Synergistic Effects of Medium-chain and Omega-3 Triglycerides on Cellular Omega-3 Fatty Acid Enrichment: Characterization, Mechanisms, and Optimization

不良的炎症反应导致细胞死亡和器官功能障碍。我们将研究急性静脉（IV）治疗含有中链甘油三酯（TG） （MCT）和富含二十碳五烯酸（EPA）和二十二碳六烯酸（DHA）的ω -3 (n-3) TG的脂质乳是否提供了一种新的治疗方法，通过靶向快速细胞n-3 FA膜富集来改善不良炎症反应。n-3脂肪酸（FA）越来越被认为是静脉喂养中的一种有效补充剂，因为它具有促进愈合和抗细胞凋亡、自由基和促炎类二十烷的产生的作用。实验动物模型研究表明，大量注射n- 3tg -富乳剂颗粒（TGRP）对降低脑卒中后脑死亡和发病率具有治疗保护作用。我们发现含有MCT和鱼油TG （8:2 wt/wt）的TGRP比其他TGRP更有效地富集白细胞和内皮细胞的n-3 FA细胞膜。利用核磁共振（NMR），我们报道了MCT纳入模型TGRP可促进模型膜脂紊乱，从而可能加速n-3的递送速率，突出了静脉注射富集n-3 FA的优势。然而，很少有研究涉及急性静脉注射n-3的急性抗炎作用。目前，没有数据表明MCT与n- 3tg的最佳混合物可以实现最快速和最高强度的抗炎作用。我们的中心假设是，同时注射含有MCT和n-3 TG的TGRPs会共同影响膜动力学，加速n-3 FA进入细胞膜，从而启动n-3介导的抗炎和抗凋亡作用。这些将在三个具体目标下进行研究。在Aim 1中，我们将使用生物物理方法结合基于质谱的脂质组学来表征含有不同n-3 TG含量的MCT- 3 TGRP的生物物理特性，以评估含有DHA、EPA或两者的特定MCT:n-3 TGRP介导的膜结构和活性的改变。在Aim 2中，我们将在培养的免疫效应巨噬细胞和小鼠模型中定义负责内化和代谢利用特异性MCT的细胞摄取途径：在Aim 1中表征的n-3 TGRP。然后，我们将在Aim 3下研究MCT:n-3 TGRP细胞内化如何影响小鼠模型中单核细胞和巨噬细胞对急性免疫刺激（LPS）的功能活动。在细胞和动物模型中提出的研究将建立创新的方法，适用于将亲脂和疏水治疗剂靶向、快速递送到特定器官或组织。我们将发现MCT和n-3 TG之间的新协同作用，调节膜脂组成，细胞脂质代谢和免疫反应。我们的研究将对许多领域具有重要意义，包括感染和炎症相关的组织损伤和器官功能障碍的免疫反应。

Adverse inflammatory response leads to cell death and organ dysfunction. We will investigate whether acute intravenous (IV) treatments of lipid emulsions containing medium-chain triglycerides (TG) (MCT) and omega-3 (n-3) TG enriched in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) provide a new therapeutic approach to ameliorate adverse inflammatory responses by targeting rapid cellular n-3 FA membrane enrichment. n-3 fatty acids (FA) are increasingly recognized as a potent supplement in IV feeding because of their pro-healing and anti-inflammatory effects against apoptosis and production of free radicals and pro- inflammatory eicosanoids. Studies in experimental animal models demonstrate that bolus injections of n-3 TG- rich emulsion particles (TGRP) are therapeutically protective in decreasing brain death and morbidity after stroke. We have discovered that TGRP containing both MCT and fish oil TG (8:2 wt/wt) enriched cell membranes of white blood cells and endothelial cells with n-3 FA more efficiently than other TGRP. Using nuclear magnetic resonance (NMR), we reported that the inclusion of MCT into model TGRP promotes model membrane lipid disorder that may accelerate n-3 delivery rates, highlighting the advantage of bolus injection for n-3 FA enrichment. However, very few studies have addressed the acute anti-inflammatory effects of acute IV n-3 administration. Currently, there is no data on optimal mixtures of MCT with n-3 TG to achieve the most rapid and highest-magnitude anti-inflammatory action. Our central hypothesis is that bolus injections of TGRPs containing both MCT and n-3 TG will cooperatively affect membrane dynamics to accelerate n-3 FA incorporation into cell membranes, which will initiate n-3-mediated anti-inflammatory and anti-apoptotic actions. These will be studied under 3 Specific Aims. Under Aim 1, we will characterize biophysical properties of MCT-containing TGRP with different n-3 TG contents using biophysical approaches coupled with mass spectrometry-based lipidomics to assess alteration in membrane structure and activity mediated by specific MCT:n-3 TGRP containing DHA, EPA, or both. In Aim 2, we will define cellular uptake pathways responsible for internalization and metabolic utilization of specific MCT:n-3 TGRP characterized in Aim 1 in cultured immune-effector macrophages and in mouse models. We will then investigate under Aim 3 how MCT:n-3 TGRP cellular internalization affects functional activities of monocytes and macrophages in response to an acute immune stimulus (LPS) in mouse models. The proposed studies in cells and animal models will establish innovative methods that are adaptable for targeted, rapid delivery of lipophilic and hydrophobic therapeutic agents to specific organs or tissues. We will uncover novel synergies between MCT and n-3 TG that modulate membrane lipid composition, cell lipid metabolism, and immune responses. Our studies will be of high significance to many fields, including immune responses to infection and inflammation-related tissue injury and organ dysfunction.