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. 2014 Jan 28;9(1):e86459. doi: 10.1371/journal.pone.0086459

Figure 6. Hypoxia regulates IGF1 expression through HIF1α, and the inhibition of HIF1α or IGF1R decreased CD133- and Oct4-positive GRPs under hypoxia.

Figure 6

A. HIF1α expression was suppressed in PC9 or HCC827 hypoxic GRPs by treatment with 50 µM, 100 µM, and 200 µM YC-1 in Lab-Tek chamber slides. Immunofluorescence staining for IGF1, phosphorylated IGF1R (pIGF1R), CD133, and Oct4 was then performed. The numbers of IGF1-, pIGF1R-, CD133-, and Oct4-positive cells were counted, and the ratio of these cells was calculated in five fields from each experiment. **p<0.001. B. PC9 or HCC827 cells were incubated with 1 or 2 µM gefitinib in the presence or absence of 0.01 µM, 0.1 µM, or 1 µM of the IGF1R inhibitor AEW541 under hypoxic conditions for 72 h in Lab-Tek chamber slides. Immunofluorescence staining for CD133 and Oct4 was then performed. The numbers of CD133- and Oct4-positive cells were counted, and the ratio was calculated in five fields for each experiment. **p<0.001. C. PC9 or HCC827 cells were plated in 10-cm plates and allowed to adhere for 24 h. Cells were then incubated with 1 or 2 µM gefitinib in the presence or absence of 0.01 µM, 0.1 µM, or 1 µM of AEW541 under hypoxic conditions for 18 or 11 days. The numbers of colonies were then counted.