Platelet Mitochondrial Epigenetics: New CVD Markers of Particle and Metal Effects

描述（由申请人提供）：生物标志物是细胞、生物化学或分子改变，可以在人体组织中轻松且非侵入性地测量，并直接或间接参与疾病途径。心血管疾病（CVD）是美国死亡的主要原因，每年造成80万人死亡。在流行病学研究中，短期和长期暴露于空气污染/有毒金属一直与CVD发病率和死亡率相关。血小板减少有助于动脉粥样硬化和动脉血栓形成的发展，这是导致CVD的主要促成事件。血小板计数增加被认为是介导颗粒物（PM）效应的原因。血小板是无核的，但线粒体DNA（mtDNA）可以很容易地从血浆血小板分离。线粒体对PM相关的氧化高度敏感，最重要的是，血小板严重依赖于线粒体中的能量产生。我们假设，空气污染物和金属暴露可能会导致血小板线粒体DNA甲基化（mtDNA）的改变，以响应吸烟诱导的全身氧化/炎症。血小板mtDNA甲基化改变可能导致或至少反映线粒体功能障碍，直接或间接参与将毒性暴露与CVD风险联系起来的途径。我们建议t检验这些假设使用的数据和生物标本从规范老化研究（NAS），一个大型的前瞻性队列的老龄化个体。我们已经修改和调整了从血浆血小板中分离mtDNA的方案，并成功地测试了我们高度时间和成本效益的方法。将在间隔4年的两个时间点测量700名参与者的血浆血小板的mtDNA氧化损伤（8-oxo-dG），标准DNA甲基化（5-mC）和新兴羟甲基化（5-OH-mC）标记的线粒体整体含量，以及一组完整的mtDNA序列中的基因座特异性甲基化。我们将确定空气污染和金属对线粒体DNA生物标志物的影响（目标1）;评估线粒体DNA生物标志物与心血管疾病风险的进一步关联（目标2）;并进行探索性目标（目标3），通过调解分析确定血小板线粒体DNA变化是否直接或间接地处于将有毒暴露与氧化/炎症标志物以及最终与心血管疾病风险联系起来的路径上。这项研究将首次描述与CVD风险相关的血小板mtDNA甲基化机制。该项目的重点是血小板，一个单一的纯细胞群体与心血管疾病病因学的关键作用，将提供易于解释的表观遗传学的发现，从而克服了目前的表观遗传学研究的主要局限性，在大的人群，这已被阻碍了使用混合细胞群体，如白色血细胞或颊细胞。由于血小板可以廉价且非侵入性地从血浆中分离出来，因此我们的研究很有可能产生未来心血管疾病的新型风险和诊断生物标志物。如果我们观察到假设的关联，我们将寻求资金，在具有CVD高风险的不同人群中进行大规模研究，以扩大这一研究领域。

DESCRIPTION (provided by applicant): Biomarkers are cellular, biochemical, or molecular alterations that can be easily and non-invasively measured in human tissues and are directly or indirectly involved in the pathway of disease. Cardiovascular disease (CVD) is the leading cause of death in the United States accounting for 800,000 deaths every year. Both short- and long-term exposure to air pollution/toxic metals has been consistently associated with CVD morbidity and death in epidemiologic studies. Platelets contribute to the development of both atherosclerosis and arterial thrombosis, which are primary contributing events leading to CVD. Increased platelet counts have been suggested to mediate particulate matter (PM) effects. Platelets are anucleate, but mitochondrial DNA (mtDNA) can be easily isolated from plasma platelets. Mitochondria are highly sensitive to PM-related oxidation and, most importantly, platelets are critically dependent on energy production in mitochondria. We hypothesize that air pollutants and metals exposure may cause alteration in platelet mitochondrial DNA methylation (mtDNA) in response to exposure-induced systemic oxidation/inflammation. Platelet mtDNA methylation changes may lead to, or at a minimum reflect mitochondrial dysfunction participating directly or indirectly to the paths linking toxic exposures to CVD risk. We propose t test these hypotheses using data and biospecimens from the Normative Aging Study (NAS), a large prospective cohort of aging individuals. We have modified and adapted protocols to isolate mtDNA from plasma platelets, and successfully tested our highly time- and cost-effective approach. Plasma platelets from 700 participants will be measured at two time points separated by 4 years for mtDNA oxidative damage (8-oxo-dG), mitochondrial global content of both standard DNA methylation (5-mC) and the emerging hydroxymethylation (5-OH-mC) mark, and locus-specific methylation in a comprehensive panel of mtDNA sequences. We will determine the effects of air pollution and metals on mtDNA biomarkers (Aim 1); evaluate further association of mtDNA biomarkers with CVD risk (Aim 2); and conduct an exploratory aim (Aim 3) to determine via mediation analysis whether platelet mtDNA changes are directly or indirectly on the paths linking toxic exposures to oxidation/inflammation markers, and ultimately to CVD risk. The proposed study will be the first to characterize mechanisms in platelet mtDNA methylation related to CVD risk. The project's focus on platelets, a single pure cell population with critical roles in CVD etiology will provide ease of interpretation of epigenetic findings and thus overcome the major limitation of current epigenetic studies in large human population, which have been hampered by the use of mixed cell populations such as white blood cells or buccal cells. Because platelets can be inexpensively and non- invasively isolated from plasma, our study has high potential to generate novel risk and diagnostic biomarkers of future CVD. If we observed the hypothesized associations, we will seek funding to expand this line of research in large-scale studies in diverse populations with high risk of CVD.