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. 2019 Sep 23;9(24):7122–7139. doi: 10.7150/thno.35729

Figure 4.

Figure 4

ALDH1A1-addicted erlotinib-resistant cells evolve an elevated anti-ROS/RCS system. (A-B) Lower levels of intracellular RCS (A) and ROS (B), analyzed by LC-QqQ-MS/MS and flow cytometry, respectively, in HCC827-ER5 cells compared to their parental counterparts. MGO, methylglyoxal; 4HNE, 4-hydroxy-2-nonenal. (C-J) HCC827-ER5 cells were resistant to erlotinib-induced intracellular RCS accumulation compared to the parental cells. The RCS were detected by LC-QqQ-MS/MS. The cells were treated with 1 μM erlotinib. (K) Upregulation of RCS and ROS mitigating enzymes, GPX4 and SOD2, respectively, in HCC827-ER5 cells assayed by western blot analysis. (L) Knockdown of ALDH1A1 induced ROS accumulation, more obviously in HCC827-ER5 cells than in their parental cells. The mock effect was shown as the curve of the black solid line compared with the treatment effect shown as the red. The cells were transfected with ALDH1A1 or control siRNA for 48 h and stained with DCFH-DA for flow cytometry analysis. (M-N) Overexpression of ALDH1A1 decreased the intracellular levels of ROS (M) and RCS (N). The cells were stained with DCFH-DA and imaged and analyzed in situ using an IncuCyte living cell cytometer.