大黄酸衍生物4B诱导内质网应激触发顺铂耐药卵巢癌细胞凋亡的活性研究

Apoptosis of cisplatin-resistant ovarian cancer cells induced by Rhein derivative 4B via trig⁃gering endoplasmic reticulum stress

目的:探究大黄酸衍生物4B杀伤顺铂耐药卵巢癌细胞的作用及相关机制。方法:以卵巢癌细胞(SKOV3)、顺铂耐药卵巢癌细胞(SKOV3/DDP)、正常卵巢细胞(IOSE80)为研究对象,采用MTT和流式细胞术分别检测大黄酸衍生物4B处理后细胞的存活率和凋亡率,苏木精-伊红(HE)染色和ER-Tracker-Red分别检测药物对细胞形态和内质网的影响,Western blotting法检测内质网相关蛋白葡萄糖调节蛋白78(GRP78)和内质网类蛋白激酶(PERK)的表达。结果:大黄酸衍生物4B在体外可明显抑制SKOV3和SKOV3/DDP的增殖,SKOV3、SKOV3/DDP、IOSE80的半数抑制浓度(IC_(50))分别为(3.68±0.71)μmol/L、(3.54±0.37)μmol/L、(5.00±0.18)μmol/L,且呈剂量依赖性地诱导SKOV3和SKOV3/DDP细胞凋亡(P<0.05)。大黄酸衍生物4B作用后,HE染色可见SKOV3和SKOV3/DDP细胞质出现大量空泡,ER-Tracker-Red染色发现空泡来源于内质网。大黄酸衍生物4B可明显上调SKOV3、SKOV3/DDP细胞GRP78和PERK蛋白表达,而内质网应激抑制剂4-PBA可有效逆转大黄酸衍生物4B引起的细胞空泡并降低其杀伤活力(P<0.05)。结论:大黄酸衍生物4B可通过激活内质网应激诱导顺铂耐药卵巢癌细胞凋亡。

Objective:To explore the killing effect of Rhein derivative 4B on cisplatin-resistant ovarian cancer cells and its possible mechanism.Methods:Ovarian cancer cells(SKOV3),cisplatin-resistant ovarian cancer cells(SKOV3/DDP)and normal ovarian cells(IOSE80)were used as the research subjects.MTT assay and flow cytometry were used to detect the survival rate and the apoptosis of cells after different treatments.Hematoxylineosin(HE)staining and ER-Tracker-Red were used to observe the changes of cell morphology and endoplasmic reticulum.The effect of derivative 4B on the expression of endoplasmic reticulum associated protein glucose regulated protein 78(GRP78)and endoplasmic reticulum protein kinase(PERK)was detected by Western blotting.Results:Derivative 4B could significantly inhibit the proliferation of ovarian cancer and cisplatin-resistant ovarian cancer strains in vitro.The IC_(50)of SKOV3,SKOV3/DDP,and IOSE80 were(3.68±0.71)μmol/L,(3.54±0.37)μmol/L,and(5.00±0.18)μmol/L,respectively.Derivative 4B induced apoptosis in SKOV3 and SKOV3/DDP cells in a dose-dependent manner.After the cells were treated with derivative 4B,a large number of cytoplasmic vacuoles were found in SKOV3 and SKOV3/DDP cells by HE staining,and ER-Tracker-Red staining which showed that the vacuoles originated from endoplasmic reticulum.Derivative 4B could significantly up-regulate the expression of GRP78 and PERK proteins in SKOV3 and SKOV3/DDP cells,while endoplasmic reticulum stress inhibitor 4-PBA could effectively reverse the vacuoles induced by derivative 4B and reduce the cytotoxicity.Conclusion:Derivative 4B can induce apoptosis of cisplatin-resistant ovarian cancer cells by activating endoplasmic reticulum stress.