TISSUE SPECIFIC NUTRITIONAL ADAPTATIONS IN RENAL FAILURE

肾衰竭时的组织特异性营养适应

• 批准号: 2430254
• 负责人: S. Russ Price
• 金额: $ 14.54万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-08-01 至 2000-05-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2430254
• 关键词: acidosis; acute renal failure; adrenalectomy; aldehyde /ketone oxidoreductase; aminoacid metabolism; ammonium chloride; branched chain aminoacid; corticosteroid receptors; dietary proteins; enzyme activity; genetic promoter element; glucocorticoids; high performance liquid chromatography; laboratory rat; liver cells; liver metabolism; malnutrition; muscle cells; muscle metabolism; nucleic acid sequence; nutrition related tag; striated muscles; tissue /cell culture; transfection

The irreversible degradation of branched-chain amino acids (BCAAs) by branched-chain ketoacid dehydrogenase (BCKAD) is regulated downward when dietary protein is limited as in the cases of anorexia or the use of low- protein diets to treat the symptoms of or slow progression of uremia. Increased BCKAD activity causing accelerated degradation of BCAA could blunt or block this nutritional response and limit the availability of BCAA for protein synthesis. Besides protein intake, acidosis accelerates BCAA catabolism; correction of the acidosis of kidney failure normalizes whole-body leucine oxidation. Glucocorticoids (GC) are involved in this process because: 1) GC are increased in acidosis and uremia; and 2) GC can modulate the activation of BCKAD. Preliminary experiments suggest that BCKAD activity and subunit mRNA levels are increased in muscles of adrenalectomized (ADX) rats fed a low-protein diet only when they are acidotic and given GC. In sharp contrast, BCKAD activity and subunit mRNA levels are decreased in liver. Our objectives are 1) to determine if conditions causing malnutrition change BCAA metabolism by varying the activity of BCKAD at both genetic and biochemical levels; and 2) to define the role of GC in these responses. Specific Aim 1: to elucidate the role of GC in regulating BCKAD enzyme activity and/or mRNA abundance in liver and muscle. Our hypothesis is that GC play a pivotal role in changing BCKAD activity and mRNA levels in different tissues. BCKAD enzyme activity, as well as subunit mRNA abundance and transcription rates will be measured in muscle and liver of ADX acidotic rats plus/minus GC supplements. To identify the role of GC in these responses, BCKAD activity and mRNA abundance will be measured in LLC-PK1 cells which lack GC receptors and in cells transfected with the GC receptor gene; we will also study BC3H1 myocytes which express GC receptors. Specific Aim 2: to determine if acute uremia, a condition characterized by loss of lean body mass, affects BCKAD activity in muscle and liver, independently of GC and/or acidosis. The hypotheses is that uremia may have independent effects on BCKAD activity, protein abundance, and mRNA levels will be measured in ADX rats with ARF, plus/minus GC supplements and/or plus/minus bicarbonate supplements. Specific Aim 3: to determine the molecular basis for two BCKAD result from different forms of BCKAD E1alpha mRNAs expressed in tissues. Moreover, this could be the basis for the observation that the percentage of total BCKAD enzyme in the active state varies dramatically in different tissues. Specific Aim 4: to characterize elements in the rat BCKAD E1alpha gene promotor. The hypothesis is that the abundance of BCKAD E1alpha mRNA is regulated by altering transcription. Results from these studies should elucidate mechanisms that influence how amino acid metabolism can change even when dietary protein restriction should suppress BCAA oxidation.

支链氨基酸的不可逆降解 支链酮酸脱氢酶（BCKAD）被下调， 饮食蛋白质是有限的，如在厌食症或使用低- 尿毒症的症状有哪些？ BCKAD活性增加导致BCAA降解加速， 减弱或阻止这种营养反应，并限制 BCAA用于蛋白质合成。 除了蛋白质摄入， 纠正肾衰竭酸中毒使其正常化 全身亮氨酸氧化。 糖皮质激素（GC）参与其中 这是因为：1）酸中毒和尿毒症时GC增加; 2）GC可 调节BCKAD的激活。 初步实验表明， BCKAD活性和亚基mRNA水平在肌肉中增加， 肾上腺切除（ADX）大鼠仅在 酸中毒并给予GC。 与此形成鲜明对比的是，BCKAD活性和亚基mRNA 肝脏中的水平降低。 我们的目标是：（1）确定 引起营养不良的条件改变支链氨基酸代谢， BCKAD在遗传和生物化学水平上的活性;和2）定义 GC在这些反应中的作用。 具体目标1：阐明作用 GC对肝脏BCKAD酶活性和/或mRNA丰度的调节 和肌肉 我们的假设是GC在改变 不同组织中BCKAD活性和mRNA水平。 BCKAD酶 活性，以及亚基mRNA丰度和转录速率将 在ADX酸中毒大鼠+/-GC的肌肉和肝脏中进行测量 补充剂. 为了确定GC在这些反应中的作用，BCKAD 将在LLC-PK1细胞中测量活性和mRNA丰度，所述LLC-PK1细胞缺乏 在GC受体和GC受体基因转染的细胞中，我们将 还研究了表达GC受体的BC3H1肌细胞。 具体目标2： 确定是否急性尿毒症，一种以瘦体丧失为特征的疾病， 质量，影响肌肉和肝脏中的BCKAD活性，独立于GC 和/或酸中毒 假设尿毒症可能与 对BCKAD活性、蛋白质丰度和mRNA水平的影响将是 在ARF、加/减GC补充剂和/或加/减GC补充剂的ADX大鼠中测量 碳酸氢盐补充剂。 具体目标3：确定分子基础 对于两种BCKAD结果，来自表达的不同形式的BCKAD E1 α mRNA 在组织中。 此外，这可能是观察的基础， 处于活性状态的总BCKAD酶的百分比变化 在不同的组织中发生了显著的变化。 具体目标4：表征 大鼠BCKAD E1 α基因启动子中的元件。 前提是 BCKAD E1 α mRNA的丰度通过改变 转录。 这些研究的结果应阐明机制 影响氨基酸代谢的变化， 蛋白质限制应该抑制BCAA氧化。