[Mechanism underlying 2-methoxyestradiol inducing apoptosis of K562 cells].
The aim of this study was to investigate apoptotic effect of 2-methoxyestradiol (2-ME2) on K562 cells and its mechanism. K562 cells were treated with different concentrations of 2-ME2. MTT assay was used to examine the effect inducing growth inhibition. DNA fragmentation assay and Annexin V-FITC/PI staining were used to detect the effect of apoptosis. The change of mitochondrial transmembrane potential was analyzed by flow cytometry. The expressions of related gene mRNA and/or proteins were detected by RT-PCR and Western blot respectively. The results indicated that the 2-ME2 inhibited proliferation of K562 cells in a time- and dose-dependent manners and the concentration of 50% growth inhibition (IC(50)) was 2 micromol/L at 48 hours. 2-ME2 induced DNA ladder and significantly increased apoptosis in K562 cells when exposed to 2 micromol/L of 2-ME2 for 24, 48 and 72 hours, the result of Annexin-V/PI staining showed that rates of the apoptotic cells were 13.78%, 22.32% and 29.43% respectively, which was remarkably higher than that of control (1.78%) (p < 0.05). The FCM analysis showed that the mitochondrial transmembrane potential in K562 cells lowered after exposed to 1, 2 and 4 micromol/L of 2-ME2 for 24 hours. 2-ME2 down-regulated the expression of bcr/abl and bcl-2, up-regulated the expression of bax mRNA, and down-regulated protein expressions of bcl-2, procaspase-3, procaspase-9, PARP (116 kD) and p-Akt, and up-regulated expression of cytoplasmic Cyto-C and PARP 85 kD apoptosis-related cleavage fragment protein, but had no effect on total Akt protein in K562 cells after treated with 2 micromol/L of 2-ME2 for 24, 48 and 72 hours. It is concluded that the 2-ME2 can induce the apoptosis and inhibit the proliferation of K562 cells by increasing the ratio of bax/bcl-2, reducing the mitochondrial transmembrane potential, releasing cytochrome C to cytoplasm, initiating the mitochondrial apoptosis pathway and leading in turn to caspase-3 activation. These findings suggest that interfere PI3K/Akt signal pathway via down-regulating the expression of bcr/abl mRNA is implicated in the effect of 2-ME2 on K562 cells.
Zhang
Xue-Ya
XY
Fujian Province Institute of Hematology; Department of Hematology, Union Hospital, Fujian Medical University, Fuzhou 350001, Fujian Province, China.
Zhan
Rong
R
Huang
Hao-Bo
HB
Yang
Ting
T
D000077584:2-Methoxyestradiol; D017209:Apoptosis / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D015536:Down-Regulation; D004958:Estradiol / Q000031:analogs & derivatives* / Q000494:pharmacology; D006801:Humans; D020014:K562 Cells; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics
D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D004958:Estradiol; D000077584:2-Methoxyestradiol; D045304:Cytochromes c; D051057:Proto-Oncogene Proteins c-akt