Magnetic Cellular Assembly and Microfluidic Conditioning for Generation of Functi

描述（由申请人提供）：心血管疾病影响7000万美国人，造成3000亿美元的经济负担，占美国所有死亡人数的近40%。常规的心肌损伤治疗不能实现心肌再生。因此，基于干细胞和组织工程的心脏组织再生方法已被积极追求。现有的基于支架的3D组织工程方法可以在非仿生环境中实现组织再生，并且对3D结构的控制有限。模拟三维细胞组织和复制在体内看到的机械加载模式将解决与传统技术相关的缺点，并显着增强体外生成的组织的功能。在本提案中，我们将启用一个平台技术，用于封装在磁性水凝胶（m - gel）中的多种细胞类型的磁性细胞组装，模拟组织水平的细胞密度，形成3D心脏组织结构。微型灌注网络将集成到3D结构中，以确保营养物质的输送，并满足心脏细胞的高代谢需求。图案构建将在微流体心脏细胞培养模型（CCCM）中培养，该模型精确地复制左心室的压力和拉伸负荷。我们之前的工作清楚地表明，我们有能力在2D和3D中构建复杂的结构，并在促进循环拉伸、细胞排列和自发同步收缩的现实机械应力下完成CCCM内的细胞培养。具体来说，我们将：(A)将心脏细胞包裹在磁性水凝胶（m -凝胶）中，围绕牺牲的孔隙网络进行组装，以获得体内类似组织水平的细胞密度和结构；(B)利用CCCM在模拟左心室压力-体积变化的细胞培养环境中培养磁组装心脏组织构建体，并在培养后提取完整组织；(C)在体外完成磁组装和微流体调节组织构建体的形态和功能特性表征，并确定3D模式的作用。在压力和拉伸负荷方案下，微尺度灌注网络和培养的整合对体外心脏发生和产生可用于修复心肌梗死的功能性心肌组织的能力有影响。虽然将使用鸡胚胎心脏细胞群来证明所提出活动的可行性，但所开发的技术将与心脏干细胞和祖细胞群兼容，以产生功能性心脏补片。

DESCRIPTION (provided by applicant): Cardiovascular diseases affect 70 million Americans, resulting in an economic burden of $300 billion and accounting for nearly 40% of all deaths in the US. Conventional treatments of myocardial injury do not achieve myocardial regeneration. Therefore, stem cell and tissue engineering based approaches for cardiac tissue regeneration have been actively pursued. Existing scaffold based 3D tissue engineering approaches accomplish tissue regeneration in non-biomimetic environments and with limited control over 3D architecture. Mimicking the 3D cellular organization and replicating mechanical loading patterns seen in vivo will address shortcomings associated with conventional techniques and significantly enhance functionality of tissues generated in vitro. In this proposal, we will enable a platform technology for magnetic cellular assembly of multiple cell types encapsulated in magnetic hydrogels (M-Gels) that mimic the tissue-level cell densities to form 3D cardiac tissue structures. Microscale perfusion networks will be integrated into the 3D constructs to ensure delivery of nutrients and meet the high metabolic needs of cardiac cells. Patterned constructs will be cultured within a microfluidic Cardiac Cell Culture Model (CCCM) that accurately replicates pressure and stretch loading seen in the left ventricle. Our prior work clearly demonstrates our ability to pattern complex architectures in both 2D and 3D and accomplish cell culture within the CCCM under realistic mechanical stresses that promote cyclic stretch, cell alignment and spontaneous synchronous contractions. Specifically we will: (A) accomplish assembly of cardiac cells encapsulated in magnetic hydrogels (M-gels) around a sacrificial porogen network to attain in vivo like tissue-level cell densities and architecture, (B) utilize th CCCM to culture magnetically assembled cardiac tissue constructs in a cell culture environment that mimics the pressure-volume changes seen in the left ventricle and enable extraction of intact tissue following culture and (C) accomplish characterization of morphological and functional properties of magnetically assembled and microfluidically conditioned tissue constructs in vitro and determine the role of 3D patterning, integration of microscale perfusion networks and culture under loading regimens of pressure and stretch on in vitro cardiogenesis and the ability to generate functional myocardial tissue that can potentially be used to repair myocardial infarctions. Though chick embryonic cardiac cell populations will be used to demonstrate the feasibility of proposed activities, the developed techniques will be compatible with cardiac stem and progenitor cell populations for generation of functional cardiac patches.