The Mechanistic Basis of Cyclin A2-Mediated Cardiac Repair

描述（由申请人提供）：刺激心肌细胞有丝分裂的概念对心脏再生以及心血管疾病具有显着影响。我们之前首次证明，如果细胞周期蛋白 A2 在转基因小鼠模型的心肌细胞中组成型表达，哺乳动物心肌细胞有丝分裂可以在出生后发生。我们最近的结果表明，在诱导心肌梗塞(M1)后，细胞周期蛋白A2介导转基因心脏中的心肌细胞有丝分裂，从而与非转基因同窝对照相比，修复心脏并显着恢复心脏功能。为了探索这些结果的潜在临床转化，我们通过灭活的腺病毒将编码细胞周期蛋白 A2 的 cDNA 给予梗塞大鼠，并注意到接受腺细胞周期蛋白 A2 的大鼠在注射部位发生细胞再生，并且与接受空腺病毒的对照组相比，保留了心输出量。我们假设有两种潜在的机制介导我们观察到的再生。首先，在我们的小鼠梗塞模型中，侧群（SP）干细胞似乎分化为心肌细胞，但这些未成熟心肌细胞中的有丝分裂仅在转基因小鼠中被发现，因此可能揭示了诱导祖细胞过度增殖的新机制。其次，我们注意到与非转基因对照相比，转基因小鼠梗塞周围区域磷酸组蛋白 H3 的表达和 BrdU 的摄取显着增加，这表明成年心肌细胞的去分化和分裂。利用来自转基因和非转基因小鼠的SP细胞，我们建议通过SP细胞衍生的祖细胞增殖的体外研究和SP细胞移植到小鼠梗塞模型的体内研究来探索第一个机制。此外，我们将使用新型工程心脏组织，通过测试转基因和非转基因小鼠的 SP 细胞来源的祖细胞和成年心肌细胞对损伤的反应来评估这两种机制。第二种机制也将通过将细胞周期蛋白 A2 cDNA 病毒转移到体外成体心肌细胞中并利用视频显微镜监测这些细胞的有丝分裂和胞质分裂来进行检查。提出这些研究是为了阐明更直接的临床治疗途径。

DESCRIPTION (provided by applicant): The concept of stimulating cardiomyocyte mitotic divisions has dramatic implications for cardiac regeneration and hence, cardiovascular disease. We previously demonstrated for the first time that mammalian cardiomyocyte mitoses can occur postnatally if cyclin A2 is constitutively expressed in cardiomyocytes in a transgenic mouse model. Our more recent results demonstrate that cyclin A2 mediates cardiomyocyte mitoses in the transgenic hearts after myocardial infarction (Ml) is induced, thus repairing the heart with significant recovery of cardiac function when compared with nontransgenic littermate controls. To explore potential clinical translation of these results, we administered cDNA encoding cyclin A2 via de-activated adenovirus to infarcted rats and noted that the rats receiving adeno-cyclin A2 undergo cellular regeneration at the sites of injection and have preserved cardiac output when compared to controls which received empty adenovirus. We hypothesize that there are two potential mechanisms mediating the regeneration we have observed. First, side population (SP) stem cells appear to differentiate into cardiomyocytes in our mouse infarction model, but mitoses in these immature cardiomyocytes are only noted in the transgenic mice, thus possibly uncovering a novel mechanism to induce hyperproliferative progenitor cells. Second, we note a significant increase in expression of phosphohistone H3 and uptake of BrdU in the peri-infarct zone of transgenic mice compared to nontransgenic controls, implicating dedifferentiation and division of adult cardiomyocytes. Utilizing SP cells from transgenic and nontransgenic mice, we propose to explore the first mechanism via in vitro studies of proliferation of SP cell-derived progenitor cells and in vivo studies of SP cell transplantation into a mouse infarction model. Additionally, we will use novel engineered cardiac tissues to evaluate both mechanisms by testing the response to injury of both SP cell-derived progenitors and adult cardiomyocytes from transgenic and nontransgenic mice. The second mechanism will also be examined using viral transfer of cyclin A2 cDNA into adult cardiomyocytes in vitro and the utilization of video microscopy to monitor mitosis and cytokinesis of these cells. These studies are proposed in order to elucidate a more directed path to clinical therapy.