Elucidating the molecular mechanisms linking maternal choline supplementation to healthy cognitive aging

阐明母体胆碱补充与健康认知衰老之间的分子机制

• 批准号: 1606833
• 负责人: Ramon Velazquez
• 金额: $ 22.19万
• 依托单位: Arizona State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-15 至 2018-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1606833&HistoricalAwards=false
• 关键词: Elucidating; molecular; mechanisms; linking; maternal

Part 1: Non-technical DescriptionThis project is supported under the SBE Postdoctoral Research Fellowships (SPRF) program. Age-related cognitive decline, hereafter referred to as cognitive aging, is a fact of life. To this end, structural and functional brain changes invariably lead to decreased cognitive functions even in otherwise healthy individuals. Although there is a large body of work on ways to reduce cognitive deficits associated with disease states, little is known about the mechanisms underlying cognitive aging. This is concerning given that life expectancy is increasing and cognitive aging leads to a deterioration of general health for the aging population. One option to reduce cognitive aging is the supplementation of the maternal diet with choline (MCS), an essential nutrient grouped with the vitamin B complex. Preliminary work has demonstrated that MCS leads to amelioration of cognitive aging. The primary goal of this postdoctoral project is to elucidate the underlying neural and molecular mechanisms linked to MCS benefits. Additionally, the PI plans to examine genetic targets within brain structures associated with memory formation to isolate genes that are differentially altered by MCS and aging. This will likely reveal various gene targets that will be the focus of future cognitive aging studies. Because choline is a non-toxic nutrient found in food and can be easily supplemented orally, the research team believes that any modifications to the recommended daily intake amount to reduce cognitive aging will be expedited. To this end, the results of this application may help establish new guidelines on how a diet regimen of MCS should be implemented in expecting women to reduce cognitive aging in their offspring. Data will be shared with both the scientific and general community through presentations at conferences and public forums. Part 2: Technical DescriptionThe loss of cognitive function is a pervasive and often debilitating feature of the aging process. To this end, structural and functional brain changes invariably lead to decreased cognitive functions even in otherwise healthy individuals. Recent work has shown that supplementation of choline, an essential nutrient grouped with the vitamin B complex, in the maternal diet (MCS) reduces cognitive aging. However, the molecular mechanisms linked to MCS benefits remains elusive. Elevated homocysteine levels correlate with cognitive aging, and aberrant gene expression mediated by reduced DNA methylation may contribute to cognitive aging. Choline is the major dietary source of methyl groups for the conversion of homocysteine to methionine, and for the production of S-Adenosyl methionine (SAM). SAM is a key substrate for epigenetic mechanisms, such as DNA methylation. Therefore, we hypothesize that MCS may reduce cognitive aging deficits by (1) reducing the buildup of homocysteine levels, and (2) by altering fetal epigenetic mechanisms during development leading to functional improvements in late life. Herein, we will breed 2-month-old C57Bl/6 mice. One-third of the breeding pairs will be kept on a CTL diet (choline normal diet, with standard choline content of 1.1 g/kg choline chloride), while the remaining mice will be kept on a maternal choline supplemented (MCS) diet (5 g/kg choline chloride), from conception through postnatal day 21. The offspring will be kept on the same choline diet as the parents until weaning at postnatal day 21. Notably, a group of dams from the MCS groups will be injected every other day with a betaine-homocysteine S-methyltransferase blocker S-(ä-carboxybutyl)-DL-homocysteine (CBHcy) that prevents the choline-mediated decrease in homocysteine levels. Thus, we will be able to determine whether the benefits of MCS are directly linked to homocysteine levels. Mice will be tested behaviorally using a longitudinal and a cross sectional strategy at 2, 8, 15 and 18 months of age to collect data at multiple time points and control for re-test effects. Tissue will be processed to (1) examine dendritic spine number and morphology within the hippocampus and (2) to examine alterations of DNA methylation in the promoter region of neuronal dendritic morphology-related genes (Dlg4, Rac1, RhoA, Doc2b). We will complement this work by using an unbiased approach to identify genes that are differentially methylated by MCS. These experiments will be done exclusively in hippocampal CA1 neurons isolated by laser-capture microdissection. If successful, our results would dissect the underlying molecular mechanisms whereby choline supplementation reduces cognitive aging. Understanding MCS benefits at the behavioral, neural and molecular level may lead to a modification in the recommended amounts of choline required for pregnant mothers for optimal cognitive functioning and prevention of cognitive aging.

第1部分：非技术性说明本项目是根据SBE博士后研究奖学金（SPRF）计划的支持。与糖尿病相关的认知能力下降，以下简称为认知老化，是生活中的一个事实。为此，大脑结构和功能的变化总是导致认知功能下降，即使在其他健康的个体中也是如此。虽然有大量的工作方法来减少与疾病状态相关的认知缺陷，但对认知老化的机制知之甚少。这一点令人担忧，因为预期寿命正在增加，认知老化导致老龄人口的总体健康状况恶化。减少认知老化的一个选择是在母亲的饮食中补充胆碱（MCS），这是一种与维生素B复合物一起归类的必需营养素。初步研究表明，MCS导致认知老化的改善。这个博士后项目的主要目标是阐明与MCS益处相关的潜在神经和分子机制。此外，PI计划检查与记忆形成相关的大脑结构内的遗传靶点，以分离出因MCS和衰老而发生差异改变的基因。这将可能揭示各种基因靶点，这些靶点将成为未来认知衰老研究的重点。由于胆碱是一种在食物中发现的无毒营养素，可以很容易地口服补充，研究小组认为，任何修改建议的每日摄入量，以减少认知老化将加快。为此，该应用程序的结果可能有助于建立关于如何实施MCS饮食方案的新指南，以期望女性减少其后代的认知老化。数据将通过在会议和公共论坛上的介绍与科学界和公众分享。第2部分：技术描述认知功能的丧失是衰老过程中一个普遍的、经常使人衰弱的特征。为此，大脑结构和功能的变化总是导致认知功能下降，即使在其他健康的个体中也是如此。最近的研究表明，在母亲饮食（MCS）中补充胆碱（一种与维生素B复合物一起的必需营养素）可以减少认知老化。然而，与MCS益处相关的分子机制仍然难以捉摸。高同型半胱氨酸水平与认知老化相关，DNA甲基化降低介导的基因表达异常可能导致认知老化。胆碱是将高半胱氨酸转化为甲硫氨酸和产生S-腺苷甲硫氨酸（SAM）的甲基的主要膳食来源。SAM是表观遗传机制（如DNA甲基化）的关键底物。因此，我们假设MCS可能通过（1）减少同型半胱氨酸水平的积累，和（2）改变发育期间的胎儿表观遗传机制，导致晚年功能改善，从而减少认知老化缺陷。 在此，我们将繁殖2个月大的C57 B1/6小鼠。三分之一的繁殖对将保持CTL饮食（胆碱正常饮食，标准胆碱含量为1.1 g/kg氯化胆碱），而其余小鼠将保持母体胆碱补充（MCS）饮食（5 g/kg氯化胆碱），从受孕到出生后第21天。后代将保持与父母相同的胆碱饮食，直到出生后第21天断奶。值得注意的是，MCS组的一组母鼠将每隔一天注射一次甜菜碱-高半胱氨酸S-甲基转移酶阻断剂S-（ε-羧基丁基）-DL-高半胱氨酸（CBHcy），以防止胆碱介导的高半胱氨酸水平降低。因此，我们将能够确定MCS的益处是否与同型半胱氨酸水平直接相关。将在2、8、15和18月龄时使用纵向和横截面策略对小鼠进行行为测试，以在多个时间点收集数据并控制重新测试的影响。处理组织以（1）检查海马内树突棘数量和形态，以及（2）检查神经元树突形态相关基因（Dlg 4、Rac 1、RhoA、Doc 2b）启动子区DNA甲基化的改变。我们将补充这项工作，使用一个公正的方法来确定基因的差异甲基化MCS。这些实验将专门在通过激光捕获显微切割分离的海马CA 1神经元中进行。如果成功，我们的研究结果将剖析胆碱补充剂减少认知老化的潜在分子机制。了解MCS在行为，神经和分子水平上的益处可能会导致孕妇最佳认知功能和预防认知老化所需胆碱的推荐量的修改。