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. 2014 Sep 16;5(20):9889–9900. doi: 10.18632/oncotarget.2475

Figure 3. The selective Chk1 inhibition sensitizes CRC cells to 5-FU induced apoptosis.

Figure 3

A, SW480 cells treated with 5-FU in presence of 500 nM of MK-8776 or 500 nM of IC-83 Chk1 inhibitors. The condensed nuclei stained with Hoechst were considered apoptotic. At least 400 cells were counted per sample in three separate experiments and represented in the bar graph. B, 5-FU-arrested SW480 cells were treated with 500 nM of MK-8776 or 10 uM of Chk2 inhibitor (Chk2i). Cells were scored for nuclear condensation. Proportion of apoptotic nuclei was represented in the bar graph. C, Detection of different forms of Chk1 phosphorylation: Ser296 (S296-P), Ser317 (S317-P) and Ser345 (S345-P) and gamma-H2AX by immunoblotting of SW480 cells treated with 500 nM of MK-8776 and 50 μM of 5-FU. D, siRNA control or Chk1 siRNA were transfected in SW480 cells and treated with 50 μM of 5-FU. Cells were analyzed and nuclear condensation was detected by immunofluorescence. Proportion of apoptotic nuclei was represented in the bar graph. E, Apoptotic nuclei were scored in cells treated with a sub-optimal 5-FU concentration of 2.5 μM in combination with MK-8776. At least 200 cells were scored per sample from three different experiments. Proportion of apoptotic nuclei was represented in the bar graph.