Deciphering a Novel LncRNA-mediated Lipid Droplet Transport System in Human Heart

摘要 这项提案的首要目标是揭示新的人类特异性分子机制， 人长链非编码RNA（lncRNA）HL 6促进人心肌细胞（CM）中脂滴（LD）转运 并显示出对代谢综合征相关心肌病的心脏保护作用。约70%的能量 成人心脏是从长链脂肪酸（FA）的氧化中获得的。平衡的脂质代谢至关重要 心脏功能。脂质代谢受损已在心肌病相关性中广泛观察到。 代谢综合征，如肥胖症和糖尿病。肥胖与糖尿病相关的显著特征 心肌病是心力衰竭发生率增加以及广泛的肌细胞内脂滴 积累LD由被磷脂单层包围的中性脂质核心组成，磷脂单层起关键作用。 在过量脂肪酸储存和动员中的作用。然而，超负荷的肌细胞内LD可引起心脏 人类患者以及肥胖和糖尿病啮齿动物模型的脂毒性和功能障碍。虽然充足 有证据表明，LD在CM中的积累可能是由于LD转运受损， LD在哺乳动物心脏中的转运仍然是难以捉摸的。特别地，CM的人类特异性LD转运系统 其在代谢综合征相关的心肌病和心力衰竭中的意义仍然未知。 最近，我们鉴定了一种人lncRNA，心脏lncRNA 6（HL 6），它在人心肌细胞中高度特异性表达。 人CM和下调人2型糖尿病心肌。HL 6基因敲除（HL 6 KO）， 通过使用CRISPR/Cas-9对人iPSC（hiPSC）进行扩增。我们发现HL 6缺乏导致广泛的LD 蓄积，LD向线粒体的转运缺陷，线粒体肿胀功能受损，以及 HL 6 KO hiPSC-CM相对于WT hiPSC-CM的CM死亡增强。有趣的是，HL 6结合LD， hiPSC-CM中的细胞骨架。此外，转基因HL 6过表达显著减轻了高脂饮食， （HFD）诱导的脂质蓄积和小鼠心脏的心功能障碍。因此，我们假设HL 6 在人CMs中，HL 6在LD转运到线粒体中起着不可或缺的作用，并且HL 6的获得表现出一种免疫调节作用。 对代谢综合征相关心肌病的心脏保护作用。提出了两个具体目标： 具体目标1：破译HL 6在人CMs脂滴转运中不可或缺的作用。具体目标二： 确定HL 6在代谢综合征相关心肌病中的心脏保护作用。

Abstract The overarching goal of this proposal is to uncover novel HUMAN-SPECIFIC molecular mechanisms by which human long non-coding RNA (lncRNA) HL6 facilitates lipid droplet (LD) transport in human cardiomyocytes (CMs) and manifests a cardioprotective role against metabolic syndrome associated cardiomyopathy. ~70% energy of adult human heart is obtained from oxidation of long chain fatty acids (FAs). Balanced lipid metabolism is crucial for heart function. Compromised lipid metabolism have been widely observed in cardiomyopathies associated with metabolic syndromes, such as obesity and diabetes. The distinct features of obese and diabetes associated cardiomyopathy are increased incidence of heart failure together with extensive intramyocyte lipid droplet accumulation. LD consists of a neutral lipid core surrounded by a phospholipid monolayer, which plays a critical role in excess fatty acid storage and mobilization. However, overwhelmed intramyocyte LDs can cause cardiac lipotoxicity and dysfunction in both human patients and rodent models of obesity and diabetes. Although ample evidence suggests that LD accumulation in CMs could be due to impaired LD transport, the cellular system for LD transport in mammalian heart remains elusive. Particularly, the human-specific LD transport system of CMs and its implication in metabolic syndrome associated cardiomyopathy and heart failure remain unknown. Recently, we identified a human lncRNA, Heart LncRNA 6 (HL6), which is highly and specifically expressed in human CMs and downregulated in human type 2 diabetic myocardium. HL6 was knocked out (HL6KO) in human iPSCs (hiPSCs) by using CRISPR/Cas-9. We found HL6 deficiency led to extensive LD accumulation, defective LD transport to mitochondria, swollen mitochondria with impaired function, and enhanced CM death in HL6KO hiPSC-CMs verse WT hiPSC-CMs. Intriguingly, HL6 binds LDs and cytoskeleton in hiPSC-CMs. Additionally, transgenic HL6 overexpression significantly mitigated high fat diet (HFD)-induced lipid accumulation and cardiac dysfunction of mouse heart. Therefore, we hypothesize that HL6 plays an indispensable role in LD transport to mitochondria in human CMs, and gain-of-HL6 manifests a cardioprotective role against metabolic syndrome associated cardiomyopathy. Two specific aims are proposed: Specific Aim 1: Decipher the indispensable role of HL6 in lipid droplet transport of human CMs. Specific Aim 2: Determine the cardioprotective role of HL6 in metabolic syndrome associated cardiomyopathy.