Cardioprotection with nuclear-targeted Akt/PKB

心力衰竭与导致收缩力丧失的心肌细胞死亡有关。先前的研究使用膜靶向 Akt（肉豆蔻酸化-Akt）（一种参与抗凋亡信号转导的酶）显示出抑制细胞死亡并预防心肌病刺激诱导的发病机制。然而，我们小组最近的研究发现激活的 Akt 在细胞核中积累，表明 Akt 活性的生物学相关靶点可能位于那里。为了检验这一假设，创建了一个靶向 Akt 构建体来确定核 Akt 积累的抗凋亡作用。通过共聚焦显微镜证实了腺病毒编码的 Akt 构建体的核定位。表达核靶向 Akt 的心肌细胞没有表现出形态重塑的证据，例如肌原纤维密度改变或肥大。在缺氧诱导的细胞死亡中，核靶向 Akt 显着提高磷酸 Akt 和激酶活性的水平，并与肉豆蔻酸化 Akt 一样有效地抑制细胞凋亡。核靶向 Akt 的转基因过表达不会导致肥厚性重塑、心肌细胞 DNA 含量或核改变、或典型细胞质 Akt 底物的磷酸化增强，但转基因心脏免受缺血再灌注损伤。基因阵列分析表明，与非转基因对照相比，Akt/nuc 心脏的转录谱发生了变化，这与转基因心脏中 Akt 表达的先前特征不同。总的来说，这些实验表明，Akt 靶向细胞核可介导细胞凋亡的抑制，而不会发生肥大性重塑，这为核靶向 Akt 抑制与心脏病相关的细胞死亡的治疗应用开辟了新的可能性。

Heart failure is associated with death of cardiomyocytes leading to loss of contractility. Previous studies using membrane-targeted Akt (myristolated-Akt), an enzyme involved in antiapoptotic signaling, showed inhibition of cell death and prevention of pathogenesis induced by cardiomyopathic stimuli. However, recent studies by our group have found accumulation of activated Akt in the nucleus, suggesting that biologically relevant target(s) of Akt activity may be located there. To test this hypothesis, a targeted Akt construct was created to determine the antiapoptotic action of nuclear Akt accumulation. Nuclear localization of the adenovirally encoded Akt construct was confirmed by confocal microscopy. Cardiomyocytes expressing nuclear-targeted Akt showed no evidence of morphological remodeling such as altered myofibril density or hypertrophy. Nuclear-targeted Akt significantly elevated levels of phospho-Akt and kinase activity and inhibited apoptosis as effectively as myristolated-Akt in hypoxia-induced cell death. Transgenic overexpression of nuclear-targeted Akt did not result in hypertrophic remodeling, altered cardiomyocyte DNA content or nucleation, or enhanced phosphorylation of typical cytoplasmic Akt substrates, yet transgenic hearts were protected from ischemia-reperfusion injury. Gene array analyses demonstrated changes in the transcriptional profile of Akt/nuc hearts compared with nontransgenic controls distinct from prior characterizations of Akt expression in transgenic hearts. Collectively, these experiments show that targeting of Akt to the nucleus mediates inhibition of apoptosis without hypertrophic remodeling, opening new possibilities for therapeutic applications of nuclear-targeted Akt to inhibit cell death associated with heart disease.