Transendothelial transport and CD36 in the dysregulated lipid trafficking of failing hearts

项目总结： 这项拟议的研究探索了心脏衰竭患者体内脂代谢失调的机制。 长链脂肪酸(LCFA)从循环到代谢的动态过程 心肌细胞(CM)。我们将探索内皮细胞(EC)递送LCFA对CMS的作用以及 CMS摄取LCFA在心力衰竭小鼠模型中的不利代谢重塑 人体心脏衰竭的代谢缺陷。心脏依赖LCFA氧化，高达70%的燃料，以满足ATP 需求。LCFA还被酯化到中性甘油三酯(TG)池中，并生成具有生理活性的酰基- 衍生品。在衰竭的心脏中，LCFA氧化减少，血脂谱变得有毒。我们已经展示了一个 心力衰竭患者中心LCFA代谢物酰基-辅酶A减少是有害的，而酰基增加 ACSL1产生辅酶A可改善代谢状态，减轻功能衰退。心脏的13C核磁共振 揭示了~(13)C LCFA进入TG的指数分量，它反映了LCFA的摄取速度，并且是敏感的 与LCFA转运蛋白CD36的活性有关。LCFA的摄取也通过代谢捕获来加速 在ACSL1的作用下，LCFA酯化为酰基辅酶A，这是一个与性别有关的过程。在小鼠身上进行的细胞实验- 具体而言，ECs(EC-CD36 KO)和CMS(CM-CD36 KO)中的CD36缺失将支持研究 竞争中ECs向CMS输送LCFA和CMS吸收LCFA对LCFA代谢的影响 途径，包括心脏衰竭时形成有害的神经酰胺。LCFA摄取和LCFA吸收的潜在差异 白蛋白结合的LCFA和脂蛋白结合的甘油三酯这两个主要生理来源之间的代谢将 被研究。假设是：a)EC CD36依赖和非依赖的跨内皮细胞的贡献 LCFAs进入正常和疾病心脏CMS的转运决定了LCFAs的代谢命运 来自CM CD36的活性，并取决于LCFA来源；b)这两种CD36都存在性别差异 LCFA的转运和通过酯化作用将LCFAs捕获到CMS中，这有助于CMS的脂毒性 心脏衰竭。具体目标是：1.阐明EC CD36在LCFA血管内皮细胞转运中的作用 雄性EC-CD36 KO小鼠与雌性EC-CD36 KO小鼠正常和衰竭心脏的摄取动力学和代谢命运。2. 区分CM-CD36和EC对正常人心脏LCFA摄取动力学和代谢去向的贡献 以及雄性CM-CD36 KO小鼠与雌性CM-CD36 KO小鼠的体温下降。3.阐明CD36转运和CD36的相互作用 ACSL1对LCFA在正常和衰竭心脏中的代谢捕获作用，通过沉默心脏中的CD36 ACSL1(MHC-ACSL1 J3)及杂交、MHC-ASCL1xEC-CD36 KO和 MHC-ASCL1xCM-CD36 KO小鼠。4.区分EC-CD36和CM-CD36对LCFA摄取的贡献 正常和衰竭野生心脏白蛋白结合来源与乳糜粒结合来源的比率和代谢 型，EC-CD36 KO和CM-CD36 KO小鼠。结果将提供关于分子事件的独特信息 调节脂质含量作为解决衰竭心脏代谢失衡的新靶点。

Project Summary: The proposed research explores mechanisms of dysregulated lipid metabolism in failing hearts within the dynamic processes of long chain fatty acid (LCFA) delivery from the circulation to metabolism in the cardiomyocyte (CM). We will explore the roles of LCFA delivery by endothelial cells (EC) to CMs and rates of LCFA uptake by CMs in adverse metabolic remodeling in a mouse model of heart failure that recapitulates key metabolic defects in failing human hearts. Hearts rely on LCFA oxidation, up to 70% of fuels, to meet ATP demand. LCFA are also esterified into the neutral triglyceride (TG) pool and into physiologically active acyl- derivatives. In failing hearts, LCFA oxidation is reduced and the lipid profile becomes toxic. We have shown a reduction in the central LCFA metabolite, acyl-CoA, in failing hearts is detrimental, and increased acyl CoA production by ACSL1 improves metabolic state and mitigates functional decline. 13C NMR of hearts revealed an exponential component of 13C LCFA entry into TG that reflects LCFA uptake rate and is sensitive to activity of the LCFA transporter, CD36. LCFA uptake is also accelerated by metabolic trapping via esterification of LCFA to acyl-CoA by ACSL1, a process that is sex-dependent. Experiments on mice with cell- specific, CD36 deletion in ECs (EC-CD36 KO) and CMs (CM-CD36 KO) will support the objective to study the separate roles of LCFA delivery by ECs to CMs and uptake by CMs on LCFA metabolism within competing pathways, including deleterious ceramide formation in failing hearts. Potential differences in LCFA uptake and metabolism between two primary physiological sources, albumin-bound LCFA and lipoprotein-bound TG, will be studied. The hypothesis is: a) the contributions of EC CD36-dependent and independent transendothelial transport of LCFAs into CMs of normal and diseased hearts determine the metabolic fate of LCFAs, separate from CM CD36 activity, and depend on the LCFA source; b) there are sex-dependent differences in both CD36 transport of LCFA and trapping of LCFAs into CMs via esterification that contribute to the lipotoxic profile of failing hearts. Specific aims are: 1. Elucidate EC CD36 contributions to transendothelial transport of LCFA uptake kinetics and metabolic fate in normal and failing hearts of male vs. female EC-CD36 KO mice. 2. Distinguish CM-CD36 from EC contributions to LCFA uptake kinetics and metabolic fate in hearts of normal and failing heats of male vs. female CM-CD36 KO mice. 3. Elucidate reciprocal effects of CD36 transport and metabolic trapping by ACSL1 on LCFA use in normal and failing hearts, by silencing of CD36 in hearts having low overexpression of ACSL1 (MHC-ACSL1 J3) and in hearts from crossed, MHC-ASCL1xEC-CD36 KO and MHC-ASCL1xCM-CD36 KO mice. 4. Distinguish contributions of EC-CD36 and CM-CD36 to LCFA uptake rates and metabolism from albumin-bound vs. chylomicron-bound sources in normal and failing hearts of wild type, EC-CD36 KO, and CM-CD36 KO mice. Outcomes will provide unique information on molecular events modulating lipid content as new targets to resolve metabolic imbalances in failing hearts.