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. 2017 Dec 5;8(67):111943–111965. doi: 10.18632/oncotarget.22955

Figure 4. Methotrexate and Etoposide induce CAV1 promoter demethylation in colon cancer cells.

Figure 4

(A) HT29(US) and (B) DLD-1 colon cancer cells were treated with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. Genomic DNA (gDNA) was denatured for 10 min at 95°C and then immunoprecipitated using the anti-5-methylcytidine antibody (5mC). Affinity purified DNA was then evaluated using qPCR analysis, defining the enrichment levels as a percentage of the input material. Specific primers were used to analyze the CAV1 proximal promoter (CAV1 promoter) or negative (CSa region) and positive control regions (IAP region). Statistically significant differences compared with the control (Basal, black bars) are indicated ***p ≤ 0.001, **p ≤ 0.01.