Quantitation of protein turnover in the human adult lens using the 14C bomb-pulse

使用 14C 炸弹脉冲定量成人晶状体中的蛋白质周转

基本信息

批准号：
7681210
负责人：
Bruce A Buchholz
金额：
$ 24.58万
依托单位：
UNIVERSITY OF CALIF-LAWRNC LVRMR NAT LAB
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-09-01 至 2012-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7681210
关键词：
Adult Age Alzheimer&apos s Disease Amyloid deposition Anabolism Appearance Attention Attenuated Back Birth Brain Calendar Carbon Carbon Dioxide Cataract Cataract Extraction Cell Nucleus Cells Cessation of life Communities Crystalline Lens Crystallins Cytoskeletal Proteins Data Deltastab Development Diffusion Disease Drops Event Exhibits Eye Fetal Proteins Genome Growth Healthcare Systems Heart Homeostasis Human Individual Label Laboratories Lens Fiber Life Maintenance Mass Spectrum Analysis Measures Membrane Molecular Myocardial Ischemia Natural regeneration Nature Neurons Nuclear Nuclear Protein Nuclear Proteins Nuclear Weapon Oceans Operative Surgical Procedures Pathology Pattern Physiologic pulse Physiology Presbyopia Procedures Production Protein Biosynthesis Protein Dynamics Protein-Folding Disease Proteins Radioactive Records Reporting Research Personnel Sampling Solubility Stress Stroke Testing Time Tissue Banking Tissue Banks Tissues Tracer Treaty Vision Water Work accelerator mass spectrometry age related aged alpha-Crystallins attenuation base falls fiber cell in vivo interest lens lens protein macromolecule middle age milligram nervous system disorder pressure prevent protein degradation protein misfolding repaired senescence stable isotope synthetic protein tool trend

项目摘要

DESCRIPTION (provided by applicant): Studying synthesis and repair of macromolecules in healthy humans is difficult. Above ground testing of nuclear weapons from 1955-1963 produced a large bomb-pulse of 14CO2, which was quickly distributed around the globe and intrinsically labeled all exchangeable carbon in the biosphere. Since the Test Ban Treaty in 1963, the atmospheric 14C bomb-pulse has been decreasing exponentially with a mean life of ~ 16 years, not due to radioactive decay, but due to diffusion and equilibration with the oceans and biosphere. All living humans have been labeled, and the 14C concentration in all organic macromolecules can sensitively identify when they were synthesized. In reality, we are all subjects in a long-term quasi-stable isotope tracer study in which molecular synthesis can be dated through the use of accelerator mass spectrometry (AMS) to count individual 14C atoms in sub-milligram samples. We seek to utilize this effective molecular chronometer to establish, quantify, and identify the specific nature of protein turnover in healthy adult human eye lenses. Preliminary data from lenses age 60 and greater suggests that the crystallin proteins of aged nuclear fiber cells contain carbon significantly younger than the cells themselves. This serves as direct evidence of in vivo protein turnover in human nuclear fiber cells and verifies the controversial hypothesis that the human eye lens maintains homeostasis at its core (at least in part) by de novo protein production. We will measure carbon turnover in purified proteins from the nuclear core of lenses formed after the peak of the bomb-pulse plus some aged controls. Compared with the data gathered from older donors, dating protein incorporation in younger lenses will provide direct evidence if this rate is attenuated when we reach middle age. A better understanding of lens maintenance could provide a useful new tool to understand and ultimately prevent the most common form of lens pathology, age-related nuclear (ARN) cataract. The expense and the demand for cataract surgery have catapulted this disease to become among the single largest burdens to our beleaguered healthcare system. Although surgical intervention is a highly effective means of restoring vision, if we could delay the onset of cataract by a decade the need for the procedure would drop by half. We will establish the baseline for specific protein turnover in the healthy adult human lens and relate the turnover rate to age. Cataracts, which can be classified as an age-related protein folding disease, require a long-term (~50 year) stable isotope tracer assessment to reveal the fundamental mechanisms a healthy lens employs to maintain the viability of its exceedingly long-lived proteins. AMS has enabled us to use elevated tissue loads of 14C produced by atmospheric nuclear testing as a means to evaluate the long term dynamics of protein in healthy lens tissue. Our preliminary data has demonstrated protein turnover at approximately 1% per year by mass in the soluble crystalline proteins of the lens core, indicating that the lens possesses a mechanism for protein repair in a region of tissue previously believed to be senescent. The aim of this study is to firmly establish, quantify, and identify the specific targets of this protein exchange in healthy lenses and bring the existence of this unknown but seemingly key feature of lens physiology to the attention of cataract researchers worldwide.

描述（由申请人提供）：研究健康人中大分子的合成和修复是困难的。从1955 - 1963年开始对核武器进行地面测试产生了大型的14CO2炸弹脉冲，该炸弹脉冲在全球范围内迅速分布，并在生物圈中固有地标记了所有可交换碳。自1963年禁止测试禁令以来，大气14C炸弹脉冲的平均寿命约为16年，而不是由于放射性衰减，而是由于扩散和与海洋和生物圈的平衡所致。所有活人都被标记了，所有有机大分子中的14C浓度均可敏感地识别它们合成的何时。实际上，我们都是长期准稳定的同位素示踪剂中的受试者，在该研究中，可以通过使用加速器质谱法（AMS）来计算亚毛皮样品中的单个14C原子，从而可以通过分子合成来记录分子合成。我们寻求利用这种有效的分子计时仪来建立，量化和确定健康成人人眼镜蛋白更新的特定性质。来自60岁及60岁镜头的初步数据表明，老化的核纤维细胞的结晶蛋白含有比细胞本身明显年轻的碳。这是人体核纤维细胞中体内蛋白质更新的直接证据，并验证了人眼镜镜的稳态（至少部分）由从头蛋白产生的核心（至少部分）。我们将从炸弹脉冲峰和一些老年对照组后形成的透镜的核心核心中测量纯化蛋白质的碳更换。与从老年捐助者那里收集的数据相比，如果我们达到中年时，年轻镜头中的约会蛋白掺入将直接证据。对晶状体维护的更好理解可以为理解并最终预防与年龄相关的核（ARN）白内障最常见形式的新工具提供有用的新工具。白内障手术的费用和需求使这种疾病成为我们陷入困境的医疗保健系统的最大负担之一。尽管手术干预是恢复视力的高效手段，但如果我们可以将白内障的发作延迟十年，那么对手术的需求将下降一半。我们将在健康的成年人类镜头中建立特定蛋白质周转的基线，并将周转率与年龄联系起来。可以将白内障分类为与年龄相关的蛋白质折叠疾病，需要长期（约50年）稳定的同位素示踪剂评估，以揭示健康晶状体采用的基本机制来维持其超持寿命的蛋白质的生存能力。 AMS使我们能够使用由大气核试验产生的14C的升高组织负荷作为评估健康晶状体组织中蛋白质长期动态的一种手段。我们的初步数据表明，在晶状体核心的可溶性晶体蛋白中，质量每年约1％的蛋白质更新，这表明该晶状体在以前认为是衰老的组织区域中具有蛋白质修复的机制。这项研究的目的是牢固建立，量化和确定健康镜片中这种蛋白质交换的特定靶标，并将这种镜头生理学的未知但看似关键特征的存在引起全球性白内障研究人员的注意。