Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2017 Jun 9;8(6):1701–1713. doi: 10.1016/j.stemcr.2017.05.013

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© 2017 The Authors

This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

PMC Copyright notice

Establishment of Differentiation Protocol of iPSCs into the Lineage of Keratinocytes

(A) Schematic of the three differentiation protocols for generation of keratinocytes from iPSCs. Protocols A and B differed in the coating agents. Protocol C was performed via embryoid body (EB) formation (iPSC-EB). DKSFM, defined keratinocyte serum-free medium; RA, retinoic acid; BMP4, bone morphogenetic protein 4; VTN, vitronectin; E8, Essential 8 medium; ESM, ESC medium.

(B) β-Galactosidase staining of iPSC-KCs at the indicated time points (days 14 and 21). Cell senescence was observed at day 21 of the induction. Scale bars, 100 μm.

(C) iPSC-derived keratinocytes with different methods (protocols A, B, and C) at passage 2 were immunocytochemically stained with anti- KRT14 antibody. Homogeneous keratinocyte-like cells were stained in protocol A. Scale bars, 100 μm.

(D) The number of cells positive for KRT14. The number of KRT14-positive cells were highest in protocol A (48.08%), compared with protocol B (39.24%) and protocol C (23.62%). Data are shown as mean ± SD of the cell number from three independent experiments. KRT14, keratin 14. ^∗∗p < 0.01.

(E) The growth rate of iPSC-derived keratinocytes with different treatment. A combination of Y-27632- and EGF accelerated cell growth. EGF, epidermal growth factor; DKSFM, defined keratinocyte serum-free medium; KSFM, keratinocyte serum-free medium.

(F) Real-time qPCR analysis of KRT14 and stem cell markers, i.e., OCT3/4 and NANOG, in iPSC-KCs at passage 2. KRT14 expression increased by the addition of epidermal growth factor (EGF). Data shown are mean ± SD of the expressions from three independent experiments. ^∗∗p < 0.01.

(G) Real-time qPCR analysis of KRT14 and stem cell markers in iPSC-KCs at passage 0. Gene expression levels of OCT3/4 and NANOG increased in the presence of Y-27632, whereas the KRT14 expression level remained unchanged by the treatment of Y-27632. Data are presented as mean ± SD from three independent experiments. ^∗p < 0.05, ^∗∗p < 0.01. NS, not significant.

(H) Schematic of the final protocol design for differentiation of iPSCs into keratinocytes. iPSC-KC, iPSC-derived keratinocytes; KSFM, keratinocyte serum-free medium; DKSFM, defined keratinocyte serum-free medium; RA, retinoic acid; BMP4, bone morphogenetic protein 4.

See also Figure S2.