Controlled gene delivery for in vivo vascularization of an engineered cardiac pat

描述(申请人提供)：心脏病仍然是发达国家的主要死亡原因。大约7000万美国人(超过四分之一的人口)患有心血管疾病，死亡人数几乎是所有癌症总和的两倍，每年用于治疗心血管疾病(www.americanheart.org/presenter.jhtml?identifier=4478).的费用约为3000亿美元组织工程学提供了在体外培养天然心肌的功能等价物用于组织修复的潜力，并探索治疗或预防这种疾病的新方法。然而，主要的挑战仍然存在，包括需要建立临床相关的人类细胞来源，以及在体外培养和体内植入后将工程组织血管化。我们建议通过控制呈现给细胞的血管内皮生长因子的水平，设计出具有高血管形成能力的心肌组织补片。我们要解决的基本问题是，在体外培养过程中和植入梗死心肌后，是否可以利用心肌细胞和表达可控水平的血管内皮生长因子的细胞共培养来调节血管网络的形成和成熟。这一应用的直接目标是通过生物反应器将心肌细胞和血管内皮生长因子转基因细胞在弹性体支架上共培养，产生具有血管形成能力的同步收缩的心肌组织。我们提出了一套有重点且协调良好的体外研究(心脏和表达血管内皮生长因子的细胞的生物反应器共培养)和体内研究(在大鼠心肌梗死模型中)，具体目的如下：(1)设计一种基于新生大鼠心肌细胞和血管内皮生长因子转基因成肌细胞的心脏补片。(2)制备释放可控的血管内皮生长因子的成人干细胞。(3)构建以新生大鼠心肌细胞和血管内皮生长因子基因转染人干细胞为基质的心脏补片。在所有情况下，细胞都将使用先进的心脏组织工程生物反应器，在高度多孔的通道弹性体支架上培养。将确定移植物的活性、功能特性、与宿主组织的整合以及血液灌注量。这项计划中的工作如果成功，将为利用成人干细胞来源的心脏和血管内皮生长因子转基因细胞设计功能性人类心脏移植物提供基础。我们目前无法将厚厚的细胞团血管化和灌流，这阻碍了构建多种类型组织的努力，最关键的是，包括心肌。

DESCRIPTION (provided by applicant): Heart disease remains the leading cause of death in developed countries. About 70 million Americans (more than one-fourth of the population) live with cardiovascular disease, with a death toll that is almost twice as high as that for all cancer combined, and the cost of ~300 billion dollars spent every year to treat the cardiovascular disease (www.americanheart.org/presenter.jhtml?identifier=4478). Tissue engineering offers a potential to grow in vitro functional equivalents of native myocardium for use in tissue repair, and to investigate new ways to treat or prevent the disease. However, major challenges remain, including the need to establish a clinically relevant source of human cells, and to vascularize the engineered tissue, both during the in vitro cultivation and following implantation in vivo. We propose to engineer a patch of myocardial tissue with high capacity for vascularization, by controlling the levels of VEGF presented to the cells. The fundamental question we address is if co-culture of cardiac myocytes and cells expressing controllable levels of VEGF can be utilized to mediate the formation and maturation of a vascular network, during in vitro culture and following implantation onto infarcted myocardium. The immediate goal of this application is to generate synchronously contracting myocardial tissues with ability for vascularization, by bioreactor co-culture of cardiomyocytes and VEGF transfected cells on elastomer scaffolds. We propose a set of focused and well coordinated in vitro studies (bioreactor co-culture of cardiac and VEGF expressing cells) and in vivo studies (in a rat heart infarction model), with the following specific aims: (1) To engineer a cardiac patch based on neonatal rat cardimyocytes and VEGF-transfected myoblasts. (2) To generate adult human stem cells with controllable release profiles of VEGF. (3) To engineer a cardiac patch based on neonatal rat cardiomyocytes and VEGF-transfected human stem cells. In all cases, cells will be cultured on a highly porous, channeled elastomer scaffold, using an advanced bioreactor for cardiac tissue engineering. The viability, functional properties, integration with the host tissue and blood perfusion will be determined. The planned work, if successful, would provide a basis for engineering functional human cardiac grafts using cardiac and VEGF transfected cells derived from adult human stem cell sources. Our current inability to vascularize and perfuse thick cell masses has hindered efforts to build many types of tissues, including, most critically, cardiac muscle.