis based on the quantification of mouse CD34–labeled vessels. Error bars indicate SD. Computer-assisted image analysis showed that the overall vascular area was not significantly greater in tumors coinjected with embryonic/fetal macrophages than in control tumors. (C) Detection of human cells by flow cytometric analysis. After administration of human glioma U87 cells into nude mice alone (control) or with hESC-derived and adult CD14⁺CD16⁺ cells (n = 6 for each group), at day 7 tumors were dissociated into single-cell suspensions and labeled with mouse CD31-PE and human CD45-APC. One representative experiment is shown. Results are mean values of the percentage of human CD45⁺ cells present in the tumors on the days of analysis (n = 6 for each group). *P < .05 for the statistical difference between the residual human CD45⁺ cells on the days of analysis after administration of adult and hESC-derived CD14⁺CD16⁺ cells. (D) Morphology of vessel sections in tumors coinjected with adult (left panel) or hESC CD14⁺CD16⁺ (middle and right) cells. Magnification is 200×. Note the larger vessel lumen after embryonic cell coadministration (→).