胃癌耐药细胞EXOs的提取鉴定及可能存在的耐药信息传递作用的初步探讨  

The role of gastric cancer resistant cells-derived exosomes in intercellular communication

作　　者：宋慧聪 邓光 张晔[1] 邓明明 胡雪君[2] 于若曦 徐玲[1] 曲秀娟[1] 刘云鹏[1] Song Huicong;Deng Guang;Zhang Ye;Deng Mingming;H u Xuejun;Y u Ruoxi;X u Ling(Laboratory I of Cancer Institute;Department of Respiratory Medicine,The First Hospital of China Medical University,Shenyang110001,China.)

机构地区：[1]中国医科大学附属第一医院肿瘤研究所一室,辽宁沈阳110001 [2]中国医科大学附属第一医院呼吸科,辽宁沈阳110001

出　　处：《现代肿瘤医学》2018年第13期2002-2005,共4页Journal of Modern Oncology

基　　金：国家自然科学基金(编号:81270036;30901736);辽宁省教育厅优秀人才支持计划项目(编号:LR2014023);辽宁省自然科学基金(编号:2013021057)

摘　　要：目的:探讨胃癌耐药细胞(SGC7901/Adr)分泌的外泌体(exosomes,EXOs)在细胞间可能存在的耐药信息传递作用。方法:选用胃癌亲本细胞SGC7901及其阿霉素耐药细胞SGC7901/Adr为模型,用PEG方法提取EXOs;透射电子显微镜观察鉴定细胞EXOs形态;Western Blot检测CD9、CD63、TSG101、Calreticulin以及P-糖蛋白(P-gp)的表达;运用激光共聚焦显微镜观察胃癌SGC7901和SGC7901/Adr细胞及加入阿霉素耐药细胞SGC7901/Adr外泌体处理后的敏感细胞内阿霉素药物浓度变化。结果:透射电子显微镜下观察到,胃癌SGC7901和SGC7901/Adr细胞来源的EXOs为直径在30～100 nm之间的囊性小泡,呈圆形或椭圆形,大小均匀一致。Western Blot检测胃癌SGC7901和SGC7901/Adr细胞来源的EXOs,携带有外泌体生物标记分子CD9(四次跨膜分子)、CD63(四次跨膜分子)以及TSG101表达,Calreticulin为阴性表达。进一步检测发现在SGC7901/Adr细胞来源的EXOs中存在P-gp蛋白表达。经过阿霉素耐药细胞SGC7901/Adr外泌体处理后的敏感细胞,胞内可见阿霉素聚集减少,核区分布减少,荧光强度减弱。结论:胃癌耐药细胞分泌的EXOs可能通过传递P-gp蛋白的形式,具有传递耐药信息的作用。Objective ：To investigate the role of exosomes （ EXOs） secreted by SGC7701/A d r cells in the drug resistancetransmission among cells. Methods ：SGC7901 and SGC7901/A d r cell lines were selected as cancer parent cell and resistant cell lin e,respectively. EXOs were isolated with PEG medium and visualized under transmission electron microscopy. Western Blot was used to detect the CD9 , CD63 , TSG101 , Calreticulin and Glycoprotein （ P - gp） . Cell uptake of adriamycin coZocal microscope of gastric cancer SGC7901,SGC7901/A d r cells and sensitive cells treated by adriamycin - resistantcells SGC7901/A d r exosomes. Results ： EXOs derived from SGC7901 and SGC7901/A d r cells were membrane -bound micro - vesicles under transmission electron microscopy. They owned a characteristic ovvid the size distribution was mainly between 3 nm and 100 nm. CD9 ,CD63 and TSG101 were identified in SGC7701 and SGC7901/A d r cells - secreted EXOs by immunoblotting while the intracellular endoplasmic reticulum protein ＂ Calreticulin’’ was confirmed absent. Further detection showed that the protein expression level of P - gp was in SGC7901/A d r cells - derived EXOs compared to parent cells - derived EXOs. Sensitive cells treated by adriamycin- resistant cells SGC7901/A d r exosome, adriamycm fluorescence intensity became weaker than that of SGC7901. Conclusion ： EXOs secreted by gastric cancer resistant cells may play an important role transmission through transferring P - gp.

关 键 词：胃癌 外泌体 多药耐药 P-糖蛋白 

分 类 号：R735.2[医药卫生—肿瘤]