Efeitos da tricostatina A sobre a acetilação de histonas, proliferação celular e diferenciação de células tronco embrionárias murinas / Effects of Trichostatin A on Histone Acetylation, Cell Proliferation and Differentiation of Murine Embryonic Stem Cells

ABSTRACT

Background: Embryonic stem cells are cells derived from early-stage embryos that are characterized by pluripotency and self- renewal capacity. The in vitro cultured murine embryonic stem cells can indefinitely propagate in an undifferentiated state in the presence of leukemia inhibitory factor (LIF). However, when stimulated, these cells can differentiate into cell lines derived from all three embryonic germ layers. The trichostatin A (TSA) is an epigenetic modifier agent and several studies have used the TSA to stimulate cellular differentiation. However, most of these studies only assessed one TSA concentration. Therefore, this study aimed to evaluate the effects of different TSA concentrations on histone hyperacetylation during in vitro cell differentiation of murine pluripotent embryonic stem cells, cultured with or without LIF, in the quest of to standardize their application on early cultures of embryonic stem cells. Materials, Methods & Results: Undifferentiated murine embryonic stem cells were plated in the presence of different TSA concentrations (0 nM, 15 nm, 50 nM and 100 nM) in the presence or absence of LIF. Thus, the treatments were evaluated in undifferentiated embryonic stem cells cultured in the presence of LIF (Control group 0 nM LIF + ; Group 15 nM LIF + ; Group 50 nM LIF + and Group 100 nM LIF + ), and in embryonic stem cells cultured in the absence of LIF (Control group 0 nM LIF - ; Group 15 nM LIF - ; Group 50 nM LIF - and Group 100 nM LIF - ). Treatment with TSA was performed for 24 h. After that the medium was replaced with fresh medium without TSA. Samples were collected at 0, 12, 24, 36 and 48 h after the beginning of the experiment. Three replicates were performed in each experimental group. The relative amount of Histone H3 lysine 9 acetylation was analyzed in all groups, as well as the cell proliferation in the embryonic stem cells cultured in the presence of LIF.