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. ? ? ?
