Gene expression and effect of demethylating treatment. Expression of EPHB4 (A), EFNB2 (B), and EFNA5 (C) genes was determined in normal BM, normal peripheral blood cells CD19⁺ (PB), primary ALL bone marrow, and leukemia cell lines. Leukemia cells were either untreated (C) or treated with DAC only, TSA only, or both (D + T) as described in “RNA extraction and real-time PCR” and “5-Aza-2′-deoxycytidine and/or trichostatin A treatment.” The relative gene expression was determined by real-time PCR assays and normalized to that of GAPDH. Methylation density (M%) for each samples and pretreatment cell lines are shown on the bottom. (D) Methylation levels of EphB4 before and after DAC treatment. Pyrosequencing was performed to determine methylation density. Error bars indicate range.