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. 2013 Jan 23;288(12):8053–8060. doi: 10.1074/jbc.M112.433995

FIGURE 5.

FIGURE 5.

COCH suppresses neural differentiation of mouse ESCs. A, R1 ESCs stably expressing GFP or COCH were cultured in normal KSR medium (ES) or N2B27 medium to allow neural differentiation for 5 days (ND). mRNA levels of the neural differentiation markers Sox1, Nes, and Tubb3 were determined by qRT-PCR. B, R1 ESCs stably expressing GFP or COCH were cultured in N2B27 medium for 5 days and were then subjected to anti-nestin immunostaining analysis. Scale bars: 50 μm. C, Coch knockdown R1 mouse ES cells (shCoch-1 and shCoch-2), and the control luciferase knockdown cells (shLuc) were cultured in normal KSR medium (ES) or N2B27 medium (ND) for 5 days, and then harvested for qRT-PCR analysis. D, Coch knockdown R1 mouse ESCs or control cells (shLuc) were cultured in N2B27 medium for 5 days, and then subjected to anti-nestin immunostaining analysis. Scale bars: 50 μm. E, Coch knockdown R1 mouse ESCs (shCoch-1 and shCoch-2) and control (shLuc) cells were cultured in N2B27 medium with or without BMP4 for 5 days and then harvested for qRT-PCR. BMP4 neural inhibition rate (+BMP4/−BMP4) was presented by the ratio of the corresponding mRNA levels of +BMP4 to −BMP4. Quantitation data are shown as mean ± S.E. (n = 3) (*, p < 0.05; **, p < 0.01; ***, p < 0.001).