Fig. 4

Differentiation of ciXEN cells into VE and their spontaneous differentiation. a The induction process of VE. b Cell morphologies after induction for 7 days (a’) and the immunofluorescence of E-cadherin (b’) in VE (bar, 100 μm (a’) and 50 μm (b’)). qPCR results for the expression of Cdh1 (c) and VE-related genes (Afp, Hnf4a, and Apoe1) (d) in VE. e ALP staining of MEFs and ciXEN cells (bar, 100 μm). f Microscopic view of the cell mass formed after hanging-drop culture for 8 days (a’), as well as the cells that emerged from the cell mass (b’) (bar, 200 μm). g Immunofluorescence of Alb (endoderm), nestin (ectoderm), and desmin (mesoderm) (bar, 50 μm). * p < 0.05. Mean ± SEM, t test, n = 3