Human Embryonic Stem Cells Acquire Responsiveness to TRAIL upon Exposure to Cisplatin

Figure 1

Cisplatin at concentrations of 1 μM and 2 μM induces DNA damage but not excessive apoptosis in hESC. Cells were treated with cisplatin at concentrations ranging from 1 μM to10 μM for 24 hours and then either harvested and fixed (24 h) or further cultured in the absence of cisplatin for 24 hours (24 + 24 h). (a) Morphology of nontreated and cisplatin-treated hESC as observed by light microscopy (10x, bar 50 μm). Both CCTL12 and CCTL14 lines of hESC were used ( for CCTL12; for CCTL14); a representative picture is shown. (b) Graphs showing cell death incidence as determined by flow cytometric analysis after double staining with Annexin-V and propidium iodide. At least 10,000 cells were analyzed per sample. Living (Annexin-/PI-), apoptotic (Annexin+/PI-), necrotic (Annexin-/PI+), and secondary necrotic cells (Annexin+/PI+) are reported as a percentage of the total cell count. The CCTL14 line of hESC was used (). (c) The presence of double-strand breaks in DNA as visualized by 53BP1 and γH2AX foci using indirect immunofluorescence (40x, bar 20 μm). Both CCTL12 and CCTL14 lines of hESC were used (); a representative picture is shown. (d) The quantity of p53 protein in cisplatin-treated hESC as demonstrated by western blot analysis. A PVDF membrane stained with 0.1% amidoblack was used as a loading control. Both CCTL12 and CCTL14 lines of hESC were used ().

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