氯硝柳胺联合顺铂抑制肾上腺皮质癌的实验研究

Study on inhibitory effect of Niclosamide combined with cisplatin on adrenocortical carcinoma

摘要:

目的 研究氯硝柳胺和低剂量顺铂联合应用对人肾上腺皮质癌SW-13 细胞株增殖、凋亡，以及 迁移力、侵袭力的影响，并探讨其作用机制。方法 体外培养SW-13 细胞，CCK-8 法检测氯硝柳胺和不同 浓度顺铂对细胞增殖的影响。设对照组、氯硝柳胺组、顺铂组、联合用药组，分别用Annexin V/PI 和吖啶 橙/ 溴化乙锭（AO/EB）染色检测细胞凋亡率的变化，Transwell 实验和细胞划痕实验检测细胞侵袭、迁移 能力的变化，Western blot 检测B 细胞淋巴瘤因子2（Bcl-2）、半胱氨酸天冬氨酸蛋白酶3（Caspase-3）、波 形蛋白（Vimnetin）、E- 钙黏蛋白（E-cadherin）表达的变化。结果 氯硝柳胺增强顺铂对SW-13 细胞增 殖的抑制作用。Annexin V/PI 和AO/EB 染色结果表明，与联合用药组比较，氯硝柳胺、顺铂促进细胞凋亡 （P <0.05）。Transwell 和划痕实验结果表明，与联合用药组比较，氯硝柳胺、顺铂降低SW-13 细胞的侵袭能 力（P <0.05）。Western blot 检测结果表明，与联合用药组比较，氯硝柳胺、顺铂抑制Bcl-2、Vimentin 的表达， 促进Caspase-3、E-cadherin 的表达（P <0.05）。结论 氯硝柳胺能够增强低剂量顺铂对SW-13 细胞增殖的 抑制作用，促进细胞凋亡，抑制侵袭及迁移能力，可能与其抑制Bcl-2、Caspase-3、E-cadherin、Vimentin 的 表达有关。

Abstract:

Objective To study the effect of Niclosamide combined with low-dosage cisplatin on the proliferation, apoptosis, migration and invasion of human adrenocortical carcinoma SW-13 cell line and its potential mechanism. Methods SW-13 cells were cultured in vitro . CCK-8 method was used to detect the influence of Niclosamide and different concentrations of cisplatin on cell proliferation. There were four groups: a control group, a Niclosamide (1 μmol/L) group, a cisplatin (16 mg/L) group, and a combined Niclosamide (1 μmol/L) plus cisplatin (16 mg/L) group. The change of cell apoptosis was detected with Annexin V/PI staining flow cytometry and AO/EB staining method respectively. The changes of cell invasion and migration abilities were detected by Transwell assay and cell scratch assay. Western blot was used to detect the changes of expressions of Bcl-2, caspase-3, E-cadherin and vimentin. Results Niclosamide enhanced the inhibitory effect of cisplatin on proliferation of SW-13 cells (P < 0.05). The results of Annexin V/PI staining and AO/EB staining showed that the combined medicine group enhanced cell apoptosis rate more significantly than the Niclosamide group and the cisplatin group (P < 0.05). Transwell assay showed that compared with the Niclosamide and cisplatin groups, the combined medicine group decreased the invasion ability of SW-13 cells (P < 0.05), and cell scratch experiments showed that compared with the Niclosamide and cisplatin groups, the combined medicine group decreased the migration ability of SW-13 cells (P < 0.05). Western blot showed that compared with the Niclosamide and cisplatin groups, the combined medicine group more efficiently inhibited the expressions of Bcl-2 and vimentin (P < 0.05), promoted the expressions of caspase-3 and E-cadherin (P < 0.05). Conclusions Niclosamide can enhance the inhibitory effect of low-dosage cisplatin on the proliferation of SW-13 cells, promote cell apoptosis, and inhibit cell invasion and migration abilities. This is probably related to its influence on expressions of Bcl-2, caspase-3, E-cadherin and vimentin.