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Development of molecular biology-based assays for human selenium status

开发基于分子生物学的人体硒状态检测方法

基本信息

批准号：
7266967
负责人：
Roger A Sunde
金额：
$ 9.37万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-08-01 至 2010-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7266967
关键词：
3&apos Untranslated Regions Abbreviations Address Ally Animal Diseases Animals Biochemical Markers Biochemistry Biological Assay Biological Markers Blood Blood Cells Codon Nucleotides Collaborations Consultations DNA Insertion Elements Development Disease England Female Goals Health Human Hydrogen Peroxide Institute of Medicine (U.S.)Intake Invasive Lactation Life Cycle Stages Link Liver Medicine Messenger RNA Methods Molecular Molecular Biology Numbers Nutritional Requirements Peptides Phospholipids Placebos Plasma Polymerase Chain Reaction Positioning Attribute Pregnancy Proteins Protocols documentation Rattus Reading Recommendation Regulation Reporting Research Reverse Transcriptase Polymerase Chain Reaction Ribonucleases Rodent Role Sampling Selenium Selenocysteine Series Structure Supplementation Thyroxine deiodinase Time Tissues Transfer RNA Translations Vertebral column Whole Blood World Health Organization base day falls glutathione peroxidase human subject improved mRNA Expression male nutrition selenoenzyme selenoprotein stem tool transposon/insertion element

项目摘要

DESCRIPTION (provided by applicant): The overall goal of this research is to develop molecular biology-based assays for human selenium (Se) status. We were the first to report that mRNA levels of Se-dependent glutathione peroxidase-1 (GPX1) fall to 10% of Se-adequate levels, thus providing a molecular explanation for why GPX1 is an effective biochemical marker for assessing Se status in humans and animals, and suggesting the hypothesis that regulation of GPX1 expression is an important aspect of the role of GPX1. Using selenoprotein mRNA levels in liver as markers, we then carefully evaluated Se requirements in rats, showing that molecular biology markers were a useful tool for assessing requirements and that understanding the regulation of these molecular markers was important when assessing requirements throughout the life cycle, such as in pregnancy and lactation. More recently, we have conducted a series of studies investigating the use of blood (a less invasive tissue), and found that GPX1 mRNA expression in rat whole blood was comparable to levels in the major tissues and could be used effectively to assess Se status. Thus we have begun to evaluate human selenoprotein mRNA levels as a potential way to assess human Se status. The advantage is that these levels appear to be homoeostatic ally controlled by Se status. Secondly, the advent of rapid molecular biology assays suggests that molecular biology markers will soon become important in assessing human health, including human nutrition. We now have found that human blood, just as in rodent blood, has selenoprotein mRNA expression at levels comparable to major tissues, suggesting that these may be the effective markers of human Se status. In this proposed research, three specific aims will be pursued: 1. To assess selenoprotein mRNA expression using RPA on human whole blood from our current (unfunded) collaboration with Reading, England, to correlate these markers with plasma GPX3 activity and plasma Se levels; 2. To develop protocols and use quantitative real-time PCR (qRT-PCR) on human whole blood to assess selenoprotein mRNA expression for a number of selenoproteins in the Reading samples; 3. To assess whole blood selenoprotein mRNA levels, along with plasma GPX3 activity, RBC GPX1 activity, and plasma Se concentration, in 120 adult human subjects in Madison WI, and then to determine the changes in selenoprotein mRNA expression 1 mo, 2 mo and 4 mo after initiation of placebo or 110 ug Se per day supplementation in 20 male and 20 female subjects. Relevance: This research is directly focused on improving our understanding and ability to use specific molecular markers to set nutrient requirements, and thus is relevant to human health. More specifically, this research will directly address the research recommendations for selenium by the 2000 Institute of Medicine Dietary Reference Intake report, including the need for biomarkers for assessment of selenium status, by developing molecular biology-based assays that are linked to homeostatic regulation of selenium status and that have the potential to be used in individualized medicine.

描述（由申请人提供）：本研究的总体目标是开发基于分子生物学的人体硒（Se）状态检测方法。我们首次报道了硒依赖性谷胱甘肽过氧化物酶-1 （GPX1）的mRNA水平下降到硒足量水平的10%，从而为GPX1为何是评估人类和动物硒状态的有效生化标志物提供了分子解释，并提出了GPX1表达调控是GPX1作用的一个重要方面的假设。使用肝脏硒蛋白mRNA水平作为标记，我们仔细评估了大鼠的硒需要量，表明分子生物学标记是评估硒需要量的有用工具，并且在评估整个生命周期（如妊娠和哺乳期）的硒需要量时，了解这些分子标记的调控非常重要。最近，我们进行了一系列研究，调查了血液（一种侵入性较小的组织）的使用，发现大鼠全血中GPX1 mRNA的表达与主要组织中的水平相当，可以有效地用于评估硒状态。因此，我们已经开始评估人硒蛋白mRNA水平，作为评估人硒状态的潜在方法。其优点是，这些水平似乎是同稳态控制硒状态。第二，快速分子生物学分析的出现表明，分子生物学标记将很快在评估人类健康，包括人类营养方面变得重要。我们现在已经发现，人类血液和啮齿动物血液一样，硒蛋白mRNA的表达水平与主要组织相当，这表明这些可能是人类硒水平的有效标志。在这项拟议的研究中，将追求三个具体目标：1。利用RPA技术评估人全血中硒蛋白mRNA的表达，将这些标记物与血浆GPX3活性和血浆硒水平联系起来。2. 制定方案，并利用人全血定量实时PCR （qRT-PCR）评估Reading样品中硒蛋白mRNA的表达；3. 评估麦迪逊WI 120名成人受试者的全血硒蛋白mRNA水平、血浆GPX3活性、红细胞GPX1活性和血浆硒浓度，然后确定20名男性和20名女性受试者在开始服用安慰剂或每天补充110 ug硒后1个月、2个月和4个月硒蛋白mRNA表达的变化。相关性：本研究的直接重点是提高我们使用特定分子标记来设定营养需求的理解和能力，因此与人类健康相关。更具体地说，这项研究将直接解决2000年医学研究所膳食参考摄入量报告中对硒的研究建议，包括需要生物标志物来评估硒的状态，通过开发基于分子生物学的分析，与硒状态的稳态调节有关，并有可能用于个体化医学。