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

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。

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(1μmol/L)group,a cisplatin(16 mg/L)group,and a combined Niclosamide(1μmol/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.