Creation of Stem Cells by Nuclear Reprogramming

描述（由申请人提供）：每年约有90万美国人患有心肌梗死。这些患者中约有225,000人死亡，其他人有慢性并发症的风险，包括充血性心力衰竭和顽固性室性心律失常。与肝脏等其他器官不同，心脏在很大程度上无法再生和替代失去的心肌细胞。现在人们正在努力寻找诱导、刺激和增强心肌再生的方法。一种有希望的方法是将未分化的干细胞移植到受损区域。干细胞移植的一个瓶颈是获得足够数量的干细胞注入患者体内。人们对使用胚胎干细胞（ES细胞）衍生的供体细胞很感兴趣。由于对破坏人类胚胎的伦理担忧，很多注意力都集中在通过其他方式获得胚胎干细胞上。一种可能的解决方案是重新编程患者自身的体细胞，使其变得更像干细胞。细胞核重编程：使终末分化细胞的细胞核变得更像未分化干细胞的细胞核的过程。本实验的目的是用爪蟾卵的提取物对分化的细胞进行重编程。我们的方法是将渗透细胞暴露于浓缩的卵提取物中，然后通过将其移植到去核的爪蟾卵中来检测重编程的程度。与接受未经处理的细胞核相比，接受提取物处理的细胞核的卵子发育改善，将表明重编程。如果我们可以通过提取物证明重编程，那么我们就可以通过用抑制剂处理提取物或通过免疫消耗消除特定蛋白质来研究各种重编程模型。它甚至有可能纯化具有重编程活性的蛋白质复合物。我们的研究结果将揭示驱动细胞分化的表观遗传变化的本质。我们还希望开发出能够有效地重编程分化细胞的提取物，以便它们可以用于细胞替代疗法。公共卫生相关性：心脏病幸存者有发展为心力衰竭的风险，因为心脏不能像其他器官一样自我修复和再生。近年来，心脏病患者一直在接受干细胞治疗，希望这些干细胞能够转化为心肌细胞，恢复心脏的泵血功能。这项提议的目的是用卵细胞提取物治疗正常细胞，看看它们能否转化为干细胞，以期用于修复心脏。

DESCRIPTION (provided by applicant): Each year approximately 900,000 Americans suffer a myocardial infarction. Approximately 225,000 of these victims die and the others are at risk for chronic complications, including congestive heart failure and intractable ventricular arrhythmias. Unlike other organs such as the liver, the heart is unable to regenerate and replace the lost myocardial cells to any significant extent. Much effort is now being expended to find ways to induce, stimulate, and augment myocardial regeneration. One promising approach is transplantation of undifferentiated stem cells into the infracted zone. One bottleneck in stem cell transplantation is the acquisition of a sufficient number of stem cells to be infused into the patient. There is much interest in using donor cells that are derivatives of embryonic stem cells (ES cells). Because of ethical concerns about destroying human embryos, much attention has been focused on obtaining ES cells by other means. One potential solution is to reprogram a patient's own somatic cells so that they become more like stem cells. Nuclear reprogramming the process by which the nucleus of terminally differentiated cells becomes more like the nucleus of an undifferentiated stem cell. The goal of the experiments in this proposal is to reprogram differentiated cells with extracts of Xenopus eggs. Our approach will be to expose permeabilized cells to concentrated egg extracts then assay the extent of reprogramming by transplanting them into enucleated Xenopus eggs. Reprogramming will be indicated by improved development of eggs that receive extract-treated nuclei compared to those that receive untreated nuclei. If we can demonstrate reprogramming by extracts, then we can investigate various reprogramming models by treating the extract with inhibitors or eliminating specific proteins by immunodepletion. It may even be possible to purify protein complexes with reprogramming activity. The results of our studies will shed light on the nature of the epigenetic changes that drive cell differentiation. We also hope to develop extracts that efficiently reprogram differentiated cells so that they can be used for cell replacement therapy. PUBLIC HEALTH RELEVANCE: Heart attack survivors are at risk for developing heart failure because the heart cannot repair and regenerate itself like some other organs. In recent years, heart attack victims have been treated with stem cells in the hope that these will be able to turn into heart muscle cells and restore the heart's pumping function. The purpose of this proposal is to treat normal cells with extracts of egg cells to see if they can be turned into stem cells in the hope that these can be used to repair the heart.