人胚胎干细胞(hESC)移植治疗心肌梗死机理的多模态分子影像学研究

The aims of this NSFC (National Natural Science Foundation of China) Grant in Molecular Imaging are to integrate the fields of molecular imaging, stem cell biology, and cardiovascular physiology to provide key biological insights into preclinical investigations on stem cell differentiation, survival, and function in vivo. In this proposal, versatile triple-reporter genes (HSV-ttk, herpes simplex virus truncated thymidine kinase; Fluc, firefly luciferase；Rluc， renilla luciferase) will be developed for tracking human embryonic stem cell（hESC） derived endothelial cells fate in vivo and molecular imaging methods, e.g., PET and bioluminescence imaging, will be used to monitor hESC transplantation in mouse myocardial ischemia models. Moreover, hESCs will be transfected with an inducible, bidirectional tetracycline (Bi-Tet) promoter driving VEGF(165) and renilla luciferase (Rluc). We hypothesize that cells transfected with inducible vascular endothelial growth factor 165 (VEGF(165)) can improve their survival and can be monitored by novel molecular imaging (Rluc) techniques. Furthermore, hESC-ECs with reporter genes (Rluc-RFP-HSV-ttk) will be transplanted into transgenic Vegfr2-Fluc mouse model. With Fluc expression under VEGFR expression, the angiogenesis of ischemia myocardium can be monitored by Fluc imaging.And near-infrared imaging will be introduced for assessment of apoptosis and angiogenesis of transplanted cells. We are hopeful that this project would lead to (1)reporter gene labeled hESC can be well tracked by multimodility molecular imaging， (2) molecular imaging serving a critical role for monitoring VEGF derived gene therapy and transplanted cells induced angiogenesis,and(3)molecular imaging provides a well established platform to investigate the transplanted cells fate in basic and clinical research.

通过对机体内部生理或病理过程在分子水平上进行无损伤的、实时的成像，分子影像技术为干细胞治疗提供了重要的评价工具。本项目将通过三融合报告基因方法标记人胚胎干细胞(hESC), 应用多模态分子影像学手段(生物发光、PET)，长时间动态的观察移植的人胚胎干细胞来源的内皮细胞(hESC-ECs)；利用VEGF条件表达的报告基因系统，阐明VEGF基因治疗与细胞存活的关系；通过转基因动物模型(Vegfr2-luc)，结合多模态分子影像学手段(生物发光、PET、近红外)，研究移植的人胚胎干细胞来源的内皮细胞(hESC-ECs)在小鼠心肌梗死模型中细胞凋亡、修复损伤组织的血管新生机理。通过本研究能够阐明多模态分子影像学方法：1)能较好的监测移植的干细胞；2)能够在分子水平上对VEGF的基因治疗、移植细胞诱导的血管新生进行无损伤的、实时的成像；3)为干细胞治疗缺血性心脏病的机理研究提供有力的评价工具。

冠状动脉粥样硬化所致的心肌缺血坏死已成为人群的主要死亡原因，干细胞移植治疗通过促进血管新生和心肌再生，能够降低病人的致残率和致死率。然而干细胞移植后的监测一直存在问题，移植细胞的动态变化难以被量化，同时干细胞在体内的过程及其作用机理尚不清楚。本项目通过应用报告基因以及转基因动物模型等多种方法，结合多模态分子影像学手段，长时间的动态的观察移植的人胚胎干细胞来源的内皮细胞治疗缺血性心脏病模型，以及其修复损伤组织的机理。本项目阐明了：1) 分子影像学方法能较好的为干细胞移植治疗提供评价标准, 能够在分子水平上对人体内部生理或病理过程进行无损伤的、实时的成像，为干细胞治疗缺血性心脏病提供了一种有效的监测手段; 2) 多模态分子影像学方法是阐明干细胞移植治疗心肌缺血组织修复机理的重要工具。