Fig. 5

ZVI@CMC nanoparticles elicited DNMT inhibition to restore the expression of SOX17 and NRF2. A, B Colony formation assay of KYSE510 pair cells treated with ZVI@CMC. The colonies were stained on day 12 after seeding (A), and the colony number was quantified (B). C Intracellular ROS level was determined by flow cytometry analysis of DCFDA fluorescence intensity after ZVI@CMC treatment for 24 h. D Immunoblotting of DNMT1, DNMT3B, SOX17, and NRF2 in ESCC cells treated with ZVI@CMC. GAPDH was used as an internal control. E Immunofluorescence staining of β-TrCP, DNMT1, NRF2, and DAPI in KYSE510 pair cells treated with ZVI@CMC. F Methylation-specific PCR (MSP) demonstrated that ZVI@CMC could reduce the methylation of SOX17 promoter. M indicates methylated PCR products, and U indicates unmethylated PCR products. G RT-qPCR analysis showed that the mRNA expressions of NRF2 downstream genes were downregulated by ZVI@CMC treatment. β-actin was used as an internal control. Data represents mean ± s.e.m. ns: non-significant; *p < 0.05; **p < 0.01; ***p < 0.001