EZH2 Knockdown Upregulates Expression of the Genes Involved in T-ALL Cell Diferentiation

EZH2 knockdown can upregulate the expression of the transcription factors involved in T cell differentiation. [...]EZH2 can halt the differentiation of immature lymphoblastic T cells. The abnormal proliferation of immune cells results in the infiltration of immature cells into other organs,2 like the central nervous system.3 Besides, tumoral lymphoblasts can pave the way for the development of severe infectious diseases.4 TFs (transcription factors) can substantially regulate the growth, differentiation, and maturation of hematopoietic stem cells; therefore, their dysregulation can facilitate the development of hematological malignancies, like T-ALL. T lymphocyte isolation Using negative selection, the isolation of autologous and analogous CD3+ T cells were done from PBMC (peripheral blood mononuclear cells) samples according to the protocol provided by the human Pan T cell Isolation Kit II (MiltenyiBiotec, Bergisch Gladbach, Germany). Briefly, cells were seeded in 96-well culture plates and incubated for 24 hours in a humidified 5% C02 incubator until they reached 75-80% confluency Then, they were transfected with scrambled siRNA, paclitaxel (as the negative and positive control groups), transfection reagent, pure siRNA, and various concentrations of EZH2-siRNA. 48 hours after transfection, 100 uL of MTT reagent (with the concentration of 0.5 mg/mL in phosphate-buffered saline) was added to the wells, and the incubation was done for 4 hours.