Cytoplasmic Glycosylation and Hypovolemic Stress

描述(由申请人提供)： 这里要检验的假设是，创伤后增加心肌细胞核质蛋白糖基化的干预措施可以减少细胞损伤，否则会导致低血容量性循环衰竭。我们的初步数据表明，在低血容量性应激大鼠模型中，注射氨基葡萄糖可显著改善创伤后功能。我们还证明了氨基葡萄糖在缺血/再灌注和钙超载的离体心模型中具有保护作用。我们认为，这种保护是由于通过己糖胺生物合成途径(HBP)增加通量而触发的自然的、应激激活的有利于生存的途径的放大。之所以能够将HBP主要产物UDP-GIcNAc的增加转化为复杂的细胞反应，是因为O-GIcNAc转移酶(OGT)被激活，并随着UDP-GIcNAc水平的增加识别不同的蛋白质。热休克蛋白(HSP)-70作为高压氧诱导的OGT蛋白底物的鉴定以及HSP与其他蛋白的结合部分是基于它们与O-GIcNAc结合的能力 处于细胞对应激反应的中心的通路。为了支持这一核心作用， O-GIcNAc在细胞保护中的作用，现在已经很清楚，各种压力都会导致UDP-GIcNAc和与蛋白质相关的O-GIcNAc水平的增加。我们建议，放大和/或加速O-GIcNAc升高的干预措施可以极大地减少应激，特别是低血容量性应激造成的损害。我们的具体目标是：通过增加UDP-GIcNAc的干预措施，优化低血容量性应激大鼠模型的恢复，并测试这些干预措施在猪身上的有效性；测试以下假设：增加UDP-GIcNAc导致其改善效果的机制是由于O-GIcNAc对核质蛋白的影响增加；记录并鉴定在低血容血症大鼠模型中，当保护措施得到优化时，心脏中优先使用O-GIcNAc衍生的核质蛋白；研究随UDP-GIcNAc增加而出现的钙内流减少在调节改善的恢复中的作用。

DESCRIPTION (provided by applicant): The hypothesis to be tested here is that interventions following trauma that increase the glycosylation of nucleocytoplasmic proteins in cardiomyocytes decrease the cellular damage that would otherwise contribute to hypovolemic circulatory collapse. Our preliminary data demonstrate that in a rat model of hypovolemic stress, the infusion of glucosamine leads to a striking improvement in post-trauma function. We also demonstrate that glucosamine is protective in isolated heart models of ischemia/reperfusion and calcium overload. We propose that this protection results from an amplification of a natural, stress-activated, pro-survival pathway triggered by increased flux through the hexosamine biosynthesis pathway (HBP). The capacity to transduce an increase in the HBP's primary product, UDP-GIcNAc, into complex cellular responses comes about because O-GIcNAc transferase (OGT) is activated and recognizes distinct proteins as levels of UDP-GIcNAc increase. The identification of heat shock protein (HSP)-70 as an HBP-induced protein substrate for the OGT and the finding that HSP's associate with other proteins in part based on their ability to bind O-GIcNAc position this pathway at the center of a primary cellular response to stress. In support of this central role for O-GIcNAc in cellular protection, it is now clear that various stresses lead to increases in levels of both UDP-GIcNAc and protein-associated O-GIcNAc. We suggest that interventions that amplify and/or accelerate increases in O-GIcNAc greatly decrease the damage resulting from stress, particularly hypovolemic stress. Our Specific Aims are: to optimize recovery in the rat model of hypovolemic stress through interventions that increase UDP-GIcNAc and to test the efficacy of these interventions in swine; to test the hypothesis that the mechanism by which increased UDP-GIcNAc leads to its ameliorating effects is because of an increase in O-GIcNAc on nuclear and cytoplasmic proteins; to document and identify the nucleocytoplasmic proteins in the heart that are preferentially derivatized with O-GIcNAc when protection is optimized in the hypovolemic rat model; to investigate the decrease in calcium influx seen with increased UDP-GIcNAc in mediating improved recoveries.