小鼠肝癌细胞外泌体对树突状细胞成熟与功能的影响  

Effects of exosomes on dendritic cell maturation and function in mouse hepatocellular carcinoma cells

作　　者：许伟兵 范璐璐[1] Xu Weibing;Fan Lulu(Dept of Oncology,The First Affiliated Hospital of Anhui Medical University, Hefei 230022)

机构地区：[1]安徽医科大学第一附属医院肿瘤内科,合肥230022

出　　处：《安徽医科大学学报》2021年第8期1197-1200,共4页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81402040)。

摘　　要：目的探讨小鼠肝癌细胞(Hepa1-6)来源的外泌体对树突状细胞成熟和功能的影响。方法提取小鼠Hepa1-6细胞的外泌体,用透射电镜来鉴定外泌体的外形,Western blot方法测定外泌体的标志蛋白CD63和TSG101的表达,使用小鼠Hepa1-6细胞外泌体冲击树突状细胞(DCs),DCs成熟相关表面分子CD40、CD80、CD83、CD86、MHC-Ⅱ的表达使用流式细胞术检测;将小鼠T淋巴细胞和小鼠Hepa1-6细胞外泌体致敏的DCs一起孵育5 d,使用CFSE染色标记和流式法来测定共孵育T细胞的分裂增殖情况。结果透射电镜可以观察到从Hepa1-6细胞的上清液中收集到的外泌体,表现为圆形或类圆形膜性囊泡样结构,符合外泌体的典型特征;Western blot检测表明小鼠Hepa1-6细胞来源的外泌体表达CD63、TSG101标志性蛋白,不表达细胞色素C;从小鼠Hepa1-6细胞提取来的外泌体致敏DCs,可上调表面MHC-Ⅱ类分子、成熟标志分子CD83和共刺激分子CD40、CD80、CD86(P<0.05)。且随着外泌体浓度的增加,DC表面分子表达增高;同时可促进T淋巴细胞的增殖(P<0.05)。结论小鼠Hepa1-6细胞提取的外泌体可促进DC2.4的朝向成熟表型分化,并促进T细胞的增殖。Objective To investigate the effects of exosomes derived from Hepa1-6 cellson dendritic maturation and function.Methods Exosomes were extracted from Hepa1-6 cells,and the shape of exosomes was identified by transmission electron microscopy.The expressions of the marker proteins of exosomes,CD63 and TSG101 were determined by Western-blot.Dendritic cellswere impacted with exosomes from Hepa1-6 cells,and the expression of maturation related surface molecules CD40,CD80,CD83,CD86 and MHC-Ⅱwas detected by flow cytometry.Mouse T lymphocytes and exosomes shock DCs from Hepa1-6 cells were co-incubated for 5 d,and the proliferation of T lymphocytes was determined by CFSE staining and flow cytometry.Results The exosomes collected from thesupernatantof Hepa1-6 cells was observed by transmission electron microscopy and exosomesshowed round or circular membranous vesicle-like structure,which was consistent with the typical characteristics of exosomes.Western-blot analysis showed that exosomesderived from Hepa1-6 cells expressed CD63 and TSG101,but did not express Cytochrome C.The exosomes extracted from Hepa1-6 cells were used to shock DCs,and the surface MHC-II molecules,mature marker CD83 and co-stimulating molecules CD40,CD80 and CD86 were up-regulated.Moreover,with the increase of exosomes concentration,the molecular expression of DC surface increased(P<0.05).Exosomes also promoted the proliferation of T lymphocytes(P<0.05).Conclusion Exosomes extracted from Hepa1-6 cells could promote the differentiation of DC2.4 toward mature phenotype and promote the proliferation of T cells.

关 键 词：肝细胞癌 外泌体 树突状细胞 

分 类 号：R735.7[医药卫生—肿瘤]