HORMONAL REGULATION OF NEONATAL MUSCLE PROTEIN SYNTHESIS

长期目标是确定激素和营养物质相互作用调节新生儿骨骼肌蛋白沉积率的机制，并确定这些机制如何被败血症改变。目标是确定新的策略，以优化营养利用的生长发育迟缓和败血症婴儿。骨骼肌质量的快速增长是新生儿生长的一个特别重要的特征。先前的研究表明，在正常新生儿中，喂养诱导的肌肉蛋白质合成刺激涉及胰岛素和氨基酸的调节。然而，由于正常新生儿肌肉蛋白合成对胰岛素和氨基酸的高度敏感性，无法辨别这两种合成代谢剂是否相互作用来调节肌肉蛋白合成。脓毒症对新生儿的发病率和死亡率有很大的影响，因为新生儿的肌肉蛋白储存有限。激素变化和底物缺乏可能导致败血症患者的肌肉萎缩。假设是：1)氨基酸增强了新生儿肌肉蛋白合成对胰岛素水平的敏感性；2)氨基酸和胰岛素的相互作用反映了它们对真核起始因子（eIFs）和信号激酶活性的独立作用；3)脓毒症通过肿瘤坏死因子α (TNFalpha)与胰岛素介导的蛋白合成的相互作用抑制新生儿肌肉蛋白合成，这种作用可以被氨基酸绕过。为了解决这些假设，我们提出了以下目标：在目标1中，将在正常新生儿中确定肌肉蛋白合成对胰岛素的剂量反应以及氨基酸对这种反应的影响。为了实现这一目标，研究将在7天大的猪中进行，使用肌肉蛋白质合成的直接动力学测量和新型激素底物夹紧技术的改进。在Aim 2中，胰岛素和氨基酸对关键eIFs活性的影响将被确定。为了实现这一目标，将使用Aim 1实验中的肌肉来确定蛋白质合成速率、参与Met-tRNA和mRNA与40S核糖体亚基结合的eif活性以及信号激酶激活之间的定量关系。这将确定体内胰岛素和氨基酸调节的特定翻译过程。在Aim 3中，肌肉蛋白合成对胰岛素在内毒素、TNFalpha、抗细胞因子治疗和增强氨基酸供应后的反应将被确定。这些研究将确定激素、细胞因子和营养物质在新生儿败血症引起的肌肉萎缩中的作用。氨基酸管理的实际效果，以避免脓毒症引起的肌肉蛋白合成胰岛素抵抗将被确定。

The long-term objectives are to identify the mechanisms by which hormones and nutrients interact to regulate the high rate of skeletal muscle protein deposition in the neonate, and to determine how these mechanisms are altered by sepsis. The goal is to identify new strategies to optimize nutrient utilization for growth in growth-retarded and septic infants. Rapid accretion of skeletal muscle mass is a particularly important characteristic of neonatal growth. Previous work has shown that the feeding-induced stimulation of muscle protein synthesis in normal neonates involves regulation by both insulin and amino acids. However, because of the exquisite sensitivity of muscle protein synthesis to insulin and amino acids in the normal neonate, it could not be discerned whether both anabolic agents interact to regulate muscle protein synthesis. Sepsis contributes substantially to the morbidity and mortality of neonates, a population with limited muscle protein stores. Hormonal changes and substrate deficiency likely contribute to the muscle wasting that occurs with sepsis. The hypotheses are: 1) amino acids enhance the sensitivity of muscle protein synthesis to insulin levels in neonates, 2) the interaction of amino acids and insulin reflects their independent effects on the activity of eukaryotic initiation factors (eIFs) and signaling kinases, and 3) sepsis inhibits muscle protein synthesis in neonates via an interaction of tumor necrosis factor alpha (TNFalpha) with insulin-mediated protein synthesis, an effect that can be circumvented by amino acids. The following aims are proposed to address these hypotheses: In Aim 1, the dose response of muscle protein synthesis to insulin and the effect of amino acids on this response will be determined in normal neonates. To achieve this, studies will be performed in 7-d-old pigs using direct kinetic measurement of muscle protein synthesis and refinements of a novel hormone-substrate clamp technique. In Aim 2, the effects of insulin and amino acids on the activity of key eIFs will be determined. To achieve this, muscles from the experiments in Aim 1 will be used to determine the quantitative relationships among protein synthesis rates, the activity of the eIFs that are involved in the binding of Met-tRNA and mRNA to the 40S ribosomal subunit, and the activation of signaling kinases. This will identify the specific translational processes regulated by insulin and amino acids in vivo. In Aim 3, the response of muscle protein synthesis to insulin following administration of endotoxin, TNFalpha, anti-cytokine therapy, and enhanced amino acid supply will be determined. These studies will identify the role of hormones, cytokines, and nutrients in the sepsis-induced muscle wasting in neonates. The practical efficacy of amino acid administration to circumvent the sepsis-induced insulin resistance of muscle protein synthesis will be identified.