Heat shock proteins and disuse muscle atrophy

描述(由申请人提供)： 项目摘要/摘要骨骼肌废用性萎缩是一种广泛的生理现象，与制动、卧床休息、失神经和太空飞行有关，或任何负重活动的普遍减少。然而，我们对在停用过程中调节肌肉质量的信号分子的理解是模糊的。因此，我们研究计划的长期目标是了解在废用过程中导致肌肉萎缩的信号通路的调节。最终，增进了解将导致确定具体干预措施的目标。热休克蛋白(HSPs)是一类在细胞中结构性表达的蛋白质，但在各种细胞应激条件下进一步快速表达。这种诱导已被证明具有多种细胞保护功能。在肌肉停用过程中，热休克家族成员Hsp70的表达持续下调，在停用过程中Hsp70的过度表达消除了核因子-B和Foxo3a反式激活的增加，并完全防止骨骼肌萎缩。这一点很重要，因为废用性肌肉萎缩需要核因子-B和Foxo3a。然而，目前尚不清楚Hsp70的过度表达是否足以特异性地抑制NF：B或Foxo3a诱导的肌肉萎缩。Hsp70基因敲除是否足以导致骨骼肌萎缩也是未知的。本提案的目的是在目标1和目标2中确定Hsp70表达的增加是否足以抑制NF：B或Foxo3a诱导的肌纤维萎缩，以及在目标3中确定Hsp70的下调是否足以导致肌肉纤维萎缩。为了达到这些特定的目的，我们将向大鼠骨骼肌注射WT IKK2+Hsp70表达载体(Aim 1)、WT Foxo3a+Hsp70表达载体(Aim 2)或产生Hsp70特异性shRNAs的质粒(Aim 3)，并测量NF-：B或Foxo3a活性、特定萎缩基因的mRNA表达和肌纤维横截面积。如果Hsp70表达的增加抑制了NF-：B活性和/或Foxo3a活性，我们将通过确定Hsp70结合的每个途径中的蛋白质来确定其机制。这些实验的发现将使人们更好地理解HSP70在骨骼肌萎缩过程中对NF-：B和Foxo3a信号的调控。 公共卫生相关性： 项目叙述由于不使用而导致的骨骼肌萎缩与固定、卧床休息、去神经和太空飞行，或任何负重活动的普遍减少有关。在这项拟议的工作中，我们将从基因上过度表达或击倒一种蛋白质，这种蛋白质被认为可以调节肌肉大小，并在因不使用而导致的肌肉萎缩过程中被下调。这将使我们能够直接确定该蛋白参与调节骨骼肌质量，并可能确定该蛋白作为治疗肌肉萎缩的新靶点。

DESCRIPTION (provided by applicant): Project summary/Abstract Skeletal muscle disuse atrophy is a widespread physiological phenomenon associated with immobilization, bed rest, denervation, and space flight, or any general reduction in weight bearing activity. However, our understanding of the signaling molecules that regulate muscle mass during disuse are ill defined. Therefore the long-range goal of our research program is to understand the regulation of signaling pathways that cause muscle atrophy during disuse. Eventually improved understanding will lead to the identification of targets for specific interventions. Heat shock proteins (Hsps) are a family of proteins that are constitutively expressed in cells, but whose expression is further, and rapidly, induced by a variety of cellular stresses. This induction has been shown to provide a variety of cytoprotective functions. During muscle disuse a member of the heat shock family, Hsp70, is consistently down-regulated and overexpression of Hsp70 during disuse abolishes the increase in NF-:B and Foxo3a transactivation, and completely prevents skeletal muscle atrophy. This is important since NF-:B and Foxo3a are required for disuse muscle atrophy. However, it is currently unknown whether Hsp70 overexpression is sufficient to specifically inhibit NF-:B-induced or Foxo3a-induced muscle atrophy. It is also unknown whether knock down of Hsp70 is sufficient to cause skeletal muscle atrophy. The objective of the current proposal is to determine in Aims 1 and 2 if an increase in Hsp70 expression is sufficient to inhibit NF-:B-induced or Foxo3a-induced muscle fiber atrophy, and in Aim 3 if knock down of Hsp70 is sufficient to cause muscle fiber atrophy. To address these specific aims we will inject WT IKK2 plus Hsp70 expression plasmids (Aim 1), WT Foxo3a plus Hsp70 expression plasmids (Aim 2), or a plasmid producing shRNAs specific for Hsp70 (Aim 3) into the skeletal muscle of rats and measure NF-:B or Foxo3a activity, the mRNA expression of specific atrophy genes and muscle fiber cross sectional area. If an increase in Hsp70 expression inhibits NF-:B activity and/or Foxo3a activity we will determine the mechanisms of this by determining the proteins in each pathway that Hsp70 binds. The findings from these experiments will lead to a greater understanding of Hsp70 in the regulation of NF-:B and Foxo3a signaling during skeletal muscle atrophy. PUBLIC HEALTH RELEVANCE: Project Narrative Skeletal muscle wasting due to disuse is associated with immobilization, bed rest, denervation, and space flight, or any general reduction in weight bearing activity. In the proposed work we will genetically overexpress or knock down a protein that is believed to regulate muscle size and is known to be down-regulated during muscle wasting caused by disuse. This will allow us to directly determine the involvement of this protein in the regulation of skeletal muscle mass, and could identify the protein as a novel therapeutic target for muscle wasting.