Fig. 3

DiR labeling does not affect ECFCs functions. A Proliferation assays. Representative images of Ki67 + (red) unlabeled (control) and DiR-labeled ECFCs (DiR-ECFCs) (left). Percentage of Ki67 + cells of unlabeled and D- ECFCs (right). B Apoptosis assay. Representative dot-plots with Annexin V (AV) and propidium iodide (PI) from flow cytometry (top). Percentage of ECFCs and D-ECFCs in early (AV + /IP-, left) and late (AV + /IP + , right) apoptosis. C Angiogenesis assay. Representative images of matrigel tubule formation results, obtained with phase-contrast microscope (left) and NIR scanning (right). D Number of meshes quantified (left) and total tubule lengths (right) measured at 24 h, 48 h and 72 h, for unlabeled and DiR-labeled ECFCs. E Flow cytometry histograms obtained to detect DiR signal (APC +) after co-culturing 24 h DiR pre-labeled EFCFs and unlabeled Jurkats. Percentage of DiR + (APC +) cells detected for DiR-ECFCs and Jurkats after 24 h co-culture. Data were represented as the mean ± SEM indicating all independent values. *p-values < 0.05. F Representative IHC images confirming the presence of DiR labeled cells (red) within the tissue, 3 days after administration, colocalizing with human CD31 + (endothelial marker, green, left image) and also detected nearby blood vessels (α-SMA staining, green)