Attenuation of denervation atrophy by nandrolone: molecular mechanisms

描述（由申请人提供）： 雄激素类固醇通过尚不清楚的分子机制减少肌肉萎缩。在研究雄激素类固醇诺龙对神经横断瘫痪肌肉萎缩的影响时，我们发现，当在神经横断后29天（35天）开始使用诺龙时，萎缩率在开始诺龙后7天内降低，这与肌肉泛素连接酶MAFbx和MuRF 1的表达降低有关。相比之下，当在神经切断时开始使用诺龙时，7天后（7天）它并没有减缓萎缩或减少MAFbx或MuRF 1的表达。我们认为，该模型提供了一个独特的机会，以了解诺龙减缓肌肉萎缩的分子机制，并了解为什么肌肉在某些条件下对诺龙的有益作用有抵抗力。有趣的是，基因分析显示，在35天而不是7天受到诺龙影响的基因中，有可能调节肌肉大小的基因是FOXO 1和RCAN 2。RCAN 2在骨骼肌中以高水平表达，在骨骼肌中它通过激活转录因子NFAT抑制钙调神经磷酸酶，钙调神经磷酸酶是一种参与肌肉肥大的蛋白磷酸酶。感兴趣的是，FOXO 1过表达也可能通过上调MAFbx降低钙调磷酸酶活性[5-7]，表明诺龙诱导的FOXO 1和MAFbx水平降低可能代表第二种平行机制，诺龙可通过该机制增加失神经肌肉中的钙调磷酸酶活性。 失神经肌肉诺龙的不敏感性在早期的时间点最有可能反映的转录调节，阻断诺龙的行动，或下调的一个是必要的it. We发现在表达水平的几个转录辅调节在失神经骨骼肌在7与35天后，去神经支配，与最大的变化，25倍的下降，是Ankrd 2的差异很大。我们建议，这些转录辅助调节对诺龙敏感性的影响的分析将提供新的见解肌肉诺龙和其他雄激素的影响的分子决定因素，并解释了一些病理生理条件下骨骼肌对雄激素的抗性这一有趣的问题。该建议的主要目的是，在去神经萎缩的大鼠模型中，确定：1）诺龙是否增加去神经肌肉中的钙调神经磷酸酶活性，以及这种增加是否是由于钙调神经磷酸酶、RCAN 2和钙调神经磷酸酶水平的上游调节剂的水平的诺龙依赖性变化（FOXO 1和MAFbx）; 2）特异性转录辅助调节因子是否介导对诺龙减少萎缩和抑制MAFbx的作用的抗性。 公共卫生相关性： 康复医学的主要目标是恢复急性疾病或损伤患者的功能，以及保护慢性疾病患者的功能以保持独立性。康复的一个重要障碍是瘫痪、固定、药物和慢性疾病导致的肌肉萎缩。减少萎缩的方法集中在治疗引起肌肉损失的潜在疾病、营养和各种形式的身体调理上。这些方法不直接调节导致肌肉损失的生物过程。开发安全有效的药物，促进肌肉恢复，阻止肌肉损失，或两者兼而有之，将是加快康复治疗和出院的一大步。我们研究的两个主要预期结果是对雄激素预防肌肉萎缩的具体机制的更深入了解，以及关于这种保护的分子决定因素的新信息。除了扩展一般知识外，这些信息还具有改善未来安全预防肌肉萎缩的治疗方法的潜力。与VHA使命的相关性肌肉损失影响退伍军人的各种医疗条件，包括中风，脊髓损伤，慢性阻塞性肺病，哮喘和风湿性疾病。它还影响到正在从烧伤和/或因受伤而无法行动中恢复的伤员。改善治疗以保持或恢复这些退伍军人的功能的知识有可能显着提高他们的恢复速度，并可能提高他们的生活质量和社区功能和融合。

DESCRIPTION (provided by applicant): Androgenic steroids reduce muscle atrophy through molecular mechanisms that are poorly understood. In studies of the effects of the androgenic steroid nandrolone on atrophy of muscle paralyzed by nerve transection, we have found that rates of atrophy are reduced within 7 days after beginning nandrolone when this steroid is started at 29 days after nerve transection (35 days) associated with reduced expression of the muscle ubiquitin ligases MAFbx and MuRF1. By contrast, when nandrolone was begun at the time of nerve transection, it did not slow atrophy or reduce expression of MAFbx or MuRF1 7 days later (7 days). We believe that this model provides a unique opportunity to understand the molecular mechanisms by which nandrolone slows muscle atrophy, and to understand why muscle is resistant to the beneficial effects of nandrolone under some conditions. Of interest, gene profiling revealed that, among genes affected by nandrolone at 35 but not 7 days, and potentially capable of regulating muscle size, were FOXO1 and RCAN2. RCAN2 is expressed at high levels in skeletal muscle, where it inhibits calcineurin, a protein phosphatase involved in muscle hypertrophy by activating the transcription factor NFAT. Of interest, FOXO1 overexpression also reduces calcineurin activity, possibly by upregulating MAFbx [5-7], suggesting that nandrolone-induced reductions in FOXO1 and MAFbx levels may represent a second, parallel mechanism by which nandrolone may increase calcineurin activity in denervated muscle. The insensitivity of denervated muscle to nandrolone at early time points most likely reflects expression of a transcriptional regulator that blocks nandrolone action, or downregulation of one that is necessary for it. We found large differences in expression levels of several transcriptional coregulators in denervated skeletal muscle at 7 versus 35 days after denervation, with the greatest change, a 25-fold decrease, being for Ankrd2. We propose that analysis of effects of these transcriptional coregulators on nandrolone sensitivity will provide new insights into molecular determinants of muscle to effects of nandrolone and other androgens and explain the intriguing problem of resistance of skeletal muscle to androgens under some pathophysiological conditions. Major Aims of this proposal are, in a rat model of denervation atrophy, to determine: 1) whether nandrolone increases calcineurin activity in denervated muscle, and whether such increases are due to nandrolone-dependent changes in levels of calcineurin, RCAN2, and upstream regulators of calcineurin levels (FOXO1 and MAFbx); 2) whether specific transcriptional coregulators mediate resistance to effects of nandrolone to reduce atrophy and repress MAFbx. PUBLIC HEALTH RELEVANCE: Significance Restoration of function in those recovering from acute illness or injury, and preservation of function in those with chronic medical conditions to maintain independence are major goals in rehabilitation medicine. A significant impediment to rehabilitation is muscle atrophy as a consequence of paralysis, immobilization, medications, and chronic illness. Approaches to minimizing atrophy focus on treating the underlying disease causing muscle loss, on nutrition, and on various forms of physical conditioning. These approaches do not directly regulate the biological processes causing muscle loss. The development of safe and effective pharmaceuticals that promote muscle recovery, block muscle loss, or both would be a huge step forward in speeding up rehabilitation therapy and discharge from the hospital. The two major expected outcomes of our studies are greater insights into specific mechanisms by which androgens prevent muscle atrophy, and new information about the molecular determinants of such protection. In addition to expanding general knowledge, this information holds the potential to improve future therapeutics that safely prevent muscle atrophy. Relevance to the VHA Mission Muscle loss affects veterans with diverse medical conditions that include stroke, spinal cord injury, COPD, asthma and rheumatological disease. It also affects wounded soldiers during recovery from burns and/or immobilization resulting from their injuries. Knowledge that improves treatments to preserve or restore function of these veterans holds the potential to significantly improve the speed of their recovery, and, possibly, their quality of life and community function and integration.