BIOENERGETIC REGULATION OF CARDIAC PROGENITOR CELLS

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 糖酵解和线粒体能量学的重组被认为是干细胞分化所必需的，这支持了表型指定需要遗传回路和发育生物能量学的协调的观点。这意味着代谢性疾病，如糖尿病，可能对心脏祖细胞(CPC)具有特别的破坏性，从而限制它们在病变心肌环境中分化和耐受压力的能力。我们计划研究有丝分裂原在调节CPC分化所需的糖酵解和线粒体能量学变化中的作用。一般的假设是，在糖尿病中，将有丝分裂原信号与能量学联系起来的发育信号被破坏，从而阻止了CPC向心肌细胞的分化。 具体目标是： 1.检测有丝分裂原对CPC能量的影响。由于胰岛素和IGF-1等有丝分裂原在CPC归巢和分化中起重要作用，我们将研究这些有丝分裂原的存在和缺失对糖酵解通量和线粒体生物能量学的影响。将检测胰岛素抵抗对能量代谢的影响，并通过共聚焦显微镜和流式细胞仪监测心肌细胞分化。 2.确定糖酵解如何调节CPC的生长和分化。利用药理学和分子干预手段(如康宁酸和PFK-2基因敲除或过表达)，我们将在模拟正常和糖尿病表型的底物条件下，研究糖酵解在CPC生长和分化中的作用。能量学将通过细胞外流量分析进行检测，向心脏谱系的分化将通过GFP-NKX2.5的表达来衡量。 3.评估再植入具有能量能力的CPC是否会提高其治疗效果。具有高糖酵解和/或线粒体能力的CPC将被重新植入梗死的心脏。心脏功能将通过超声心动图进行评估，植入将在心脏组织切片上通过共聚焦显微镜进行分析。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. A restructuring of glycolysis and mitochondrial energetics is thought to be required for stem cell differentiation, supporting the view that phenotype specification requires the coordination of genetic circuits and developmental bioenergetics. This implies that metabolic diseases such as diabetes may be particularly destructive to cardiac progenitor cells (CPCs), thereby limiting their ability to differentiate and tolerate stress in the milieu of the diseased myocardium. We plan to examine the role of mitogens in mediating the changes in glycolysis and mitochondrial energetics required for CPC differentiation. The general hypothesis is that the developmental cues that relate mitogen signaling with energetics are disrupted in diabetes, thereby preventing differentiation of CPCs into cardiomyocytes. The specific aims are to: 1. Examine the effects of mitogens on CPC energetics. Because mitogens such as insulin and IGF-1 play important roles in CPC homing and differentiation, we will examine how the absence and presence of these mitogens affect glycolytic flux and mitochondrial bioenergetics. The effect of insulin resistance on energetics will be examined, and cardiomyocyte differentiation will be monitored by confocal microscopy and flow cytometry. 2. Determine how glycolysis regulates CPC growth and differentiation. Using pharmacological and molecular interventions (e.g., koningic acid and PFK-2 knockdown or overexpression), we will examine the role of glycolysis in CPC growth and differentiation under substrate conditions that mimic the normal and diabetic phenotype. Energetics will be examined by extracellular flux analysis and differentiation into the cardiac lineage will be measured by expression of GFP-Nkx2.5. 3. Assess whether reimplantation of energetically competent CPCs increases their therapeutic efficacy. CPCs with high glycolytic and/or mitochondrial capacities will be reimplanted in the infarcted heart. Myocardial function will be evaluated by echocardiography and engraftment will be analyzed by confocal microscopy in heart tissue sections.