Role of Lipoprotein Assembly in Maternal-Fetal Transport of Beta-Carotene

脂蛋白组装在 β-胡萝卜素母胎转运中的作用

• 批准号: 10390463
• 负责人: M Mahmood Hussain
• 金额: $ 37.02万
• 依托单位: RUTGERS, THE STATE UNIV OF N.J.
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-12 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10390463
• 关键词: Adult; Affect; Anabolism; Animal Model; Apolipoproteins B; Beta Carotene; Biological Assay; Biological Process; Blood Circulation; Carotenoids; Cells; Congenital Abnormality; Data; Developed Countries; Developing Countries; Development; Dietary Component; Dietary Practices; Dose; Elements; Embryo; Embryonic Development; Event; Fetal health; Fetus; Generations; Genetic Transcription; HNF4A gene; Health; Human; In Vitro; Incidence; Intake; Intervention; Intraperitoneal Injections; Knowledge; Link; Lipoproteins; Mediating; Metabolism; Micronutrients; Molecular; Mothers; Mus; Nutrient; Oxides; Oxygenases; Placenta; Pregnancy; Pregnant Women; Premature Birth; Production; Public Health; Publishing; Regulation; Reporter; Retinaldehyde; Retinoids; Risk; Role; Small Interfering RNA; Source; Time; Tretinoin; Vitamin A; Vitamin A Deficiency; Vitamins; Work; base; chromatin immunoprecipitation; dietary; embryo tissue; enhancing factor; fetal; human model; in vivo; inhibitor; insight; intervention program; knock-down; microsomal triglyceride transfer protein; mouse model; novel; promoter; transcription factor; trophoblast

Embryonic vitamin A deficiency or excess have been linked to congenital defects in humans and animal models, owing to the defective transcriptional action of retinoic acid (RA), the active form of vitamin A, during embryogenesis. β-carotene (BC) is the most abundant dietary vitamin A precursor. In the embryo, BCO1 cleaves intact BC, from the maternal circulation, to generate retinaldehyde, which is then oxidized to RA. Asymmetric cleavage of BC by BCO2, generates β-apo-10'-carotenal (apo10AL) which can serve as a precursor of retinoids (vitamin A and its derivatives), but also antagonize RA action. Our data suggest that lipoproteins synthesized within placenta mediate the transfer of BC towards the fetus. We hypothesize that apo10AL, generated in placenta by BCO2 from BC, upregulates microsomal triglyceride transfer protein (Mttp; MTP) through HNF4α and/or COUP-TFs, by antagonizing the action of RA. We propose three Specific Aims to understand the molecular mechanisms of BC placental-fetal transfer under a normal maternal vitamin A status. Aim 1. To investigate in vivo the regulation of placental lipoprotein biosynthesis by apo10AL and RA. We will assess placental carotenoids and retinoids levels and metabolism, and lipoprotein biosynthesis in WT, Bco1-/- and Bco2-/- mice. Aim 1A: To define the molecular events related to carotenoid metabolism and lipoprotein production that occur in placenta after a single maternal administration of BC. Dams will be sacrificed at various time points post-BC administration (at 13.5 dpc, by IP injection). Aim 1B: To establish the effects of a single maternal BC administration at various times during gestation on the molecular events related to carotenoid metabolism and lipoprotein production. Dams will be administered BC at various times during gestation and sacrificed after 4 and 24 h. Aim 2. To establish that placenta MTP deficiency abrogates accumulation of embryonic BC after its administration to the mother. A mouse model with placental-specific inactivation of Mttp will be used. Dams will be sacrificed 4 and 24 h after BC administration at 13.5 dpc, and placentas analyzed as in Aim 1. Aim 3. To investigate in vitro the regulation of human placental lipoprotein biosynthesis by apo10AL and RA. In this Aim, we will expand our studies to human models. Aim 3A: To study the apo10AL-mediated regulation of MTP function in human placenta explants and Bewo cells. We will: (i) determine the time course and dose-dependent effect of BC, apo10AL and RA on carotenoid and retinoid metabolism and lipoprotein biosynthesis; (ii) inactivate MTP by siRNA or an MTP inhibitor and (iii) Bco1 and Bco2 by siRNA; (iv) study the competing effects of apo10AL and RA on MTP expression. Aim 3B: To identify the cis-elements in the Mttp promoter and the transcription factors required for apo10AL-mediated increase in Mttp transcription. Promoter reporter assays, transcription factors knockdown, and ChIP assays will establish how placental Mttp expression is regulated by apo10AL through HNF4a and/or COUP-TFs.

在人类和动物模型中，由于维生素A的活性形式维甲酸(RA)在胚胎发育过程中转录活性的缺陷，胚胎维生素A的缺乏或过剩与先天性缺陷有关。胡萝卜素(β-胡萝卜素，BC)是膳食中含量最丰富的维生素A前体。在胚胎中，BCO1从母体循环中裂解完整的BC，生成视黄醛，然后被氧化为RA。BC被BCO2不对称裂解，产生载脂蛋白-10‘-胡萝卜素(β-apo-10’-胡萝卜素，apo10AL)，它可以作为维甲酸(维生素A及其衍生物)的前体，但也能拮抗RA的作用。我们的数据表明，胎盘中合成的脂蛋白介导了BC向胎儿的转移。我们假设，由BC产生的BCO2在胎盘中产生的apo10AL通过HNF4α和/或COUP-TF上调微粒体甘油三酯转移蛋白(MTTP；MTP)，从而拮抗RA的作用。我们提出了三个特定的目标，以了解在正常母亲维生素A状态下BC胎盘-胎儿移植的分子机制。目的1.研究apo10AL和RA对胎盘脂蛋白生物合成的调节作用。我们将评估WT、Bco1-/-和Bco2-/-小鼠胎盘类胡萝卜素和维甲酸的水平和代谢，以及脂蛋白的生物合成。目的1 A：明确孕妇单次服用BC后，胎盘中与类胡萝卜素代谢和脂蛋白产生有关的分子事件。大坝将在BC管理后的不同时间点被牺牲(在13.5DPC，通过IP注射)。目的1B：探讨孕期不同时间单次给药对类胡萝卜素代谢和脂蛋白生成相关分子事件的影响。在妊娠期间的不同时间给予DAMS，并在4小时和24小时后处死。目的2.证实胎盘MTP缺乏症消除了母体给药后胚胎BC的蓄积。将使用胎盘特异性MTTP失活的小鼠模型。分别于BC给药后4小时和24小时处死大鼠，取胎盘进行组织学分析。目的1.研究apo10AL和RA对人胎盘脂蛋白生物合成的调节作用。为了达到这个目标，我们将把我们的研究扩展到人体模型。目的3A：研究apo10AL对人胎盘组织和Bewo细胞MTP功能的调节。我们将：(I)确定BC、apo10AL和RA对类胡萝卜素和类维A酸代谢以及脂蛋白生物合成的时程和剂量依赖效应；(Ii)通过siRNA或MTP抑制剂来灭活MTP；(Iii)通过siRNA来灭活Bco1和Bco2；(Iv)研究apo10AL和RA对MTP表达的竞争作用。目的3B：鉴定MTTP启动子中的顺式元件以及apo10AL介导的MTTP转录上调所需的转录因子。启动子报告分析、转录因子敲除和芯片分析将确定apo10AL如何通过HNF4a和/或COUP-TF调节胎盘MTTP的表达。