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. 2020 Nov 5;29(3):1151–1163. doi: 10.1016/j.ymthe.2020.11.004

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© 2020 The Author(s)

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

PMC Copyright notice

Cardiac Differentiation Protocol Using Sequential Stimulation and Then Inhibition of LPAR4 in Mouse ESCs and Human iPSCs

(A) To maximize the cardiac differentiation efficiency, LPA or ODP was used as LPAR4 stimulants. LPAR4 antagonist combination (BrP-LPA and AM966) or p38 MAPK blocker was used as LPAR4 inhibitors. Mouse ESCs were differentiated into CMCs and confirmed using the cardiac lineage marker Mesp1 at cardiac differentiation day 7 and cTnT at cardiac differentiation day 10 by real-time PCR. Besides, beating CMCs were counted (displayed as a bar graph) to confirm the cardiac differentiation efficiency. (B) The human cardiac differentiation protocol is shown in a schematic figure (upper panel). The effects of ODP and the p38 MAPK blocker on the differentiation efficiency of human iPSCs into CMCs were evaluated. The cardiac lineage markers were analyzed by real-time PCR at cardiac differentiation day 17 (lower panel). Statistical analyses were performed using one-way ANOVA (Newman-Keuls). ∗∗∗p < 0.001. All experiments were conducted at least in triplicate.