Fig. 5.

Role of lipid rafts on the proliferation of mouse ES cells. (A) Mouse ES cells were incubated with or without 5 mM Mβ-CD for 24 h and then cyclin E and cyclin D1 protein expression was detected by Western blot. (B, C) Cells were transfected for 24 h with different doses (0–50 nM) of SMARTpool of caveolin-1 siRNA or nontarget control siRNA, and cyclin E and cyclin D1 expression was detected by Western blot. Zero represents no siRNA. Each example shown is representative of three experiments. The graphs denote the mean ± SE of three experiments for each condition determined from densitometry relative to β-actin. (D) Mouse ES cells were incubated with 5 mM Mβ-CD and 50 nM caveolin-1 siRNA, harvested, and subjected to PI staining for cell cycle analysis by flow cytometry. The gates were configured manually to determine the percentage of cells in S phase based on the DNA content. The data is calculated using proliferation index [(S + G2/M)/(G0/G1 + S + G2/M)] and 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.