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. 2014 Nov 20;9(11):e113262. doi: 10.1371/journal.pone.0113262

Figure 4. Ectopic expression of HDAC1 and 3 inhibits the differentiation into the mesodermal lineage in EBs.

Figure 4

(A) Bright-field images and alkaline phosphatase staining of ESCs in control, HDAC1-overexpression (HDAC1-OE), and HDAC3-overexpression (HDAC3-OE) ESCs. (B) Western blotting verification and QRT-PCR analysis of the overexpression of HDAC1 and HDAC3 in stable E14 cell lines. GAPDH was used as a loading control. (C) QRT-PCR analysis for the mRNA levels of mesoderm genes in HDAC1-OE ESCs, HDAC3-OE ESCs and control cells during EB differentiation. (D) Western blotting analysis of the Gata4 and α-SMA protein levels in HDAC3-OE ESCs and control cell lines during EB differentiation. (E) Representative immunofluorescence images for the GATA4 expression level in control, HDAC1-OE, and HDAC3-OE cells after 9 days of EB formation. Red, Gata4; blue, Hoechst 33342 for nuclei staining. Data are expressed as means ± SD. Statistical significance was assessed by two-tailed Student's t test. ***, P<0.001; **, P<0.01; *, P<0.05.