Effects of 2-methoxyestradiol on the proliferation and apoptosis of B16 malignant melanoma cells
10.3760/cma.j.issn.0412-4030.2015.03.006
- VernacularTitle:2-甲氧基雌二醇对恶性黑素瘤B16细胞株增殖与凋亡的影响
- Author:
Caixia HU
;
Lianmei ZHAO
;
Guoqiang ZHANG
;
Fei TIAN
;
Wenqing WANG
;
Shunqiang GAO
- Publication Type:Journal Article
- Keywords:
Estradiol;
Melanoma,experimental;
Cell line,tumor;
Cell proliferation;
Apoptosis;
Genes,myc;
Gadd45 gene
- From:
Chinese Journal of Dermatology
2015;48(3):166-170
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the effects of 2-methoxyestradiol (2-ME) on the proliferation and apoptosis of a mouse malignant melanoma cell line B16,and to explore their mechanism.Methods B16 cells were cultured in vitro,and divided into a negative control group receiving no treatment and several intervention groups treated with 2-ME at final concentrations of 5,10,20,40 mmol/L,respectively.After different durations of treatment,inverted phase-contrast microscopy was conducted to observe the morphologic change of B16 cells,sulforhodamine B (SRB) assay to evaluate proliferative activity and to draw growth curve of B16 cells according to the absorbance value at 490 nm,flow cytometry to detect cell cycle and apoptosis,and reverse transcription PCR and real-time PCR were performed to measure the expressions of the apoptosis-inducing gene gadd45b and proto-oncogene c-myc.Results As repeated measures analysis of variance showed,there were significant differences in the inhibitory effect on B16 cell proliferation among different concentrations (5,10,20,40 mmol/L) and different treatment durations (24,48,72 hours) of 2-ME (F =1170.94,1843.04,respectively,both P ＜ 0.01),and there was a significant interaction effect between these concentrations and treatment durations (F =272.79,P ＜ 0.01).After 48-hour treatment with 2-ME at 10,20 and 40 mmol/L,the apoptosis rate of B16 cells was increased to (4.13 ± 1.12)％,(11.25 ± 2.380)％ and (19.46 ± 2.9)％ respectively,compared to (0.23 ± 0.5)％ in the negative control group (all P＜ 0.01); the proportion of B16 cells in G0/G1 phase was increased to (59.5 ± 5.6)％,(63.4 ± 8.2)％ and (70.8 ± 4.4)％ respectively,compared to (44.1 ± 3.4)％ in the negative control group.There was a significant difference in the proportion of B16 cells in G0/G1 phase among the negative control group and intervention groups (F =13.56,P ＜ 0.05).Moreover,the mRNA expression of gadd45b was significantly enhanced after 24-hour treatment with 2-ME at concentrations of 20 and 40 mmol/L (both P＜ 0.01),while that of c-myc was significantly weakened after treatment with 2-ME at 10,20 and 40 mmol/L (all ＜ 0.05) compared with the negative control group.Conclusion 2-ME can inhibit the proliferation of B16 cells in vitro,upregulate the expression of gadd45b gene and downregulate the expression of C-myc gene.
