IDENTIFYING ROADBLOCKS TO LIMB REGENERATION

项目概要（摘要） 人类天生的肢体再生能力极其有限。因受伤或疾病而丧失肢体 是一个严重的健康问题。目前约有200万美国人生活在肢体丧失的后果中， 这一数字预计将上升，因为糖尿病等主要危险因素的流行率增加， 影响脉管系统的疾病。截肢的后果是深刻的病人，大多数必须 依靠的是并不完美的假肢再生医学的方法可能有一天是可行的，如果它是 了解如何自然地实现全肢体置换。为了获得这种理解，我们 使用动物模型，蝾螈蝾螈，它可以完全再生肢体， 即使是成年人也要截肢蝾螈的四肢在解剖学上与人类的四肢相似，它们最初的发育 也很相似因此，他们提供了一个蓝图，如何一个复杂的，三维的肢体可以再生， 在截肢后功能性地整合到现有的残肢中。必须解决的关键问题， 范例是要翻译到人类论坛是如何axolotls激活和培养的祖先， 新的肢体此外，细胞和分子的力量，可能拮抗成功的再生 也必须理解为这些可能通常存在于人类患者中，从而阻止再生。 未来的研究可能会阐明指导这些事件的分子和细胞力量是否不是 在哺乳动物中激活，或者它们是否过早终止，或者它们是否被其他 因素方法是首先彻底了解肢体是如何再生的，然后再使用这个方法。 这些信息可以为未来可能的治疗方法提供假设。 在这个提议中，我们利用了我们最近的发现，即蝾螈的能力可能会受到损害， 在反复截肢后再生肢体。这一发现提供了一个独特的机会， 这可能会限制再生或拮抗它。我们将研究祖细胞的激活 在连续截肢后，以确定这些细胞是否在再生失败中耗尽。我们还将 考虑巨噬细胞和肌成纤维细胞在重复截肢后再生失败中的作用。我们 我们将测试我们发现的再生限制是否在局部水平上运作，在肢体本身和接近肢体的地方。 截肢部位，或者如果它们更系统地作用，则在身体的其他部位。最后，我们将调查 两个基因的活动，其表达变得失调后，反复截肢，双调蛋白 和眼睛缺失2，两者都具有人类相关性。这项研究将利用机会 我们的新模型提出，希望增加对再生限制的理解， 这对未来患者的再生医学方法至关重要。

PROJECT SUMMARY (ABSTRACT) Humans have exceedingly limited natural limb regenerative abilities. Limb loss due to injury or disease is a major health problem. About two million Americans currently live with the consequences of limb loss, and this number is expected to rise because of increased prevalence of key risk factors such as diabetes and other diseases that affect vasculature. The consequences of amputation are profound for patients and most must rely on prosthetics, which are not perfect. A regenerative medicine approach may one day be feasible if it were understood how total limb replacement can be naturally achieved. To gain this understanding, we are employing an animal model, the axolotl salamander, which can completely regenerate limbs following amputation, even as adults. Axolotl limbs are anatomically similar to human limbs, and their initial development is similar as well. Thus, they offer a blueprint for how a complex, three-dimensional limb can be regrown and functionally integrated into the existing stump following amputation. Key issues that must be resolved if this paradigm is to be translated into the human forum are how axolotls activate and cultivate the progenitors for the new limb. Additionally, the cellular and molecular forces that might antagonize successful regeneration must also be understood as these might normally exist in human patients and thereby prevent regeneration. Future research could elucidate whether the molecular and cellular forces guiding these events are not activated in mammals, or whether they terminate prematurely, or whether they are overtly blocked by other factors. The approach is to first thoroughly understand how limbs do regenerate, and then later use this information to develop hypotheses for future possible therapies. In this proposal, we leverage our recent finding that axolotls can be compromised in their ability to regenerate limbs following repeated amputation. This finding presents a unique opportunity to identify factors that may be limiting in regeneration or may antagonize it. We will examine activation of progenitor cells following successive amputations to determine if these cells are exhausted in regenerative failure. We will also consider the role of macrophages and myofibroblasts in regenerative failure following repeated amputation. We will test if the regenerative limitations we uncovered operate at a local level, within the limb itself and close to the site of amputation, or if they act more systemically, elsewhere in the body. Finally, we will investigate the activities of two genes whose expression becomes dysregulated following repeated amputation, amphiregulin and eyes absent 2, both of which have human correlates. This research will capitalize on the opportunities presented by our new model with the hope that increased understanding of regenerative limitations will be essential for future regenerative medicine approaches in patients.