Cardiac Regeneration through Growth Factor Eluting Microrod Scaffolds

描述(由申请人提供)： 美国有500万心力衰竭患者由于收缩心肌细胞不可逆转的损伤而心泵功能不佳。因此，通过细胞和组织工程来恢复心脏功能是非常可取的。本申请中提出的新方法结合了生物工程和基于细胞生物学的技术，直接针对心肌再生，这是一个重要的临床问题，目前的治疗方法并没有很好地解决这一问题。我们系统地提供了一种人工稳定形式的机械生长因子(MGF)(24个氨基酸，来自前激素的E结构域)，它是胰岛素样生长因子(IGF)家族的成员，随着常驻祖细胞的动员，在衰竭的小鼠心脏中显示出功能的恢复。我们利用心脏的自然修复能力，提供了一种微型支架(MRS)，不仅提供了天然的、快速降解的MGF，而且还提供了适当的细胞连接和分化所需的局部机械和地形图线索。我们的总体假设是，MRS局部定时释放MGF和IGF-1可以再生和加强受损的心肌，而不会产生有害的副作用。这在祖细胞/干细胞和培养的心肌细胞和动物模型上进行了测试。具体目标1确定了硬度变化对生长在3D微棒支架中的细胞的影响。我们优化了MRS特性，以调节干细胞和心肌细胞的增殖、谱系承诺、分化、收缩成熟度和连接性。特定目标2确定了在模拟正常和缺血心脏的环境条件下，携带生长因子(GF)的MRS的效果。我们研究了洗脱MRS的包封率、GF生物稳定性和脱细胞释放动力学。我们测定了从MRS释放的GF对祖细胞/干细胞和新生大鼠心室肌细胞增殖和迁移的影响，以建立在模拟正常和缺血心脏的培养条件下基因表达和细胞存活的变化。具体目标3确定了在MRS将GFS输送到梗死边缘区域后，在心肌梗死后不同阶段发生的细胞、分子和功能收益。我们研究了体内GF释放如何影响心脏祖细胞的迁移和分化。我们研究了局部MGF多肽对心功能、防止心肌细胞凋亡、防止不利的心脏重塑和减少瘢痕形成的有益效果。我们的长期目标是开发支持心肌再生的微创MGF疗法，以恢复衰竭心脏的心功能。 公共卫生相关声明(由申请人提供)：心力衰竭是一种常见的疾病，具有很高的残疾和死亡率负担。目前的估计是，在美国，心力衰竭每年造成大约100万次住院和100亿美元的医疗费用。心力衰竭的主要原因是缺血性心脏病、高血压和特发性扩张型心肌病，其共同的病理生理特征是功能心肌细胞数量不足。这项建议开发和测试了一种新的治疗性3D洗脱微支架(MRS)系统，以提供一种天然的心脏生长因子，以帮助培养和动物模型中受损心肌的再生和恢复。

DESCRIPTION (provided by applicant): Five million heart failure patients in the US have poor cardiac pumping due to irreversible damage of the contractile myocytes. Thus, recovery of cardiac function by cell and tissue engineering is highly desirable. The novel approach proposed in this application combines bioengineering and cell biology-based techniques to directly target the regeneration of heart muscle, which is an important clinical problem not well addressed by current therapies. We have systemically delivered an artificial stabilized form of the mechano-growth factor (MGF), (24 amino acid, E-domain peptide from the prohormone) that is a member of the insulin-like growth factor (IGF) family and shown recovery of function in failing mouse hearts along with the mobilization of resident progenitor cells. We take advantage of the natural repair capacity of the heart by providing a microrod scaffold (MRS) to not only deliver the native, rapidly degradable MGF, but also provide local mechanical and topographic cues necessary for proper cellular connectivity and differentiation. Our overall hypothesis is that timed release of MGF and IGF-1 delivered locally by the MRS regenerates and strengthens the damaged myocardium without harmful side effects. This is tested on progenitor/stem cells and cardiac myocytes in culture and in animal models. Specific Aim 1 determines the effect of stiffness variation on cells grown in 3D microrod scaffolds. We optimize MRS characteristics for regulation of cell proliferation, lineage commitment, differentiation, contractile maturity and connectivity of stem cells and cardiac myocytes. Specific Aim 2 determines the effect of growth factor (GF)-loaded MRS in environmental conditions that mimic the normal and ischemic heart. We characterize encapsulation efficiency, GF biostability, and acellular release kinetics of the eluting MRS in vitro. We determine effects of GF release from MRS on proliferation and migration of progenitor/ stem and neonatal rat ventricular myocytes to establish changes in gene expression and cell survival under culture conditions that mimic the normal and ischemic heart. Specific Aim 3 determines the cellular, molecular and functional gains that occur at different stages following myocardial infarction after MRS delivery of GFs to the border zone of an infarct. We examine how GF release affects the migration and differentiation of cardiac progenitor cells in vivo. We examine the beneficial effects of localized MGF peptide delivery on cardiac function, prevention of cardiac myocyte apoptosis, prevention of adverse cardiac remodeling, and reduced scar formation. Our long-term goal is to develop microrod MGF therapy that supports the regeneration of cardiac muscle to regain cardiac function in the failing human heart. Public Health Relevance Statement (provided by applicant): Heart failure is a common condition carrying a high burden of disability and mortality. Current estimates are that heart failure accounts for approximately one million hospitalizations and $10 billion in health care costs in the United States per year. The main underlying causes of heart failure are ischemic heart disease, hypertension and idiopathic dilated cardiomyopathy, whose common pathophysiologic characteristic is an inadequate mass of functional myocytes. This proposal develops and tests a novel therapeutic 3D eluting microrod scaffold (MRS) system to deliver a natural cardiac growth factor for aiding in the regeneration and recovery of damaged cardiac muscle in culture and animal models.