Cardiac regeneration in situ: Is it possible?

描述（由申请人提供）：修复受损心脏的策略主要集中在输注、直接注射或移植各种类型的干细胞/祖细胞。我们最近报道了心肌细胞中GSK-3b的条件缺失导致心肌细胞增殖。此外，我们发现，在胸主动脉收缩（产生左心室收缩压显著增加）或心肌梗死（MI）引起的应激环境下，增殖程度被放大。事实上，我们已经发现，GSK-3b的缺失（或抑制）导致小鼠胚胎干细胞衍生的胚胎样体、培养的新生大鼠心室肌细胞、发育中的心脏心肌细胞、正常心脏的心肌细胞，以及，如上所述，特别是应激心脏的心肌细胞的增殖。如果这种增殖足够强劲，就可以诱导有意义的再生，而不需要移植（即原位再生）。我们将以三个具体目标来解决这一假设。我们将首先通过量化正常心脏和应激心脏的增殖率和心肌细胞质量来确定GSK-3b缺失导致的心肌细胞增殖有多强。然后，我们将通过确定是否对左心室功能有有益影响来检查这种增殖的生理后果。最后，我们将确定GSK-3b的缺失是否会对中晚期心力衰竭小鼠产生有益的影响——总结晚期心力衰竭患者的临床情况，对于这些患者，除了心脏移植或目的地LVAD治疗之外，治疗选择很少。在“未来方向”一节中，我们还讨论了GSK-3b在驱动未成熟心肌细胞向成熟心肌细胞分化中所起的关键作用。我们建议在增殖反应产生新的肌细胞后，通过腺相关病毒基因转移到心脏来恢复GSK-3b的表达。因此，任何因GSK-3b缺失而形成的未成熟肌细胞都将分化为成熟的、功能齐全的肌细胞，从而最大限度地改善左室功能。我们相信，这一申请完全符合R21机制的目标，即支持突破性研究，有可能显著推进心力衰竭患者的治疗选择——这种疾病多年来没有新的重要药物上市。

DESCRIPTION (provided by applicant): Strategies to repair injured hearts have predominantly focused on infusion, direct injection, or transplantation of any of a variety of types of stem/progenitor cells. We have recently reported that conditional deletion of GSK-3b specifically in cardiomyocytes leads to proliferation of cardiomyocytes. Furthermore, we find that the degree of proliferation is amplified in the setting of stress induced by thoracic aortic constriction (producing marked increases in left ventricular systolic pressure) or myocardial infarction (MI). In fact, we have found that deletion (or inhibition) of GSK-3b leads to proliferation of cardiomyocytes in mouse embryonic stem cell-derived embryoid bodies, in neonatal rat ventricular myocytes in culture, in cardiomyocytes in the developing heart, in cardiomyocytes in the normal heart, and, as noted above, particularly in cardiomyocytes in the stressed heart. If this proliferation is sufficiently robust, one could induce meaningful regeneration without the need for transplantation (i.e. regeneration in situ). We will address this hypothesis with three Specific Aims. We will first determine how robust is the cardiomyocyte proliferation resulting from deletion of GSK-3b by quantifying proliferation rates and cardiomyocyte mass in both the normal heart and in the stressed heart. We will then examine the physiologic consequences of this proliferation by determining whether there is a beneficial effect on left ventricular function. Finally, we will determine whether deletion of GSK-3b can lead to beneficial effects in mice with moderately advanced heart failure- recapitulating the clinical scenario of patients with advanced heart failure for whom there are very few treatment options besides heart transplant or destination LVAD therapy. In a "future directions" section, we also discuss the critical role played by GSK-3b in driving differentiation of immature cardiomyocytes into mature cardiomyocytes. We propose studies to restore expression of GSK-3b via adeno-associated virus gene transfer to the heart after the proliferative response has created new myocytes. Thus any immature myocytes formed in response to the deletion of GSK-3b will be differentiated into mature, fully functional myocytes, maximizing the chances of improving LV function. We believe that this application perfectly matches the goals of the R21 mechanism of supporting groundbreaking research with the potential to significantly advance therapeutic options for heart failure patients- a disease that has had no new important classes of agents come to market in many years. PUBLIC HEALTH RELEVANCE: Herein we propose to determine whether it is possible to drive cardiomyocyte proliferation sufficiently to result in cardiac regeneration in situ (i.e. without the need to utilize exogenous stem/progenitor cells). Based on our extensive studies in mouse models, we hypothesize that by manipulating a single signaling pathway- the glycogen synthase kinase-3b (GSK-3b) signaling pathway- in vivo, we will indeed be able to achieve regeneration. If we can achieve this, it would be a significant step forward in the treatment of patients with end- stage heart diseases of many etiologies.