心脏干细胞植入心脏脱细胞基质构建人工生物心脏的探索性研究

Recent progresses in cardiac stem cell research cast a new light on the construction of bioartificial heart(BAH). Researchers from our group and others have isolated cardiac stem cell in adult heart. Two distinct subpopulations of endogenous cardiac stem cells (CSCs) are identified: myogenic CSCs (mCSCs), which are characterized by the presence of c-kit receptor, reside in the myocyte niche surrounded by those matured cardiomyocytes, whereas vasculogenic CSCs (vCSCs), which express c-kit and KDR, are stored in the vascular niche found in the vessel wall. They both possess the fundamental properties of stem cells: self-renewing and multipotent. In vitro, mCSCs preferentially differentiate into myocyte lineage, while vCSCs tend to differentiate into vascular endothelial or smooth muscle lineages. In our study, we plan to generate BAH with coronary circulation by seeding vCSC and mCSC into the vascular and chamber part of decellularized hearts separately in the Bi-ventricle heart perfusion system which can faithfully mimic in-vivo heart working condition. By comparative analysis of the stucture and function of normal hearts and BAHs, we try to obtain critical experiment evidences for construction of more simulated, more transplantable BAH and the following transplantation experiment in rat. Here, directed differentiation of CSC into cardiac tissue and autologous isolation of CSC from human auricular appendage make CSC a best candidate cells for clinic application， and separate and localized seeding of vCSC and mCSC may provide a better way to vascularize the artificial tissue or organ.

近年来心脏干细胞（CSC）的研究进展为人工生物心脏（BAH）的构建带来了新的曙光。本课题组和其他的研究结果显示，在成年动物、人体心脏内均含有CSC,部分CSC（即vCSC c-kit+/KDR+）存在于心脏血管壁小生境，主要分化为平滑肌和内皮细胞；部分CSC（即mCSC,c-kit+/KDR-）存在于心肌组织间小生境，主要分化为心肌细胞。本课题拟利用CSC这一特性，在双心室工作心灌流系统中分步、定位种植大鼠vCSC和mCSC于心脏脱细胞支架的血管和心肌部分，模拟心脏体内工作环境，构建拥有冠脉系统的再细胞化心脏（RCH）。通过对比分析正常心脏和RCH的结构和功能，探讨我们的研究是否能构建出更具可移植性的BAH用于BAH移植的动物实验。课题首次选用分化能力好，将来可于人自体心耳提取的CSC构建BAH，极具临床应用前景；而分步、定位种植vCSC和mCSC可以巧妙的解决心脏血管化这一难题。

治疗心衰一直是心血管领域的重要研究课题，人工生物心脏备受人们的关注。近年来心脏干细胞的研究对人工生物心脏的制备带来了新的曙光。本课题组和其他的研究结果显示，在成年动物、人体心脏内均含有CSC。CSC具有良好的分化能力，可向血管内皮、平滑肌及心肌方向分化。我们通过冠脉灌流成功制作出无细胞的心脏支架，通过电镜、免疫荧光和免疫组化对支架进行了鉴定，支架完整的保持了心脏原有的结构并除去了细胞成分。我们成功从小鼠心脏中分离出心脏干细胞并进行培养，将其种入制作好的支架中，但是发现细胞的存活率较低。在心脏干细胞方面我们做了关于增殖、分化等方面的功能实验，发现地塞米松等因素的干预可促进干细胞向心肌细胞方向分化并且这些干预因素可促进干细胞的增殖。