线粒体靶向小分子IR-780对顺铂耐药膀胱癌细胞的杀伤效应

Anti-tumor effect of mitochondrial targeted small molecule IR-780 on cisplatin-resistant bladder cancer cells

目的探讨线粒体靶向小分子IR-780对顺铂耐药T24膀胱癌细胞株(T24/DPP)的杀伤效应及其作用机制。方法通过顺铂反复间断刺激T24细胞,构建T24/DPP耐药细胞株,以不同浓度的IR-780处理细胞,CCK-8法检测细胞活力,划痕实验检测细胞迁移能力,荧光共聚焦确定IR-780的亚细胞器定位,流式细胞术测定细胞凋亡以及线粒体活性氧水平。对6~8周龄雄性裸鼠皮下注射T24/DPP细胞建立动物荷瘤模型,将荷瘤小鼠完全随机化分组:对照组、阿霉素组和IR-780组分别腹腔注射5 mg/kg PBS、阿霉素和IR-780,隔天1次。通过近红外活体成像以及荧光共聚焦检测IR-780的蓄积特性,并对肿瘤生长情况进行监测。结果①细胞实验结果显示,IR-780抑制T24/DPP细胞增殖与迁移,且抑制作用呈明显的剂量效应关系(P<0.05)。IR-780选择性积聚在T24/DPP细胞的线粒体中。随着IR-780浓度的增加,T24/DDP细胞线粒体活性氧的产生和细胞凋亡率均随之增加(P<0.05)。②体内实验结果显示,近红外活体成像和肿瘤组织冰冻切片显示IR-780可优先聚集于裸鼠T24/DDP皮下移植瘤,并且IR-780组小鼠肿瘤的体积及质量均明显小于对照组(P<0.05)和阿霉素组(P<0.05)。结论IR-780能显著抑制T24/DPP细胞和裸鼠皮下肿瘤的生长,且线粒体通路可能参与了IR-780诱导T24/DPP细胞发生凋亡的过程。

Objective To study the anti-tumor effect of IR-780,a mitochondrial targeting small molecule,on cisplatin-resistant T24 cell line(T24/DPP).Methods T24/DPP cell line was constructed by repeated and discontinuous stimulation of T24 cells with cisplatin.After the drug-resistant cells were treated with different concentrations of IR-780,cell viability was detected by CCK-8 assay and cell migration was measured by wound healing assay.The suborganelle localization of IR-780 was determined by confocal fluorescence microscopy with aid of mitotracker treatment.Cell apoptosis and production of mitochondrial reactive oxygen species(ROS)were determined by flow cytometry.T24/DPP cells were injected subcutaneously into 6-~8-week-old male nude mice to establish a tumor-bearing nude mouse model.The tumor-bearing mice were randomly divided into 3 groups,including control group(5 mg/kg PBS),DOX group(5 mg/kg DOX)and IR-780 group(5 mg/kg IR-780).After intraperitoneal administration of IR-780,the accumulation of IR-780 were detected by near-infrared in vivo imaging system and confocal fluorescence microscopy,while the tumor volume and weight were measured.Results①In vitro,IR-780 inhibited the proliferation and migration of T24/DPP cells in a dose-dependent manner(P<0.05).IR-780 was selectively accumulated in the mitochondria of T24/DPP cells.With the increase of IR-780 concentration,the production of mitochondrial ROS and the rate of apoptosis in T24/DDP cells were increased(P<0.05).②In vivo,near-infrared imaging and frozen sections of tumor tissue showed that IR-780 preferentially accumulated in the tumor of tumor-bearing nude mouse model.The tumor volume and weight of the IR-780 group were significantly less and lower when compared with the control group(P<0.05)and DOX group(P<0.05).Conclusion IR-780 can significantly inhibit the growth of T24/DDP cells and subcutaneous tumor xenografts in nude mice,and the mitochondrial pathway might be involved in the apoptosis of T24/DPP cells induced by IR-780.