Fig. 4.

Role of lipid rafts in the maintenance of pluripotency of mouse ES cells. (A) Mouse ES cells were cultured with or without 10 mM Mβ-CD for 1 h everyday for 4 days. (B, C) Cells were transfected for 24 h with different doses of SMARTpool of caveolin-1 siRNA or nontarget control siRNA (0–50 nM), and expressions of caveolin-1 and Oct4 protein were detected. Zero represents no siRNA. (D) Cells were cultured with or without LIF for 5 and 10 days, and Oct4 protein expression levels were detected by Western blot. Each example is representative of three experiments. The graphs denote the mean ± SE of three experiments for each condition determined from densitometry relative to β-actin. (E–H) Mouse ES cells were harvested after culture in normal culture condition for 5 days, cultured with 10 mM Mβ-CD for 1 h every day for 5 days in normal culture condition, and cultured in normal culture condition for 4 days after transfection with 50 nM caveolin-1 or nontarget control siRNA for 24 h, respectively. Real-time RT-PCR quantification of Oct4, Sox2, FoxD3, and Rex1 was carried out. The data is reported as the mean ± SE of three independent experiments, each conducted in triplicate. ^*P < 0.05 versus control. ES cell, embryonic stem cell; LIF, leukemia inhibitory factor; Mβ-CD, methyl-β-cyclodextrin; ROD, relative optical density; siRNA, small interfering RNA; STAT3, signal transducer and activator of transcription 3.