Figure 2.
Conversion of Human Fibroblasts into a Transient Neural Precursor Stage
(A) Scheme of experimental setup for conversion into induced Schwann cells.
(B) Pretreatment with valproic acid (VPA) results in increased sphere size. Columns show mean sphere diameter ± SD at day 3 of suspension culture (n = 4).
(C) Secondary spheres at day 11 with bipolar cells growing out of spheres. Cells express neural plate markers SOX1 and NESTIN (top left). No expression of NSC marker SOX2 and of neural crest markers SNAI1, SOX10, FOXD3, and PAX3 was detected. Scale bars, 50 μm.
(D) At day 18, cells express neural crest markers SNAI1, SOX10, FOXD3, and PAX3. Scale bars, 20 μm.
(E) Flow cytometry of precursors (d18) and fibroblasts (d0) revealed downregulation of fibroblast marker CD29 and upregulation of neural crest marker CD271. Panels show quantification of mean fluorescence intensity (MFI) (n = 3).
(F–H) Nonneural differentiation of transient precursors. (F) Adipocyte formation analyzed by oil red O staining. (G) Formation of SMA+ smooth muscle cells. (H) Formation of chondrocytes analyzed by Alcian blue staining. Inset shows closeup of chondrogenic pellet. Scale bars, 50 (F), 100 (G), and 20 μm (H, inset).