Novel roles of the intestine to regulate whole body lipid homeostasis using pathways of nutritional physiology and molecular regulation.

肠道利用营养生理学和分子调节途径调节全身脂质稳态的新作用。

• 批准号: RGPIN-2021-03871
• 负责人: Proctor, Spencer
• 金额: $ 2.4万
• 依托单位: University of Alberta
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2021
• 资助国家: 加拿大
• 起止时间: 2021-01-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=742762
• 关键词: Novel; roles; intestine; regulate; whole

Hypothesis and Significance The working hypothesis of this program is that the intestine is a primary regulator of lipid and steroid precursors for whole body lipid metabolism and employs compensatory mechanisms through a network of novel mechanistic pathways, including inflammation. By transforming this new integrated biology, we will reveal and reconcile novel pathways for homeostatic and perturbed nutritional states. Primary Objectives 1. Assess the nutritional and physiological significance of intestinal-derived (lymph) HDL to whole body cholesterol homoeostasis using a higher order animal model (swine). Short- and long-term objectives include comparing HDL and its primary protein (apolipoprotein AI) synthesized by both the intestine and the liver; understand its contrition and kinetics to cholesterol transport and excretion; map the profile of miR's associated within the HDL fraction from lymph and discover their contributions to regulate metabolism. We will utilize the indwelling lymph-cannulation in swine (developed by our group) to assess the impact of lymphopenia on HDL/cholesterol metabolism; employ tracers to determine apoAI kinetics derived from hepatic and mesenteric lymphatics; continue to develop miR arrays for lymphatic samples from swine and characterize their profile under different nutritional states. 2. Delineate the novel role of miR-150 in mesenteric lymph and impact to lipid and HDL metabolism. We will determine the immune response under fed and fasted conditions; investigate the relationship of miR-150 during impaired lipid metabolism to assess its role pro-inflammatory responses during this state. We will use both rodent and swine models to generate lymph samples under different nutritional states. Inflammatory status will be assessed using ex vivo cell proliferation techniques (with and without miR-150), as well as cell sorting (FACS), cell-surface markers and gating approaches. We also propose to develop new functional methodological approaches to validate the relevance of miR-150 such as using isolated lymphocytes pre-incubated with antagomir-150 or antagomir-scr at varying concentrations. 3. Reveal the molecular mechanisms that regulate TICE within the enterocyte identified by our preliminary data; (i) intracellular transcription, (ii) HSP90 chaperone cycle for steroid hormone receptors, (iii) enterocytic transport and metabolism and (iv) cell-cell communication. We have applied proteomic and gene array technologies to isolates of whole enterocytes as well as specific brush boarder membrane (BBM) and used system mapping to identify the canonical pathways involved in TICE. We have developed nutritional and pharmacological agents that can impair or stimulate TICE in vivo. We will combine these integrated biological approaches. Exciting preliminary data suggests that HDL derived from the lymphatics maybe an effective donor for TICE (ex vivo Ussing Chamber methods) and represents a pioneering perspective for the field.

假设和意义 该计划的工作假设是，肠道是全身脂质代谢的脂质和类固醇前体的主要调节器，并通过包括炎症在内的新型机制途径网络采用补偿机制。通过改造这种新的综合生物学，我们将揭示并协调稳态和扰动营养状态的新途径。主要目标 1. 使用高级动物模型（猪）评估肠源性（淋巴）HDL 对全身胆固醇稳态的营养和生理意义。短期和长期目标包括比较 HDL 及其由肠道和肝脏合成的主要蛋白质（载脂蛋白 AI）；了解其对胆固醇转运和排泄的影响和动力学；绘制淋巴 HDL 部分中相关 miR 的谱图，并发现它们对调节代谢的贡献。我们将利用猪体内留置淋巴插管（由我们小组开发）来评估淋巴细胞减少症对HDL/胆固醇代谢的影响；使用示踪剂来确定源自肝和肠系膜淋巴管的 apoAI 动力学；继续开发猪淋巴样本的 miR 阵列，并表征其在不同营养状态下的特征。 2. 描述miR-150在肠系膜淋巴中的新作用以及对脂质和HDL代谢的影响。 我们将确定进食和禁食条件下的免疫反应；研究 miR-150 在脂质代谢受损期间的关系，以评估其在此状态下促炎症反应的作用。我们将使用啮齿动物和猪模型来生成不同营养状态下的淋巴样本。将使用离体细胞增殖技术（有或没有 miR-150）以及细胞分选（FACS）、细胞表面标记物和门控方法来评估炎症状态。我们还建议开发新的功能方法来验证 miR-150 的相关性，例如使用与不同浓度的 antagomir-150 或 antagomir-scr 预孵育的分离淋巴细胞。 3.揭示我们初步数据确定的肠上皮细胞内调节TICE的分子机制； (i) 细胞内转录，(ii) 类固醇激素受体的 HSP90 伴侣循环，(iii) 肠细胞转运和代谢，以及 (iv) 细胞间通讯。我们已将蛋白质组学和基因阵列技术应用于整个肠细胞以及特定刷状寄生膜 (BBM) 的分离，并使用系统图谱来识别 TICE 中涉及的典型途径。我们开发了可以削弱或刺激体内 TICE 的营养和药物制剂。我们将结合这些综合的生物学方法。令人兴奋的初步数据表明，源自淋巴管的 HDL 可能是 TICE（离体 Ussing Chamber 方法）的有效供体，并代表了该领域的开拓性前景。