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胞外囊泡中linc-VLDLR表达与食管癌发生及耐药的关系
作者:刘亮 胡月阳 巨英超 周荣秒 
单位:050011 石家庄 河北医科大学第四医院肿瘤研究所(刘亮、周荣秒) 实验动物中心(胡月阳、巨英超) 
关键词:食管肿瘤 胞外囊泡 长链非编码RNA 多药耐药 三磷酸腺苷结合转运蛋白G2 
分类号:R735.1
出版年,卷(期):页码:2018,43(1):12-16
摘要:

[摘要] 目的 探讨胞外囊泡(EVs)linc-VLDLR的表达与食管癌的发生及耐药的关系。方法 以不同浓度阿霉素(ADM)作用于食管鳞癌Eca109细胞24h,采用MTT法检测ADMEca109细胞的半数抑制浓度(IC50)。以IC50浓度为基准设置3ADM浓度干预Eca109细胞24h,提取培养液中EVs。荧光定量RT-PCR检测EVslinc-VLDLR mRNA表达。以EVs干预Eca109细胞48h,随后以不同浓度ADM再作用于细胞24hMTT法检测IC50EVs干预Eca109细胞48h,流式细胞术检测细胞周期;荧光定量RT-PCR检测Eca109细胞中linc-VLDLR及三磷酸腺苷结合转运蛋白G2(ABCG2) mRNA表达。结果 ADM作用Eca109细胞24hIC500.44±0.02μg/ml,选取00.20.40.8μg/ml ADM浓度作用Eca109细胞24h后提取上清液中的EVs(EVs1–4)EVs4linc-VLDLR表达水平明显高于EVs1–3(P<0.01)EVs1–4干预Eca109细胞48hEVs4IC50值明显高于EVs1–3及对照组(P<0.05)。流式细胞术检测显示EVs4Eca109细胞增殖指数(PI)明显高于EVs1–3及对照组(P<0.01)EVs4干预Eca109细胞48h后,Eca109细胞中linc-VLDLRABCG2基因表达水平明显高于EVs1–3及对照组 (P<0.05)结论 linc-VLDLRABCG2基因在食管癌细胞中高表达,参与了食管癌耐药形成。耐药细胞释放的EVs可以上调食管癌细胞中ABCG2表达,调控细胞耐药性,其作用与EVs上携带的linc-VLDLR基因有关。

[Abstract] Objective To investigate the relationship between the expression of linc-VLDLR in extracellular vesicles (EVs) and the development and drug resistance of esophageal carcinoma. Methods Fifty percent of inhibitory concentration (IC50) of adriamycin (ADM) for Eca109 cells was detected by MTT assay, after the treatment of esophageal squamous cell carcinoma Eca109 cell line with di?erent concentrations of ADM for 24h. The culture medium was treated with 3 concentrations of ADM based on the IC50 for 24h for extracting EVs in Eca109 cells. linc-VLDLR mRNA expression in EVs was detected by qRT-PCR assay. IC50 of ADM for Eca109 cells intervened by EVs for 48h was detected by MTT assay. Cell cycle was detected by FCM and linc-VLDLR and ABCG2mRNA expressions in Eca109 cells were detected by qRT-PCR after the treatment of the EVs for 48h. Results IC50 of ADM acting on Eca109 cells for 24h was 0.44±0.02μg/ml, so ADM concentrations of 0.2, 0.4, 0.8μg/ml were choosed in the following studies. EVs were extracted from the supernatant after the treatment of 0, 0.2, 0.4, 0.8μg/ml ADM for 24h and were labeled as EVs1, 2, 3 and 4 respectively. Linc-VLDLR mRNA expression in EVs4 was significantly higher than that in EVs1-3 (P<0.01). ADM IC50 for Eca109 cells in EVs4 group was significantly higher than that in other groups after the treatment of EVs1-4 on Eca109 cells for 48h (P<0.05). Flow cytometry results showed that the proliferation index of Eca109 cells in EVs4 group was significantly higher than that in EVs1-3 and control groups (P<0.01). Linc-VLDLR and ABCG2 mRNA expression levels in Eca109 cells of EVs4 group were significantly higher than these of EVs1-3 and control groups (P<0.05). Conclusions High expression of linc-VLDLR and ABCG2 gene in esophageal cancer cells is involved in the formation of esophageal cancer resistance. EVs released by drug-resistant cells can upregulate the expression of ABCG2 in esophageal cancer cells and regulate the drug resistance of esophageal cancer cells, which is related to the linc-VLDLR gene carried by EVs.

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