骨髓间充质干细胞定向迁移恶性胶质瘤的多模态报告基因成像研究

Mesenchymal stem cells (MSCs) have emerged as a promising cellular vehicle for gene therapy of malignant gliomas due to their property of tumor tropism. However, MSCs may show bidirectional and divergent effects on tumor growth that are highly dependent on their intricate and varying mutual interactions with tumor cells and with other stromal constituents in the tumor microenvironment. Hence, monitoring their biological behavior, including their migration, distribution, fate and therapeutic effects, in a real-time, accurate and sensitive manner is very important in developing an effective MSC- based therapeutic strategy for gliomas. Based on our previous studies, in this study, we will construct a lentivirus vector with FTH-Fluc gene overexpression. Rat MSCs will be genetically modified by using FTH-Fluc gene overexpressed vector to confer the capacities of MR and bioluminescent visualization to MSCs. After the genetically modified MSCs transplanted in rats bearing intracranial orthotopic gliomas, the feasibility, efficacy and biological safety of this genetic modification will be determined at the levels of in vitro and in vivo living animals. The interactions between MSCs and tumor microenvironments will be accessed. The migration, location, survival and differentiation of transplanted MSCs will be dynamic observed by using in vivo MRI and optical imaging. In light of the multidisciplinary integration of multimodality molecular imaging, tissue engineering and molecular biology, the aim of this project is to establish a new means to in vivo accurately and sensitively monitor the long-term biological processes of transplanted MSCs, and thereby to provide more experimental evidence to promote the clinical transformation of the MSC-based therapy for gliomas.

骨髓间充质干细胞（mesenchymal stem cells, MSCs）具有主动穿越血脑屏障而定向趋化至胶质瘤的特性，可作为胶质瘤靶向性治疗的理想载体。然而迁移到肿瘤的MSCs在肿瘤复杂的微环境中发挥作用不明确，对MSCs定向迁移至胶质瘤的生物学行为进行活体、动态、精准的可视化监测是MSCs治疗胶质瘤亟待解决的问题。本项目拟在前期研究基础上，构建铁蛋白-荧光素酶慢病毒载体，对大鼠MSCs进行双模态报告基因标记，将基因修饰的MSCs移植入红色荧光标记的大鼠原位胶质瘤模型中，利用MRI、生物发光成像对MSCs移植后在胶质瘤内迁移、定植、存活及转归情况进行多模态成像示踪，明确MSCs与肿瘤微环境的交互作用。本项目通过分子影像学、基因工程、分子生物学等多学科交叉融合，建立能长期、准确、定量示踪干细胞的技术体系，阐明MSCs定向迁移的生物学行为规律，为发展胶质瘤靶向治疗的新方法提供实验依据。

骨髓间充质干细胞（mesenchymal stem cells, MSCs）虽能主动穿越血脑屏障而定向趋化至胶质瘤，可作为胶质瘤靶向性治疗的理想载体。然而迁移到肿瘤的MSCs在肿瘤复杂的微环境中发挥作用不明确，对MSCs定向迁移至胶质瘤的生物学行为进行活体、动态、精准的可视化检测是MSCs治疗胶质瘤亟待解决的问题。本项目在前期研究基础上，成功构建铁蛋白（Ferritin heavy chain, FTH）-荧光素酶（Fluc）慢病毒载体，用于对大鼠MSCs进行MRI和生物发光双模态报告基因修饰，使得MSCs能被MRI和光学成像双模态精准示踪。体外细胞实验显示，FTH-Fluc能对MSCs进行高效标记，转染率达90%以上，同时明显上调MSCs的FTH及Fluc表达，在MRI和生物发光成像中显示出良好的成像效果，而转染不影响MSCs的增殖、凋亡水平、干性、分化及迁移能力。动物活体实验表明，将基因修饰的MSCs移植入红色荧光标记的大鼠原位胶质瘤模型中，其在胶质瘤内的定植效率和存活率能够被MRI、生物发光成像活体、动态监测至移植后7天，明确经肿瘤同侧颈内动脉注射的标记MSCs对胶质瘤的趋化效率是经瘤内注射的1/8，在到达肿瘤的第4天，MSCs大量死亡，而FTH-Fluc修饰的MSCs对胶质瘤生长未见促进或抑制作用。本项目通过分子影像学、基因工程、分子生物学等多学科交叉融合，建立能长期、准确、定量示踪干细胞的技术体系，阐明MSCs定向迁移的生物学行为规律，为发展胶质瘤靶向治疗的新方法提供研究依据。