Enhancing the effectiveness of human cardiac stem cell therapy

描述(由申请人提供)：缺血性心脏病仍然是美国最大的死亡原因。C-kit阳性的人心脏干细胞(HCSCs)移植是修复心肌梗死的一种非常有前景的治疗方法，但供体细胞存活率低严重限制了移植的效果。一氧化碳(CO)是血红素加氧酶-1(HO-1)的副产物，对心肌细胞的缺血/再灌注损伤具有明显的抗细胞凋亡和细胞保护作用。然而，目前尚不清楚HO-1/CO对人神经干细胞是否具有细胞保护作用。我们的前期研究发现，HO-1/CO在调节HCSC存活能力中起着重要作用。因此，本研究的应用重点在于验证我们的假设，即HO-1/CO通过激活存活信号通路和细胞因子效应促进人心脏干细胞存活，并增强移植到免疫缺陷小鼠心肌梗死后的干细胞治疗效果和心功能。因此，我们将检测经HO-1诱导剂(COPP)或CO释放分子(CORM-3)预处理的hCSCs在细胞存活、增殖、迁移、内皮和心肌分化方面表现出更强的能力(目标1)，并了解HO-1/CO对HCSC存活信号通路、抗凋亡、ROS产生和细胞因子释放的影响的分子机制(目标2)。本方案的目的3将在免疫缺陷小鼠(SCID)心肌梗死模型上测试移植预适应的EGFP+-hCSC是否能改善体内HCSC的存活和归巢、内源性小鼠CSC的增殖和分化以及心脏的结构和功能。上述实验结果将为提高HCSC移植后存活率的策略提供重要的启示，从而为临床修复心肌梗死、用于缺血性心脏病患者的细胞治疗提供重要的启示。 公共卫生相关性：供体细胞存活率低是干细胞治疗缺血性心脏病的主要挑战之一。本项目调查了有效的战略，以 使用小分子，包括钴原卟啉(CoPP)，一种促进内源性一氧化碳生成的血红素加氧酶1诱导剂，或一氧化碳供体CORM-3，提高人类心脏干细胞的治疗效果。这些研究将展示CoPP/CORM-3如何在体外细胞培养模型中影响人类心脏干细胞的存活能力，以及心肌内注射经CoPP/CORM-3处理的人心脏干细胞如何改善缺血再灌注损伤后体内免疫缺陷小鼠的心功能。这一结果和广泛的结论将直接关系到提高缺血性心脏病患者心肌梗死细胞治疗的临床疗效。

DESCRIPTION (provided by applicant): Ischemic heart disease remains as the single largest cause of mortality in the United States. Delivery of c-kit positive human cardiac stem cells (hCSCs) is a very promising therapeutic approach in repairing the infarcted heart, but is severely limited by the poor survival of donor cells. Carbon monoxide (CO), a byproduct of heme oxygenase 1 (HO-1), has a potent anti- apoptotic, cytoprotective effect against ischemia/reperfusion injury to cardiomyocytes. However, it is not known whether HO-1/CO is cytoprotective for hCSCs. Our pilot studies found that HO-1/CO plays an important role in mediating hCSC survival ability. Therefore, this application focuses on testing our hypothesis that HO-1/CO promotes human cardiac stem cell survival vial the activation of survival signal pathways and the cytokine effects, as well as enhances the stem cell therapy efficacy and heart function after transplantation into immunodeficient mouse heart following myocardial infarction. Thus, we will examine whether hCSCs pretreated with an HO-1 inducer (CoPP), or a CO releasing molecule (CORM-3), will exhibit greater abilities in cell survival, proliferation, migration, endothelial and cardiomyogenic differentiation (Aim 1), and understand the molecular mechanisms that underlie the effect of HO-1/CO on hCSC survival signal pathways, resistance to apoptosis, ROS generation and cytokine release (Aim 2). Aim 3 of this proposal will test whether transplantation of the preconditioned eGFP+-hCSCs will result in improvement in invivo hCSC survival and homing, endogenous mouse CSC proliferation and differentiation, and cardiac structure and function with immunodeficient mouse (SCID) model following myocardial infarction. Knowledge gained from the above experiments will provide the essential implications on the strategies to enhance hCSC survival after transplantation and, therefore, their efficacy in clinical repairing infarcted myocardium for the cell therapy of patients with ischemic heart disease. PUBLIC HEALTH RELEVANCE: Poor survival of donor cells is one of the major challenges in stem cell therapy for ischemic heart disease. This project investigates the effective strategies to enhance the human cardiac stem cell therapy efficacy with small molecules, including cobalt protoporphyrin (CoPP), a heme oxygenase 1 inducer, which promotes endogenous carbon monoxide generation; or a carbon monoxide donor, CORM-3. These studies will show how CoPP/CORM-3 affects the survival ability of human cardiac stem cells in an in vitro cell culture model, and how intramyocardial injection of CoPP/CORM-3 pretreated human cardiac stem cells improve heart function in an in vivo immunodeficient mouse model following ischemia-reperfusion injury. The results and broad conclusions will be directly relevant to enhance the efficacy in clinical repairing the infarcted myocardium for the cell therapy of patients with ischemic heart disease.