Effect of glucose regulated protein 78 on autophagy and apoptosis in ovarian epithelial carcinoma
10.3760/cma.j.issn.0529-567x.2015.11.011
- VernacularTitle:葡萄糖调节蛋白78基因在卵巢上皮性癌细胞自噬与凋亡中的作用
- Author:
Min LI
;
Jing TIAN
;
Quanxin QU
- Publication Type:Journal Article
- Keywords:
Ovarian neoplasms;
Heat-shock proteins;
Autophagy;
Apoptosis;
Cisplatin
- From:
Chinese Journal of Obstetrics and Gynecology
2015;50(11):848-853
- CountryChina
- Language:Chinese
-
Abstract:
Objective To explore the effect and mechanism of glucose regulated protein 78 (GRP78) on autophagy and apoptosis in ovarian carcinoma, and to investigate the influence on the growth and sensitivity to cisplatin on the ovarian cancer cells.Methods The human ovarian cancer cell line SKOV3 were treated by the GRP78 regulator BAPTA-AM and A23187, which were used to decrease or increase the expression levels of GRP78, respectively.The experiment were divided into three groups.Cells in the group of BAPTA-AM were treated by BAPTA-AM at the final concerntration of 40 μ mol/L for 1 hour.Cells in the group of A23187 were treated by A23187 at the final concerntration of 4 μmol/L for 24 hours.While, cells in the control group were treated by culture medium without any GRP78 regulator for 24 hours.The expressions of GRP78, beclin1, Bcl-2 and CHOP mRNA and protein were detected by reverse transcription (RT)-PCR and western blot.The autophagy levels was observed by green fluorescent protein-microtubule-associated protein 1 light chain 3-Ⅱ (GFP-LC3-Ⅱ) fluorescence staining.The flow cytometry was used to analyse the apoptosis rates of cells.The effect on cell growth and the sensitivity to cisplatin of SKOV3 were accessed by methyl thiazolyl tetrazolium (MTT).Results (1)The mRNA expressions of GRP78, beclin1, Bcl-2 and CHOP in the group of BAPTA-AM were 0.583±0.025, 0.860± 0.055, 0.714±0.032 and 0.811±0.004, respectively.The mRNA expressions of GRP78, beclin1, Bcl-2 and CHOP in the group of A23187 were 0.840± 0.044, 0.654 ± 0.065, 0.908 ± 0.047 and 0.620 ± 0.062, respectively.The mRNA expressions of GRP78, beclin1, Bcl-2 and CHOP in the control group were 0.687± 0.032, 0.772 ±0.029, 0.845 ±0.018, 0.712 ± 0.077, respectively.While the protein expressions of GRP78, beclin 1, Bcl-2 and CHOP in the group of BAPTA-AM were 0.423±0.035, 0.952±0.022, 0.385±0.032, 0.681± 0.095, respectively.The protein expressions of GRP78, beclin1, Bcl-2 and CHOP in the group of A23187 were 0.743 ±0.032, 0.638±0.025, 0.596±0.029, 0.431 ±0.095, respectively.The protein expressions of GRP78, beclin1, Bcl-2 and CHOP in the control group were 0.617±0.031, 0.789±0.083, 0.492±0.036, 0.531 ± 0.003, respectively.The mRNA and protein expressions of beclin and CHOP in the group of BAPTA-AM were both higher than those in the control group (P＜0.05).While, the mRNA and protein expressions of beclin and CHOP in the group of A23187 were both lower than those in the control group (P＜ 0.05).(2) The autophagy fluorescence of SKOV3 in the group of BAPTA-AM, A23187 and the control group were 706±117, 473±128, 595± 126, respectively, in which there were significant differences among three groups (P＜0.05).(3) The apoptosis rate of SKOV3 in the group of BAPTA-AM was (27.4±2.2)％, which was higher than that in the control group [(19.6± 1.4)％, P＜0.05].The apoptosis rate of SKOV3 in the group of A23187 was (12.2± 1.9)％, which was lower than that in the control group (P＜0.05).(4) The comparison of the sensitivity to cisplatin in 3 groups of SKOV3.The 50％ inhibition concentration (IC50) of SKOV3 to cisplatin was (3.02±0.62) mg/L.After treated by BAPTA-AM + cisplatin, the IC50 was (2.00±0.17) mg/L and the sensitivity of SKOV3 to cisplatin was increased by 33.8％, and there was significant difference (P＜0.05), compared with the control group.And after treated by A23187 + cisplatin, the IC50 was (4.91±2.52) mg/L and the sensitivity of SKOV3 to cisplatin was decreasd by 62.6％;and there was significant difference (P＜0.05), compared with the control group.Conclusion GRP78 could regulate autophagy and apoptosis of ovarian cancer cells by regulating the expressions of beclin1, Bcl-2 and CHOP, thereby affecting the sensitivity to cisplatin in ovarian carcinoma, which may be a new method for the treatment and improvement of the sensitivity to cisplatin in ovarian carcinoma.
