Cardiac Differentiation of Adipose Tissue-Derived Stem Cells by Light Stimulation

描述（由申请人提供）：心肌梗死表现为肌肉纤维的丧失，最终无法再生肌肉组织。基于细胞的治疗可以恢复缺血区域，脂肪组织可以为这种治疗提供一个简单的细胞来源。我们以前已经表明，人类脂肪组织中含有有可能分化成心血管谱系的细胞。现有分化方案的主要缺点之一是脂肪组织源性干细胞（ASCs）的产量非常低，生化刺激产生随机空间组织和电非同步心肌细胞菌落。因此，显然需要新的方法来理解和操纵分化过程，以获得更高产量的同步心肌细胞，从而有效地植入受损心脏。为了实现这一目标，我们将利用一种新的光遗传学系统，该系统最近被证明可以在一定的时间尺度上调节神经元的激活和抑制。具体来说，在我们的目标中，我们提出：1)鉴定心包脂肪组织（PTA）中特定的心脏祖细胞亚群。根据上述初步数据，我们的工作假设是PTA含有充足的多能细胞来源。2)确定光遗传学在调节人类脂肪干细胞（hASCs）向心血管细胞分化中的作用。我们在初步数据的基础上再次假设，连续的膜电位去极化促进了hASCs的承诺，促进了CMs的产生。3)评估移植的电同步hASCs对缺血心肌再生的贡献。已有研究表明，移植的hASCs-CMs可改善心脏功能，本研究假设同步hASCs-CMs可显示更高的植入率，并直接促进心脏收缩功能。这些实验将有助于我们了解ASCs分化的机制，并为自体干细胞移植治疗缺血性心肌病提供依据。

DESCRIPTION (provided by applicant): Myocardial infarction shows a loss of muscle fibers with ultimate failure to regenerate muscle tissue. Cell based therapies may restore ischemic areas and adipose tissue can provide a facile source of cells for such therapy. We have previously shown that human adipose tissue contains cells which have the potential to differentiate into cardiovascular linage. One of the main disadvantages of the existing differentiation protocols is that yields of Adipose Tissue-Derived Stem Cells (ASCs) have been very low and biochemical stimulation produces random spatial organization and electrically non-synchronization cardiomyocytes colonies. Therefore there is an obvious necessity for new approaches to understand and manipulate the differentiation process in order to achieve higher yields of synchronous cardiomyocytes that can engraft effectively within the damaged heart. To approach this goal we will utilize a novel optogentics system that has been recently shown to allow tunable neuronal activation and inhibition over a range of timescales. Specifically, in our aims we propose to: 1) Identify specific sub-populations of cardiac progenitor cells within Pericardial Adipose Tissue (PTA). Based on preliminary data referred to above, our working hypothesis is that PTA contains an ample source of pluripotent cells. 2) Determine the role of opto-genetics, in regulating human adipose stem cells (hASCs) differentiation to cardiovascular cells. We postulate again on the basis of our preliminary data, that continuous membrane potential depolarization promote hASCs commitment to facilitate the generation of CMs and 3) Evaluate the contribution of transplanted electrically synchronous hASCs to ischemic myocardial regeneration. It has been shown that transplanted hASCs-CMs improve cardiac function, here we hypothesize that synchronous hASCs-CMs can show higher rate of engraftment and directly contribute to the systolic function. These experiments should help us understand the mechanisms of ASCs differentiation and provide the basis for autologous stem cell transplantation for the treatment of ischemic cardiomyopathy. PUBLIC HEALTH RELEVANCE: The project is relevant to the NIH mission because patients with ischemic cardiomyopathy will benefit from therapy whereby injected differentiated cardiomyocytes can be incorporated into the heart muscle. We have already shown that perivascular adipose stromal cells have a potential in differentiating into cardiomyocytes and that using our novel approach we can stimulate cells to redirect them into cardiac linage cells. Hereby we propose to characterize and enhance the potential of these cells and explore how they can help in cardiac muscle regeneration using optogenetic stimulation. This research could provide a basis for autogenous tissue being used as a source of cells for the treatment of end-stage ischemic heart disease.