Figure 1
Nuclear PKM2 expression in CRC cell lines correlates with their resistance to gefitinib.
(A) Representative CRC cell lines were treated with varying concentrations of gefitinib. MTS assays were conducted and IC50 values were calculated after 72 h (*P < 0.05, **P < 0.01). (B) The expression of PKM2, phosphorylation and total EGFR were detected in nuclear extracts (NE) and cytoplasmic extracts (Cyto) of three representative CRC cell lines using Western blot analysis. Lamin B or β-actin were loading controls. GAPDH in nuclear extracts was analyzed as a control showing there were no cytoplasmic protein contaminations. (C) The protein levels of nuclear PKM2 were quantified by densitometry analysis and the reading was normalized to Lamin B. The error bars represent the S.D. of the mean values of three experiments. A Student’s t-test was used to assess the significance of the differences in the nuclear PKM2 between the cells lines. (D) The association between nuclear PKM2 protein expression levels and gefitinib sensitivity of CRC cells was evaluated using Pearson analysis. The coefficient of determination (R2) in the linear regression model was calculated. (E,G) The expression of PKM2 in HT29 (E) and C2BBel (G) cells were analyzed by immunoblot of nuclear extracts (NE) and whole-cell lysates (WCL) of cells in which nuclear PKM2 was overexpressed or knocked down. (F,H) The viability was analyzed using the MTS assays in HT29 (F) and C2BBel (H) cells treated with different concentrations of gefitinib. Nuclear PKM2 was overexpressed or knocked down in HT29 (F) and C2BBel (H) cells, respectively.
