乳腺癌MDA-MB-231细胞源exosomes对血管内皮细胞EGFR表达的影响  

Effect of exosomes derived from breast cancer cell line MDA-MB-231 on EGFR expression in HUVECs

作　　者：谢莹珊[1] 沈宜[1] 隆霜[1] 范维柯[1] 姜蓉[1] 陈黎[1] 

机构地区：[1]重庆医科大学干细胞与组织工程研究室,400016

出　　处：《重庆医学》2012年第26期2694-2697,I0001,F0002,共6页Chongqing medicine

基　　金：重庆市渝中区科委资助项目(2009)

摘　　要：目的研究人乳腺癌MDA-MB-231细胞源exosomes介导表皮生长因子受体(EGFR)对人脐静脉内皮细胞株(HUVECs)EGFR表达影响,探讨肿瘤组织血管内皮细胞异常表达EGFR的机制。方法超速离心及密度梯度离心法提取MDA-MB-231细胞源exosomes;免疫细胞化学法检测MDA-MB-231细胞EGFR的表达;Western blot法检测MDA-MB-231细胞、exosomes及HUVECs EGFR蛋白表达;免疫细胞化学法检测HUVECs与exosomes共培养24h后EGFR的表达;RT-PCR法检测MDA-MB-231细胞与实验组HUVECs细胞EGFR mRNA的表达。结果 EGFR在MDA-MB-231细胞中呈高表达,MDA-MB-231细胞及exosomes可见相对分子质量为170×103的EGFR蛋白呈阳性反应带,HUVECs呈阴性反应带;HUVECs与exosomes共培养24h后,镜下可见实验组部分HUVECs细胞质有淡黄色或棕黄色颗粒,EGFR阳性表达率为(21.4±3.1)%,与对照组(无EGFR蛋白表达)比较,差异有统计学意义(P<0.01)。MDA-MB-231细胞EGFR mRNA表达阳性,而实验组HU-VECs EGFR mRNA表达阴性。结论 MDA-MB-231细胞源exosomes携带癌基因EGFR,并能介导其向周围血管内皮细胞转移,这可能是肿瘤内皮细胞异常表达EGFR的一种方式。Objective To study the effect of exosomes derived from breast cancer cell line MDA-MB-231 on epidermal growth factor receptor（EGFR） expression in human umbilical vein endothelial cells（HUVECs） and to research the possible manner of abnormal expression EGFR of tumor endothelial cell.Methods Exosomes were purified by serial ultracentrifugation and sugar density ultracentrifugation.The expression of EGFR in MDA-MB-231 was detected by immunocytochemistry and Western blotting.The expression of EGFR in exosomes and HUVECs was detected by Western blotting.The expression of EGFR in HUVECs with exosomes co-cultured 24 h was detected by immunocytochemistry.RT-PCR was used to detect the mRNA expression of EGFR in MDA-MB-231 and experimental group.Results MDA-MB-231 cells expressed high level of EGFR.In the results of Western blotting,the positive reaction band was found in 170 kD which coincided with the molecular weight of EGFR in MDA-MB-231 and exosomes,and negative reaction in HUVECs.When co-cultured with HUVECs and exosomes for 24 h,the result of immunocytochemistry showed that the part of the experimental group was found straw color or brown madder particle in HUVECs cytoplasm,the ratio of positive expression was 21.4±3.1%.There was significant increase compared with the control group（P〈0.01）.RT-PCR exhibited positive of EGFR mRNA in MDA-MB-231,and negative in the experimental group of HUVECs.Conclusion Exosomes derive from MDA-MB-231 cell lines with oncogene EGFR cargo,and could mediate the cargo to endothelial cell.It may be the possible manner of abnormal expression EGFR in tumor endothelial cells.

关 键 词：乳腺肿瘤 内皮细胞 表皮生长因子 脐静脉 

分 类 号：R737.9[医药卫生—肿瘤]