relative expression and promoter methylation of IGSF4 in cell lines after culture with decitabine. Graph showing IGSF4 mRNA expression in cell lines ML-2, HL-60, and MONO-MAC-1 at different time points during culture with demethylating agent decitabine (DAC). (A) Solid lines correspond to untreated conditions of cell lines ML-2 (t(6;11)) and HL-60 (AML-other), and dotted lines reflect values from treated conditions of the same cell lines. Shown is > 1000-fold up-regulation of IGSF4 expression during culture with decitabine, whereas in control conditions expression remained stable. (B) Treatment of cell line MONO-MAC-1 with decitabine. The solid line reflects values from untreated condition, and the dotted line from corresponding treated samples. A 10-fold up-regulation was found during a treatment period of 8 days. (C) Results for MS-PCR in cell lines cultured with demethylating agent decitabine for 2-9 days. Left column, results for 9M primers; right column, for 9U primers. Under control conditions ML-2 and HL-60 mainly show a methylated promoter region. Shortly after start of treatment unmethylated bands are visible, and methylated bands decrease in intensity. MONO-MAC-1 shows both bands at the start of the experiment, and the methylated band clearly weakens during treatment.