Induction of differentiation and apoptosis by ATRA and ATO in NB4 cells. NB4 cells were cultured in medium in the presence or absence of ATRA (10−6 M), ATO (10−6 M), or their combination, for 0, 4, 8, 24, 48, 72, and 96 hours. (A) May-Grünwald-Giemsa staining was performed on cytospins after the indicated treatments for 96 hours. ATRA-treated cells showed typical differentiation features, such as wider cytoplasm and more lobulated nuclei. ATO-treated cells showed evident signs of apoptosis, including fragmentation of the nuclei and formation of apoptotic bodies. Original magnification ×400. (B) After 72 hours of ATRA and/or ATO treatment, CD11b expression was analyzed by flow cytometry to determine differentiation. Mean fluorescence intensity (MFI) of the life-gated cells is plotted. (C) Annexin V and propidium iodide (PI) staining was performed to determine apoptosis after 72 hours of ATRA and/or ATO treatment. Analysis was performed on the ungated cells. The percentages of negative, single-positive, and double-positive cells are depicted.
