Fig. 6
Detection of human endothelial cells and perfused vessels in grafts 5 days post-implantation. a, b Immunofluorescent-stained paraffin sections of self-assembled (SA) and µV+SA grafts: a human embryonic stem cell-derived endothelial cells (hESC-ECs) (DsRed+, red) and perfused rat and human endothelium (Lectin+, green). White arrows in merged images denote double positive cells. Scale bar, 100 μm. b hESC-ECs (DsRed+, red) and perfused human endothelium (UEA I+, white). Yellow arrows in merged images denote double positive cells. Scale bar, 100 μm. c Quantification of vessel density (number of vessels per mm2) and average vessel size for hESC-EC vessels (DsRed+, red circles) and all perfused vessels (Lectin+, blue circles) in the grafts. N = 6 biologically independent animals for both SA and µV+SA grafts. p = 0.045 for lectin+ vessels for SA and µV+SA, p = 0.018 for DsRed+ and lectin+ for µV+SA, p > 0.05 for all others (two-tailed t test). d Quantification of vessel density and average vessel size of perfused human vessels (UEA I+, grey circles) in the grafts. N = 6 biologically independent animals for both SA and µV+SA grafts, with one animal with µV+SA graft excluded from vessel size calculation due to lack of UEA I+ vessels. p = 0.023 for vessel size for SA and µV+SA, p > 0.05 for vessel density (two-tailed t test). Representative images for a, b from six biologically independent animals containing SA grafts and six biologically independent animals containing μV+SA grafts, with similar results. Hoechst-stained nuclei, blue. Data were collected from at least three confocal images of randomly selected regions per sample and analyzed by a custom lumen identifying code for vessel density and size. Error bars, mean ± SEM. *p < 0.05 determined using two-tailed t test. UEA Ulex europaeus Agglutinin I
