Impact of Lipids On Intestinal Mucus Transport And Structural Properties

描述(由申请人提供)：本项目的总体目标是阐明胃肠道(GI)中的脂质和与食物相关的物理化学变化对肠道粘液屏障特性的影响。这些信息将激发控制粘膜屏障的战略，实现有效的药物载体系统(如粘膜疫苗)，增强营养，并潜在地抑制病原体入侵。胃肠道粘液是一种天然水凝胶，提供了一种微调的、令人惊讶的选择性屏障，保护底层上皮免受肠腔内严酷的物理化学变化(如胆盐浓度升高、pH降低)的影响，选择性地抑制微生物的运输，从而实现高效 吸收营养物质和水分。尽管粘液在健康和疾病中的作用和潜在的影响非常重要，但人们对其屏障特性的了解相对较少。初步数据表明，肠道粘液的性质受到与食物相关的脂质和与食物相关的物理化学变化(即pH和[Ca2]的变化)的显著调节。为了研究这些现象并使其能够设计出用于治疗目的的策略，将使用多粒子跟踪技术和电子顺磁共振(EPR)的新应用来分析微小颗粒、模型微生物和小分子化合物的传输。在不同的pH和钙浓度下，颗粒、微生物和化合物将暴露在含有食物相关脂的介质中的粘液表面以及模型禁食状态的肠道缓冲液中。这些分析将在天然粘液和纯化的粘蛋白凝胶上进行，以深入了解粘液结构成分对特定食物相关刺激的反应。同时进行的微观和宏观流变学分析将表明，在观察到的传输现象中，粘液凝胶结构变化与颗粒/分子和凝胶成分之间发生的相互作用的相对重要性。还将使用先进的显微技术，包括快速冷冻深度蚀刻显微镜(QF/DEM)，检查与食品相关的脂类暴露和物理化学环境变化相关的粘液凝胶的结构变化。将使用EPR和氮氧化物探针标记的胆盐和脂肪来检查粘液层内食物相关脂形成的胶体结构的完整性，因为目前尚不清楚这些结构是否在粘液中保持完整，这可能会显著影响通过这种天然水凝胶的运输性质。跨学科研究团队拥有成功了解和开始利用脂质对胃肠道粘液屏障的影响所需的专业知识，其中包括PI，一名在药物输送和粘液传输现象方面有专长的化学工程师，一名在病原体通过粘液和粘液神学转运方面有专长的生物化学家，一名在生物基质结构高级微观分析方面有专长的机械工程师，以及一名在膜小分子流动性电子顺磁共振分析方面有专长的化学家。

DESCRIPTION (provided by applicant): The overall goal of this project is to elucidate the impact of lipids and food-associated physicochemical changes in the gastrointestinal (GI) tract on intestinal mucus barrier properties. This information will motivate strategies for mucosal barrier control, enabling efficient drug carrier systems (e.g. mucosal vaccines), enhanced nutrition, and potentially inhibited pathogen invasion. Gastrointestinal mucus is a natural hydrogel providing a finely-tuned and amazingly selective barrier, protecting the underlying epithelium from harsh physicochemical changes in the intestinal lumen (e.g. elevations in bile salt concentration, lowered pH), selectively inhibiting microbial transport, and enabling efficient absorption of nutrients and water. Despite the significance of mucus's role and potential implications in health and disease, its barrier properties are relatively poorly understood. Preliminary data indicates that intestinal mucus properties are significantly modulated by food-associated lipids and physicochemical changes associated with food (i.e. changes in pH and [Ca+2 ]). To study these phenomena and enable design of strategies to exploit them for therapeutic purposes, the transport of micro-particulates, model microbes and small molecular weight compounds will be analyzed using multiple particle tracking techniques and a novel application of electron paramagnetic resonance (EPR). Particles, microbes and compounds will be exposed to mucus surfaces in media containing food-associated lipids as well as model fasted state intestinal buffer, at varied pH and Ca+2 concentrations. These analyses will be performed on both native mucus as well as purified mucin gels to provide insight into mucus structural components responding to specific food-associated stimuli. Micro- and macro-rheological analyses performed in parallel will indicate the relative significance of mucus gel structural changes vs. interactions occurring between particles/molecules and gel constituents in observed transport phenomena. Structural changes in mucus gels associated with exposure to food-associated lipids and physicochemical environmental changes will also be examined using advanced microscopic techniques, including quick-freeze deep etch microscopy (QF/DEM). The integrity of colloidal structures formed by food-associated lipids within the mucus layer will be examined using EPR and nitroxide-probe labeled bile salts and lipids, as it is currently not understood whether these structures stay intact within mucus, which could significantly impact the nature of transport through this natural hydrogel. The interdisciplinary research team possesses the expertise required to successfully understand and begin to exploit the impact of lipids on the GI mucus barrier, including the PI, a chemical engineer with expertise in transport phenomena in drug delivery and mucus, a biochemist with expertise in pathogen transport through mucus and mucus theology, a mechanical engineer with expertise in advanced microscopic analysis of biological matrix structure, and a chemist with expertise in electron paramagnetic resonance analysis of small molecule mobility in membranes.