稳定转染红色荧光蛋白基因的人脑胶质瘤起始细胞可自发融合宿主骨髓间充质干细胞并诱导恶性转化  

Human glioma-initiating cells stably transfected with red fluorescent protein gene can spontaneously fuse with host-derived bone marrow mesenchymal stem cells and induce malignant transformation

作　　者：韩辉 代兴亮 程宏伟[2] 兰青[1] Han Hui;Dai Xingliang;Cheng Hongwei;Lan Qing(Department of Neurosurgery, the Second Affiliated Hospital of Soochow University,Suzhou 215004, Jiangsu Province, China;Department of Neurosurgery, the First Affiliated Hospital of Anhui Medical University, Hefei 230022, Anhui Province, China)

机构地区：[1]苏州大学附属第二医院神经外科,江苏省苏州市215004 [2]安徽医科大学第一附属医院神经外科,安徽省合肥市230022

出　　处：《中国组织工程研究》2019年第25期3944-3950,共7页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(81702457),项目负责人:代兴亮;苏州市科技计划项目(SYS201723),项目负责人:代兴亮~~

摘　　要：背景:异种移植瘤中宿主来源间质细胞恶性转化已有报道,但是对恶变的机制知之甚少。目的:旨在使用双色荧光示踪的体内模型研究宿主来源肿瘤间质细胞的恶性转化,并探讨可能的机制。方法:将稳定转染红色荧光蛋白基因的人脑胶质瘤起始细胞(glioma-initiatingcellstransfectedwithred fluorescent protein gene,GICs-RFP)接种到表达绿光荧光蛋白的裸鼠体内,建立具有2种荧光的动物模型;通过激光共聚焦观察移植瘤的荧光表达;流式细胞仪检测和分选各种荧光细胞,包括绿色荧光蛋白细胞、红色荧光蛋白细胞和表达这2种荧光的融合细胞;体外亚克隆得到具有恶性肿瘤细胞特征的同时表达绿色荧光蛋白/红色荧光蛋白两种荧光的融合细胞;体外鉴定细胞来源、表面标记物和恶性特征,裸鼠皮下移植验证融合细胞致瘤特性。实验经苏州大学动物实验伦理委员会批准,批准号为ECSU-201800090。结果与结论:GICs-RFP在绿色荧光蛋白裸小鼠的体内致瘤率均为100%(14/14);动物移植瘤模型中均可观察到融合细胞,分选、克隆到的融合细胞不仅共表达绿色荧光蛋白和红色荧光蛋白,而且共表达GICs-RFP标记物Nestin和骨髓间充质干细胞标记物CD44、CD105和CD29;融合细胞在体外表现出高度增殖活性,较GICs-RFP更具侵袭和迁移特征;融合细胞(1×105个/只)在无胸腺裸小鼠的致瘤率为100%(4/4)。提示GICs-RFP自发融合宿主骨髓间充质干细胞并诱导恶性转化,为肿瘤异质性的细胞来源提供了新的双色荧光示踪的证据。BACKGROUND: Malignant transformation of host-derived stromal cells in xenograft tumors has been reported, but little is known about the mechanisms of malignant transformation.OBJECTIVE: To investigate the malignant transformation of host-derived tumor stromal cells and to explore the possible mechanisms in an in vivo model of two-color fluorescent tracing.METHODS: Human glioma-initiating cells stably transfected with red fluorescent protein gene(GICs-RFP) were inoculated into nude mice expressing enhanced green fluorescent protein(EGFP). An in vivo model of two-color fluorescence tracer was established. The fluorescent expression of transplanted tumors was observed by confocal laser scanning. Flow cytometry was used to detect and classify various fluorescent cells, including EGFP cells, RFP cells and EGFP/RFP fusion cells. The fusion cells co-expressed EGFP/RFP and had malignant proliferation characteristics because of in vitro subcloning. Origin, surface markers and malignant features of the cells were identified in vitro.The tumorigenicity of fusion cells was verified by subcutaneous transplantation in nude mice. The study was approved by the Animal Ethics Committee of Soochow University(approval No. ECSU-201800090).RESULTS AND CONCLUSION: The tumorigenic rates of GICs-RFP in EGFP nude mice were 100%(14/14). Fusion cells were observed in the transplanted tumors of all the animal models. The fusion cells not only co-expressed EGFP and RFP, but also co-expressed GICs-RFP marker Nestin and bone marrow mesenchymal stem cell markers CD44, CD105 and CD29. Fusion cells showed high proliferative activity and more invasive and migratory characteristics in vitro as compared with GICs-RFP. The tumorigenicity of fusion cells(1×105 cells per mouse) in thymus-free nude mice was 100%(4/4). These results indicate that GICs-RFP spontaneously fuses with host-derived bone marrow mesenchymal stem cells and induces malignant transformation, which provides new evidence for two-color fluorescence tracing of heterogeneous tumor cell

关 键 词：荧光蛋白示踪 细胞融合 恶性转化 胶质瘤起始细胞 荧光裸小鼠 肿瘤微环境 动物模型 间充质干细胞 肿瘤干细胞 

分 类 号：R441[医药卫生—诊断学] R364[医药卫生—临床医学]