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. 2018 Jan 1;145(1):dev159905. doi: 10.1242/dev.159905

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© 2018. Published by The Company of Biologists Ltd

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed.

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Fig. 4. — Enforced Wnt inhibition promotes NkxEC fate from hPSC-derived CPCs. (A-C) hPSCs were sequentially stimulated with factors that promote cardiovascular differentiation with the Wnt inhibitor Endo-IWR1 included for 2 days (days 2-4) or for an extended period (days 2-7). Colored contour plots showing representative flow cytometry measurements of the two conditions at day 7 (B) and quantification of the CP/CM, NkxEC and nEC populations (C). (D) CP/CMs were isolated at day 7 of differentiation and cultured for an additional 5 days in different concentrations of Endo-IWR1 or CHIR99021. The percentage of CD31⁺ cells in resultant cultures was determined by flow cytometry. (E) Day 7 CP/CMs were isolated and cultured in increasing concentrations of the Wnt inhibitor XAV-939. The percentage of CD31⁺ cells was quantified after 5 days. (F,G) CP/CMs were isolated at day 7 (F) or 10 (G) of differentiation and plated at clonal density. Clones exhibiting the presence of EC derivatives after 6 days were quantified in response to control, CHIR99021 or Endo-IWR1 conditions. Error bars represent s.d. between eight (C) or six (D,E) biological replicates or (F,G) between three groups of clones with the aggregate number of clones for all three groups shown beneath the bar. *P<0.05, **P<0.01, relative to control conditions.