Fig. 1

Impact of hyperthermia on Oct4 expression in mouse blastocysts and ESCs. a Immunostaining of Oct4, Klf4, Sox2, and Nanog in the ICMs of E3.5 blastocysts. The blastocysts were incubated for 1 h at the indicated temperature, and the nuclei were shown by DAPI staining (blue). b, c Western blot analysis of Oct4, Nanog, Sox2, Klf4, c-Myc, and Lin28 from E14 and JM83A mESCs. Lamin B and GAPDH were used as controls. C, 37 °C; H, 42 °C for 1 h; R, recovery at 37 °C for 6 h following 42 °C treatment (b). Quantification data represent the mean ± SD of three independent experiments. ∗p < 0.05, ∗∗p < 0.01 (c). d, e Western blot analysis of Oct4, Nanog, Sox2, and Klf4 from P19 cells incubated for 1 h at each indicated temperature (d). Quantification data represent the mean ± SD of three independent experiments. ∗p < 0.05, ∗∗p < 0.01 (e). f, g Western blot analysis of ectopically expressed FLAG-tagged Oct4 from HEK293T cells (f). Quantification data represent the mean ± SD of three independent experiments. ∗p < 0.05, ∗∗p < 0.01 (g). h Colony formation assays of JM8A3 mESCs treated as indicated. The colonies were visualized using methylene blue staining. i Hyperthermia produced no obvious effects on the mRNA expression of oct4, nanog, sox2, or klf4 in E14 mESCs, as revealed by real-time RT-PCR assays. Each bar represents the mean value obtained from at least three independent experiments normalized against gapdh mRNA; the S.D. is shown on top of each bar