APOPTOSIS OF CARDIAC MUSCLE AS A TARGET FOR GENE THERAPY

程序性细胞死亡（凋亡）越来越被认为是一种 缺血/再灌注心肌细胞丢失的原因 损伤、心肌梗死和长期心力衰竭。 而 在这些环境中启动细胞凋亡的分子机制仍然存在， 未经证实，已经提出Bcl-2家族成员，包括 Bcl-2本身和Bax是细胞凋亡的一般调节因子， 神经酰胺途径、DNA损伤和其他各种原因。 使用 腺病毒基因转移到体外心肌细胞，研究者 证实了Bcl-2的抗凋亡作用， 腺病毒E1 B（Bax的抑制剂），在这个细胞背景。 其他 已建立的手段，以覆盖凋亡信号，至少在培养的 哺乳动物细胞包括：白细胞介素-1 β转化抑制剂 酶（ICE）/CED-3家族蛋白酶;细胞因子受体突变 Fas/APO-1及相关死亡结构域蛋白;哺乳动物细胞凋亡家族 或凋亡蛋白的杆状病毒抑制剂（IAP）;和显性- 肝白血病因子（HLF）的阴性形式，一种哺乳动物同源物 对于转录因子CES-2，保守的细胞死亡规范 秀丽隐杆线虫中的蛋白质。 然而，目前很少或 关于保护哺乳动物的潜力， 心室肌细胞凋亡，通过一个或多个这些 机制等 在目前的建议中，腺病毒基因转移到成人， 小鼠心肌将用于测试以下假设： (1)Bcl-2和Bcl-Xl，（2）ICE/CED-3蛋白酶抑制剂，（3）显性- 干扰死亡结构域蛋白，（4）IAP，和（5）显性- 干扰CES-2转录因子将保护免受 体内心脏凋亡。 心脏限制性转基因将成为 并行使用。 因此，这项建议将系统地测试基因 转移以保护冠状动脉结扎后的细胞凋亡， 五种互补的蛋白质， 细胞凋亡，至少在体外。 迄今为止，已知只有E1 B和Bcl-2 抑制心肌细胞凋亡，即使在培养中。 鉴于 细胞凋亡作为心肌死亡原因的证据程度， 特别是在梗塞和再灌注损伤的急性情况下， 迫切需要进行研究， 在这些环境中驱动细胞凋亡的途径， 干预以阻止体内凋亡途径。

Programmed cell death (apoptosis) is recognized, increasingly, as a contributing cause of cardiac myocyte loss with ischemia/reperfusion injury, myocardial infarction, and long-standing heart failure. While the molecular mechanisms initiating apoptosis in these settings remain unproven, it has been suggested that Bcl-2 family members including Bcl-2 itself and Bax are general regulators of apoptosis arising from the ceramide pathway, DNA damage, and other diverse causes. Using adenoviral gene transfer to cardiac myocytes in vitro, the Investigator has substantiated the predicted anti-apopotic effect of Bcl-2 and adenoviral E1B (an inhibitor of Bax), in this cell background. Other established means to override apoptotic signals at least in cultured mammalian cells include: inhibitors of interleukin-1beta converting enzyme (ICE)/CED-3 family proteases; mutations of the cytokine receptor Fas/APO-1 and related death domain proteins; the family of mammalian or baculovirus inhibitor of Apoptosis Proteins (IAPs); and a dominant- negative form of hepatic leukemia factor (HLF), a mammalian homologue for the transcription factor CES-2, a conserved cell death specification protein in Caenorhabditis elegans. At present, however, little or nothing is known concerning the potential to protect mammalian ventricular muscle from apoptosis, through one or more of these mechanisms. In the present proposal, adenoviral gene transfer to adult mouse myocardium will be used to test the hypotheses that delivery of: (1) Bcl-2 and Bcl-Xl, (2) ICE/CED-3 protease inhibitors, (3) dominant- interfering death domain proteins, (4) IAPs, and (5) dominant- interfering CES-2 transcription factors will be protective against cardiac apoptosis in vivo. Cardiac-restricted transgenes will be utilized in parallel. Thus, this proposal will systematically test gene transfer for protection from apoptosis after coronary artery ligation, for five complementary classes of protein with proven ability to inhibit apoptosis, at least in vitro. To date, only E1B and Bcl-2 are known to inhibit apoptosis of cardiac myocytes, even in culture. Given the extent of evidence for apoptosis as a cause of cardiac muscle death, especially in the acute settings of infarction and reperfusion injury, there is a compelling need for research that would define the molecular pathways driving apoptosis in these settings and for potential interventions to arrest the apoptotic pathway in vivo.