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

. 2019 Dec 18;9:19389. doi: 10.1038/s41598-019-55620-x

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

© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

PMC Copyright notice

The Gi(I/M)Wi hiPSC-CM differentiation protocol is highly efficient for hiPSCs of different origins and passages. (a) The differentiation of hiPSCs into CMs progresses through stages of mesendoderm, mesoderm, and cardiac mesoderm/CPC specification, and each stage is associated with the expression of specific sets of genetic markers. (b) The GiWi and Gi(I/M)Wi protocols are illustrated schematically, including the time points and concentrations of CHIR and IWR1 treatment. In the Gi(I/M)Wi protocol, the cells were differentiated with the following initial (I) and maintenance (M) CHIR doses: Gi(10/0), I = 10 μM and M = 0 μM; Gi(10/1), I = 10 μM and M = 1 μM; Gi(10/2), I = 10 μM and M = 2 μM; Gi(10/3), I = 10 μM and M = 3 μM; Gi(10/4), I = 10 μM and M = 4 μM; Gi(10/5), I = 10 μM and M = 5 μM. (c) Twelve days after differentiation was initiated, expression of the CM marker cTnT was evaluated in cells from all six Gi(I/M)Wi treatment groups via flow cytometry; then, (d) the proportion of cells that expressed cTnT was calculated and presented as a percentage of the total number of cells (n = 3 different batches of differentiated cells). *P < 0.05 vs. Gi(10/0); ^#P < 0.05 vs. Gi(10/1); ^†P < 0.05 vs. Gi(10/2).(e) hiPSCs reprogrammed from cardiac (hciPSCs) or dermal (hdiPSCs) fibroblasts were differentiated via the Gi(I/M)Wi protocol with CHIR initiation and maintenance doses of 10 μM and 2 μM respectively. Fourteen days after differentiation was initiated, the cells were immunofluorescently stained and imaged for the expression of cTnT, MYL2, α-actinin, and Cx43 (bars = 50 μm) and also verified by transmission electron microscopy (SM: sarcomere, Z: Z-band, Mito: mitochondrion; bars = 2 μm). (f,g) hciPSCs and hdiPSCs were passaged 10 (P10) or 50 (P50) times and then differentiated via the GiWi or Gi(10/2)Wi protocols; then, cTnT expression was evaluated via flow cytometry. (f) The proportion of cells that expressed cTnT was presented as a percentage of the total number of cells (n = 10 different batches of differentiated cells), and (g) the total number of cTnT-expressing cells per well was quantified (n = 7 different batches of differentiated cells). *P < 0.05 vs GiWi.