可显著提高干细胞心肌梗死模型疗效的新型细胞移植体系研究

Cardiac stem cell therapy has been used exclusively in clinical trials and animal studies to improve function of the hearts injured by myocardial infarction (MI). Its efficacy is directly dependent on the number of delivered cells that are retained and survived in the infarct heart. To date, it is impossible for clinical application of cardiac stem cell therapy because of its low efficacy caused by inferior cell retention and survival. A major hurdle in improving the efficacy of current cardiac stem cell therapy is lack of suitable cell carriers. Existing cell carriers, mostly saline and injectable hydrogels, are not ideal for cardiac stem cell therapy. This is because: 1) they cannot solidify quickly enough in the heart tissue to prevent delivered cells from squeezing out of the heart; 2) most injectable hydrogels solidify in blood, and thus can block blood vessels in the heart causing further damage; 3) most hydrogels are not stretchable and are likely in a broken state during heart beating. These hydrogels are thus unable to provide efficient mechanical support to the heart tissue; and 4) current cell carriers cannot promote cell survival under harsh condition of the infarct heart. The goal of this project is to create a new class of hydrogel-based cell carriers that address the above limitations in order to significantly improve the efficacy of current cardiac stem cell therapy. Towards this goal, we have created novel hydrogels that have high gelation rate and stretchability, and do not solidify in blood. The specific objective of this proposal is to create a prosurvival microenvironment in these hydrogels to enhance cell survival under harsh condition of the infarct hearts. We hypothesize that a prosurvival microenvironment that addresses the hypoxic condition of the infarct hearts will significantly improve cell survival. To test the hypothesis, we will create a microenvironment capable of releasing oxygen to oxygenate cells, and evaluate its efficacy in improving survival of an emerging cell type for cardiac therapy, cardiosphere-derived cells (CDCs). CDCs improve heart function mainly by paracrine effect. We expect that enhancing CDC survival will provide prolonged paracrine effect for heart function improvement.Aim 1 will create a prosurvival microenvironment to enhance CDC survival and paracrine effect under hypoxic condition.Aim 2 will evaluate efficacy of the developed cell carriers in infarct hearts.

心肌梗死造成大量心肌细胞死亡。干细胞移植是目前最有潜力的疗法之一，但面临着移植效率极低的巨大挑战。造成移植效率低的关键原因是：细胞载体粘度低，大量细胞在注射过程中流失，因而细胞滞留率低；同时滞留的细胞处于坏死心肌的低氧环境，其生存率低。当前尚无有效措施可解决以上问题。我们设想用一种可注射、高粘度、可在受损心肌中快速凝固、且能改善低氧环境的细胞载体解决此问题。我们率先合成了可注射、可快速凝固又不堵塞血管的水凝胶载体，可显著提高细胞滞留率。近期我们又制备出了能释放氧气的微球，可大幅提高低氧条件下细胞存活。但目前微球的释放功能仅能维持2周，且在动物体内的效果不详。本项目旨在建立水凝胶与氧气释放微球结合的、可提高细胞滞留率和长期生存率的新型移植载体。通过观察干细胞在急性心肌梗死动物中的滞留率和生存率，以及心肌的血管化、心脏功能和水凝胶的生物相容性来验证新载体的功能，以期为细胞治疗提供新的科学依据。

缺血性疾病是老年人的常见疾病，也是世界上致残率最高的疾病之一。干细胞移植作为一个新兴的治疗手段开始应用于缺血性疾病的治疗。移植干细胞的载体是影响干细胞滞留率的一个主要因素，目前国内外尚无理想的应用于干细胞移植的专用载体，因此亟需创建一种新型高效的细胞载体。本课题组制备出温敏、pH敏感且机械性能适宜的水凝胶作为干细胞移植的载体，提高移植干细胞的滞留率；同时构建了氧气释放微球，并引入该载体，可以为移植干细胞供氧3周以上，以期达到提高移植干细胞及周围的肌肉细胞在缺血、缺氧环境下的存活率，从而促进肌细胞再生。引入氧气释放微球的干细胞移植体系，体外检测，dsDNA浓度测定随时间增加而增高，且明显高于对照组，与活细胞荧光成像结果一致，证实新的移植体系能大大提高干细胞的生存率；并且发现氧气释放微球结合干细胞治疗显著促进缺血组织中的肌肉细胞及血管内皮细胞方向分化；苏木精-伊红染色、梅森三色染色、多普勒血流检测、跑步测试等方法证实此体系有效的改善了缺血肢体肌肉萎缩情况，有助于肌肉功能的恢复。氧气释放微球结合干细胞治疗体系能大大提高干细胞的滞留率和生存率，并能有效的分化成所需细胞系，改进干细胞治疗下肢缺血性疾病的效果。本研究的研究结果为干细胞治疗缺血性血管病提供了新的理论基础。