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The suspension was centrifuged at 1000 rcf for 3 min, and the cell pellet was resuspended in tissue-specific culture media, including Advanced DMEM/F-12 (Gibco, 12634028) with glutamax (Invitrogen, 35050061), B27 (Gibco, 17504044), 1.25 mM N-acetylcysteine (Macklin, N800425), 50 ng/mL Recombinant Human EGF (R&D, 236-EG-01M), 100 ng/mL Recombinant Human FGF-10 (Peprotech, 100-26-5), 100 ng/mL Recombinant Human FGF-4 (Peprotech, 100-31-25), 100 ng/mL Recombinant Human Noggin (R&D, 6057-NG), 10 M SB202190 (R&D, 1264), 250 nM CHIR99021 (Sigma, SML1046), anti-mouse CD16/32, 10 M Y-27632 (APExBIO, A3008), and 500 nM A 83-01(MCE, HY-10432)

Vail and Roepke, 2019)

The stock solution was sequentially diluted with complete DMEM to prepare drug-containing working solutions with low concentrations (6.25400 g/mL) and high concentrations (506400 g/mL)

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