Atherosclerotic Risk of Branched Chain Amino Acids in a Tissue Engineered Blood Vessel Model

组织工程血管模型中支链氨基酸的动脉粥样硬化风险

基本信息

批准号：
10536528
负责人：
Ellery Jensen Jones
金额：
$ 3.95万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10536528
关键词：
Address Adhesions Affect Amino Acids Arterial Fatty Streak Atherosclerosis Autophagocytosis Autophagosome Biological Biological Assay Biological Models Blood Blood Vessels Branched-Chain Amino Acids Cardiovascular Diseases Cardiovascular system Cause of Death Cells Chondroitin Sulfates Clinical Collaborations Collagen Conflict (Psychology)Dermal Development Diabetes Mellitus Disease Disease Progression Disease model Encapsulated Endothelium Environment Event Extracellular Matrix Fibroblasts Foam Cells Functional disorder Glucose Goals Health Heart Diseases Heart failure Human Hydrogels Impairment Incidence Inflammation Inflammatory Insulin Resistance Isoleucine Lesion Leucine Leukocytes Link Lipids Lipoprotein Binding Low-Density Lipoproteins Medial Mediating Metabolic Metabolic Diseases Metabolic syndrome Minority Groups Mitochondria Modeling Molecular Myocardial Infarction Onset of illness Orthopedics Outcome Study Oxidative Stress Pathologic Pathology Patients Phenotype Physiological Physiology Prevalence Proteins Proteoglycan Reactive Oxygen Species Recovery Research Risk Role Serum Stress Stroke System Testing Time Tissue Engineering United States Valine Vasodilation Western Blotting Woman amino acid metabolism atherogenesis atherosclerosis risk cardiogenesis cardiovascular risk factor cell growth chondroitin sulfate glycosaminoglycan cytokine disability disease phenotype endothelial dysfunction human tissue in vitro Model interest low and middle-income countries member metabolomics monocyte mouse model negative affect neonatal human novel novel therapeutics oxidized low density lipoprotein polysulfated glycosaminoglycan prevent therapeutic target vascular inflammation

项目摘要

Elevated levels of the branched chain amino acids (BCAA) leucine, isoleucine, and valine are highly correlated with the development of CVD and adverse cardiovascular events, such as heart attacks. Some mechanisms linking BCAA, metabolic disease and heart failure have been documented. However, it remains unclear if BCAAs directly interact with vascular cells to enable CVD onset and progression. Mouse models that have been developed to study CVD and BCAA both have limitations in their applicability to human physiology. Current in vitro models of atherosclerosis are limited to early stages of atherogenesis, and it is challenging to recapitulate more advanced atherosclerotic changes. To address this issue, we will develop a model of an intermediate atherosclerotic lesion using a human tissue engineered blood vessel system (TEBV) model with a disease-pathology collagen extracellular matrix enriched with the glycosaminoglycan chondroitin sulfate (CS), treated with modified low-density lipoprotein (LDL). We will use collagen TEBVs treated with modified LDL as a model for early atherosclerosis. These platforms will be used as a platform to study BCAA mechanism in CVD. The TEBV model is made of collagen or CS-collagen vessels with encapsulated human neonatal dermal fibroblasts serving as the medial cells in the vascular wall and an endothelialized inner lumen of endothelial colony-forming cells (ECFCs). It is perfused with medium at a rate of 2 mL/minute and can be cultured for up to 6 weeks. We have demonstrated that the CS-enriched vessels have an enhanced sensitivity to the inflammatory effects of modified LDL and demonstrate increased vessel vasoactive dysfunction and endothelial-leukocyte interactions compared to collagen TEBVs. We have also demonstrated that elevations of the BCAA within the physiological milieu are sufficient to induce an atherosclerotic phenotype in the endothelium. Preliminary results demonstrate that treatment of ECFCs with elevated BCAA and oxidized low- density lipoprotein causes increased mitochondrial oxidative stress, as well as decreased expression of LC3B, an autophagosome protein. This induced early atherosclerotic events in the TEBV system: impaired endothelium-controlled vasodilation and leukocyte adhesion to the endothelium. Autophagic flux will be further assayed via Western Blot for LC3-I to LC3-II conversion. Vascular cells, especially the endothelium, may be sensitive to the effects of elevated BCAA because they do not significantly contribute to BCAA metabolism, leading to intracellular buildup of metabolites and mitochondrial stress. This will be tested via metabolomic analysis of BCAA and their downstream metabolites in the vascular cells. Using the TEBVs, we will determine the role of BCAA in early versus intermediate atherosclerosis. At the end of the study, we will have an intermediate lesion model of atherosclerosis, the mechanisms linking BCAA to cardiovascular events will become clearer, and potential therapeutic targets for atherosclerosis will be identified.

分支链氨基酸（BCAA）亮氨酸，异亮氨酸和瓣膜的升高水平高度相关随着CVD和不良心血管事件的发展，例如心脏病发作。一些机制已经记录了联系BCAA，代谢疾病和心力衰竭的信息。但是，尚不清楚是否 BCAA直接与血管细胞相互作用，使CVD发作和进展。具有的鼠标模型已开发用于研究CVD和BCAA都对人类生理的适用性都有局限性。当前的动脉粥样硬化体外模型仅限于动脉粥样硬化的早期阶段，并且具有挑战性概括更先进的动脉粥样硬化变化。为了解决这个问题，我们将开发一个模型使用人体组织工程血管系统（TEBV）模型的中间动脉粥样硬化病变疾病病理学胶原蛋白外基质富含糖胺聚糖软骨素硫酸盐（CS），，，用改良的低密度脂蛋白（LDL）处理。我们将使用经过修改的LDL处理的胶原蛋白TEBV作为早期动脉粥样硬化的模型。这些平台将用作研究CVD中BCAA机制的平台。 TEBV模型由胶原蛋白或CS-胶原蛋白容器制成成纤维细胞用作血管壁中的内侧细胞和内皮的内皮化内腔菌落形成细胞（ECFC）。它以2 ml/分钟的速度灌注培养基，可以培养 6周。我们已经证明，富含CS的血管对修饰的LDL的炎症作用，表明血管活性功能障碍增加和与胶原蛋白TEBV相比，内皮 - 白细胞相互作用。我们还证明了生理环境中的BCAA足以在诱导动脉粥样硬化表型中内皮。初步结果表明，用BCAA升高并氧化低 - 的ECFC处理密度脂蛋白会导致线粒体氧化应激增加，并降低LC3B的表达自噬体蛋白。这引起了TEBV系统中的早期动脉粥样硬化事件：受损内皮控制的血管舒张和白细胞粘附于内皮。自噬通量将进一步通过Western blot分析LC3-I为LC3-II转换。血管细胞，尤其是内皮细胞，可能是对BCAA升高的影响敏感，因为它们没有显着促进BCAA代谢，导致细胞内积聚代谢物和线粒体应激。这将通过代谢组进行测试分析血管细胞中BCAA及其下游代谢产物。使用TEBV，我们将确定 BCAA在早期与中间动脉粥样硬化中的作用。在研究结束时，我们将有一个动脉粥样硬化的中间病变模型，将BCAA与心血管事件联系起来的机制将变得更清晰，将确定动脉粥样硬化的潜在治疗靶标。