Determination of methylmercury metabolism and elimination status in humans

 描述(申请人提供)：甲基汞(MeHg)是一种持久性环境毒素，积聚在鱼类和海鲜中。了解甲基汞的风险与鱼类饮食中必需营养素的益处是人类健康的优先问题。甲基汞的毒性是由其缓慢的消除速度决定的，众所周知，这一速度在人与人之间变化很大。量化人体内甲基汞排除量的方法严重不发达。此外，消除甲基汞所需的代谢机制还没有完全了解。我们的目标有两个：1)建立非侵入性方法来量化 (2)验证肠道细菌酶在甲基汞去甲基化过程中的假定作用，从而促进甲基汞的消除。我们的长期目标是使用这种方法来研究影响甲基汞代谢的遗传和饮食因素。我们将使用一个首先在啮齿动物模型中建立的模型，根据该模型，肠道细菌对甲基汞的去甲基化是其消除的限速步骤。然而，肠道细菌在人类体内甲基汞去甲基化和消除中的作用仍有待鉴定。人类微生物群中的一些细菌物种在mer基因座上含有汞解毒基因。对我们的假设特别重要的是MerB(有机汞裂解酶)和MERA(汞还原酶)基因，它们执行甲基汞的还原去甲基化。我们的工作假设是：1)人体内甲基汞的消除是由甲基汞去甲基化的限速步骤决定的，2)人类体内的大部分甲基汞去甲基化是通过细菌汞解毒酶在肠道中发生的。根据这些假设，我们预测不同个体之间甲基汞清除率的差异将与肠道微生物组中merB和mera基因的丰度和活性相对应。在这项研究中，我们将评估以自然产生的甲基汞水平喂食金枪鱼的受试者的结果。在目标1中，我们将确定人类食用鱼粉后甲基汞清除率与甲基汞去甲基化状态的相关性。甲基汞的清除率将通过头发样本的纵向汞分析来确定。甲基汞去甲基化状态将以粪便中汞的百分比来确定。在目标2中，我们将确定细菌merB和meA的丰度和活性对甲基汞消除速率和甲基汞去甲基化状态的影响。我们将对服用金枪鱼粉的受试者的粪便样本进行调查，以了解MerB和Mera DNA的丰富性和多样性，以及MerB和Mera变体的特定酶活性。将确定MerB和MERA丰度和活性与甲基汞清除率和粪便中去甲基化状态的统计相关性。我们预计，这项研究的发现将产生一种宝贵的方法来识别甲基汞代谢“快”或“慢”的个体，并进一步解决通过改变肠道细菌组成来改变甲基汞代谢的潜在机制。

 DESCRIPTION (provided by applicant): Methylmercury (MeHg) is a persistent environmental toxin that accumulates in fish and seafood. Understanding the risks of MeHg versus the benefits of essential nutrients in a fish diet is a priority human health issue. Toxicity of MeHg is dictate by its slow rate elimination rate, which is known to be highly variable from person to person. Methods to quantify MeHg elimination in people are grossly underdeveloped. Furthermore, the mechanisms of MeHg metabolism that are required for its elimination are not fully understood. Our objectives are two-fold: 1) to establish non-invasive methods to quantify the variable trait of MeHg metabolism and elimination in individual people consuming a normal fish diet and 2) to validate the putative role for intestinal bacterial enzymes in de-methylation of MeHg which facilitates its elimination. Our long-term goal is to use this method to investigate genetic and dietary factors that affect MeHg metabolism. We will work with a model first established in rodent models whereby de-methylation of MeHg by bacteria in the gut is a rate-limiting step in its elimination. However, the role of gut bacteria in MeHg de-methylation and elimination in humans remains to be characterized. A number of bacterial species in the human microbiome harbor Hg-detoxifying genes contained in the mer locus. Of particular importance to our hypothesis are the merB (organomercurial lyase) and merA (mercury reductase) genes, which carry out reductive de-methylation of MeHg. Our working hypotheses are that: 1) elimination of MeHg from the human body is dictated by the rate-limiting step of MeHg de-methylation and 2) the majority of MeHg de-methylation in humans occurs in the gut via bacterial mercury de-toxifying enzymes. With these hypotheses we predict differences in MeHg elimination rate between individuals will correspond with the abundance and activity of the merB and merA genes in the gut microbiome. In this study we will evaluate outcomes in human subjects fed meals of tuna with naturally occurring levels of MeHg. In Aim 1, we will determine the correlation of MeHg elimination rate with MeHg de-methylation status in human subjects subsequent to fish meal consumption. MeHg elimination rate will be determined by longitudinal Hg analysis of hair samples. MeHg de-methylation status will be determined as the percent Hgi in feces. In Aim 2, we will determine the influence of the abundance and activity of bacterial merB and merA on MeHg elimination rate and MeHg de-methylation status. We will investigate fecal samples of subjects receiving tuna meals for the abundance and diversity of merB and merA DNA and as well as the specific enzyme activity of merB and merA variants. Statistical correlations of merB and merA abundance and activity with MeHg elimination rate and with de-methylation status in feces will be determined. We anticipate the findings from this study will yield an invaluable method to identify individuals with a "fast" or "slow" MeHg metabolism, and furthermore resolve a potential mechanism for altering MeHg metabolism via changes to the gut bacterial composition.