摘要
目的探究肿瘤来源的细胞外囊泡(TEVs)中lnc RNA AC116025.2的表达对食管癌(EC)细胞5-FU耐药的影响。方法采用差速离心法分离人正常食管上皮细胞(HEEC细胞)和EC细胞的细胞外囊泡(EVs),并用TEVs干预TE-1细胞和耐药株TE-1/5-FU。采用q RT-PCR和Westernblot检测EC细胞及其EVs中lnc RNA AC116025.2的表达,EC细胞内mi R-4496及SEMAF蛋白的表达差异。采用CCK-8实验、克隆形成实验及流式细胞实验检测TEVs或5-FU干预对EC细胞增殖和凋亡的影响。此外,还进行了双荧光素酶报告基因实验,以阐明TEVs lnc RNA AC116025.2调控mi R-4496/SEMAF信号轴对食管癌5-FU耐药的可能机制。最后,在裸鼠体内验证了TEVs lnc RNA AC116025.2调控食管癌5-FU耐药中的作用。结果EC细胞(特别是TE-1/5-FU)EVs中lnc RNA AC116025.2的表达显著升高。在EC细胞(特别是TE-1/5-FU)中AC116025.2和SEMAF表达上调、mi R-4496表达下调。5-FU+EVs-oe-AC116025.2干预后TE-1细胞的活力和克隆形成增加、凋亡减少,而5-FU+EVs-sh-AC116025.2干预后TE-1/5-FU细胞活力和克隆形成下降、细胞凋亡升高。体内外实验表明,TEVs干预降低了EC细胞中mi R-4496的表达、增加AC116025.2和SEMAF的表达,促进食管癌5-FU耐药的发展。结论TEVs AC116025.2靶向mi R-4496促进SEMAF的表达,从而促进了食管癌对5-FU耐药的发生。
AIM To explore the relationship between the expression of lncRNA AC116025.2 in tumor-derived extracellular vesicles(TEVs)and 5-FU resistance of esophageal cancer(EC)cells.METHODS Differential centrifugation was used to separate the EVs from EC cells.TEVs were used to intervene with TE-1 and drug-resistant strains of TE-1/5-FU.The qRT-PCR and Western blot were used to detect the the difference between the expression of lncRNA AC116025.2in EC cells and their EVs,also detect the difference between the expression of miR-4496 and that of SEMAF in EC cells.The effects of TEVs or 5-FU intervention on EC cells proliferation and apoptosis were detected by CCK-8 assay,clone formation assay and flow cytometry assay.In addition,dual-luciferase reporter experiments were performed to elucidate the possible mechanism by which TEVs lncRNA AC116025.2 regulated the miR-4496/SEMAF signaling axis and influences 5-FU resistance in esophageal cancer.Finally,the role of TEVs lncRNA AC116025.2 in regulating 5-FU resistance in esophageal cancer was verified in nude mice.RESULTS The expression of lncRNA AC116025.2 was significantly increased in EVs from EC cells(especially TE-1/5-FU).The expressions of AC116025.2 and SEMAF were up-regulated and the expression of miR-4496 was down-regulated in EC cells(especially TE-1/5-FU).After 5-FU and EVs-oe-AC116025.2 intervention,the viability and clonogenicity of TE-1 cells increased and apoptosis decreased,while 5-FU and EVs-sh-AC116025.2 intervention decreased the viability and clonogenicity of TE-1/5-FU cells and increased apoptosis.In vitro and vivo experiments showed that TEVs intervention reduced the expression of miR-4496 and increased the expression of lncRNA AC116025.2 and SEMAF in EC cells,promoting the development of 5-FU resistance in esophageal cancer.CONCLUSION TEVs AC116025.2 promotes the expression of SEMAF by targeting miR-4496 and promotes 5-FU resistance in esophageal cancer.
作者
赵承宽
刘苏麟
许城城
张秋珍
陈美霓
郑丹玲
张述耀
ZHAO Chengkuan;LIU Sulin;XU Chengcheng;ZHANG Qiuzhen;CHEN Meini;ZHENG Danling;ZHANG Shuyao(Guangzhou Red Cross Hospital Pharmacy Department,Guangzhou 510220,China;Shantou University Medical College,Shantou 515041,China;Medical Department,The First Affiliated Hospital of Shantou University Medical College,Shantou 515041,China)
出处
《中国临床药学杂志》
CAS
2022年第7期528-535,共8页
Chinese Journal of Clinical Pharmacy
基金
2020年广东省科技专项资金(“大专项+任务清单”项目,汕府科﹝2020﹞53号,编号200115105879476)
2018年广东省科技创新战略专项资金(“纵向协同管理方向”项目,汕府科﹝2018﹞157号,编号180916134960437)
关键词
食管癌
5-氟尿嘧啶
细胞外囊泡
耐药
esophageal cancer
5-fluorouracil
extracellular vesicles
drug resistance
