Journal: Stem Cell Research & Therapy
Article Title: Exosomes secreted by human-induced pluripotent stem cell-derived mesenchymal stem cells attenuate limb ischemia by promoting angiogenesis in mice
Figure Lengend Snippet: Efficient differentiation of mesenchymal stem cells (MSCs) from human induced pluripotent stem cells (iPSCs) and characterization of iMSCs-Exo. (A) Phase-contrast image of long-term cultured iPSC clone before differentiation (i), intermediate phase of induced iPSCs (ii), and iMSCs (iii). (B) Quantitative reverse-transcriptase polymerase chain reaction analysis of the expression level of pluripotent genes (Nanog, Oct4, and Msx1) in iPSCs during differentiation (i) and in iPSCs and iMSCs (ii). (C) Flow cytometric analysis of mesenchymal positive markers, such as CD29, CD44, CD73, CD90, CD105, and CD146, and negative markers, such as CD34, CD45, CD133, and HLA-DR. Black histograms represent the isotype controls, and the red solid peak represents the marker indicated. (D) Multi-differentiation potential of iMSCs. (i) Alizarin Red staining of osteogenic mineralization (day 14). (ii) Oil Red O staining of small lipid droplets (day 21). (iii) Toluidine Blue staining of cartilaginous extracellular matrix (day 21). (E) Morphology of iMSCs-Exo under transmission electron microscopy. (F) Western blotting analysis of exosomal CD63, CD81, and CD9 protein in iMSCs and iMSCs-Exo. The culture medium served as the control. GAPDH, glyceraldehyde 3-phosphate dehydrogenase; iMSC, induced pluripotent stem cell-derived mesenchymal stem cell; iMSCs-Exo, exosomes derived from induced pluripotent stem cell-derived mesenchymal stem cells.
Article Snippet: The following human primers were used: Nanog, Oct4, Msx1, hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor-A (VEGFA), VEGFB, placental growth factor (PGF), basic fibroblast growth factor (bFGF), transforming growth factor beta 1 (TGFB1), Angiogenin (Angio), VEGF receptor 2 (kinase insert domain receptor, or KDR), bFGF receptor (bFGFR), and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (all from Sangon Biotech, Shanghai, China).
Techniques: Cell Culture, Polymerase Chain Reaction, Expressing, Flow Cytometry, Marker, Staining, Transmission Assay, Electron Microscopy, Western Blot, Derivative Assay