The role of sphingosine kinase 2 in mitochondrial dysfunction and NAFLD

鞘氨醇激酶 2 在线粒体功能障碍和 NAFLD 中的作用

• 批准号: 10607244
• 负责人: Kaitlyn Georgene Jackson
• 金额: $ 4.13万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-10 至 2027-04-09
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10607244
• 关键词: Adenovirus Vector; Adult; Affect; Anti-Inflammatory Agents; Bile Acids; Biochemical; Biological; Biology; Cardiac Myocytes; Cell Nucleus; Cell physiology; Cells; Clinical; Complex; Data; Disease; Disease Progression; Drug Targeting; Evaluation; Exhibits; Exposure to; FDA approved; Fatty Liver; Flow Cytometry; Future; Genes; Genetic Transcription; Goals; Hepatic; Hepatocyte; Histone Deacetylase Inhibitor; Human; Immune; Immune response; In Vitro; Incidence; Inflammation; Inflammatory; Inflammatory Response; Innate Immune Response; Intervention; Knockout Mice; Knowledge; Kupffer Cells; Link; Lipids; Literature; Liver; Location; Malignant Neoplasms; Measures; Mediator; Membrane Potentials; Metabolic Diseases; Metabolic stress; Metabolism; Mitochondria; Modeling; Molecular; Nuclear; Ontology; Oxidative Stress; Palmitic Acids; Pathogenesis; Patients; Pattern; Peripheral; Pharmacotherapy; Population; Prevalence; Protein Isoforms; Reactive Oxygen Species; Regulation; Reporting; Risk; Role; Saturated Fatty Acids; Signal Transduction; Signaling Molecule; Source; Sphingolipids; Sphingosine-1-Phosphate Receptor; System; Testing; Up-Regulation; Variant; Wild Type Mouse; complex IV; cytochrome c; cytochrome c oxidase; cytokine; end stage liver disease; endoplasmic reticulum stress; fatty acid oxidation; hepatocyte injury; immune cell infiltrate; immunoregulation; improved; in vivo; innovation; lipid metabolism; liver inflammation; mitochondrial dysfunction; mitochondrial membrane; mouse model; new therapeutic target; non-alcoholic fatty liver disease; nonalcoholic steatohepatitis; novel; novel therapeutic intervention; overexpression; pharmacologic; prevent; prohibitin; protective effect; response; sphingosine 1-phosphate; sphingosine kinase; trafficking; transcriptome sequencing; western diet

Non-alcoholic fatty liver disease (NAFLD) is a fast-growing disease with no FDA-approved treatments. Numerous studies have linked NAFLD-associated hepatocyte lipotoxicity with mitochondrial dysfunction during the last few decades. Though the onset of inflammation that drives the progression from NAFL to NASH is considered multifactorial, damage-associated molecular patterns (DAMPs) and reactive oxygen species (ROS) released by injured hepatocytes are essential in activating Kupffer cells and promoting peripheral immune cell infiltration. Sphingolipids are biochemical signaling mediators and can regulate mitochondrial function in metabolic diseases, including NAFLD. Sphingosine-1-phosphate (S1P) is a well-studied molecule with dualistic roles in cell function, dependent on the location of the sphingosine kinase (SphK) isoforms, SphK1 and SphK2. While SphK1- generated cytosolic S1P signaling is thoroughly described in the literature, few studies assess the role of SphK2- derived S1P. SphK2 is localized to the cell nucleus and mitochondria. In the context of NAFLD, nuclear SphK2 is anti-inflammatory by inhibiting histone deacetylases (HDACs) and preventing the transcription of proinflammatory genes. In contrast, the role of SphK2 within the mitochondria is not well defined. Previous in vitro studies evaluated the effect of cytosolic S1P, rather than mitochondrial S1P, on mitochondrial function. While SphK2 function has been assessed in hepatocytes, these studies have only defined short-term effects of SphK2 on hepatic steatosis. Our long-term goal is to understand sphingolipid regulation and define the mechanisms that influence NAFLD disease progression. The central hypothesis of this application is that dysregulation of mitochondrial SphK2 promotes NAFLD disease progression. Two specific aims are proposed to test the hypothesis. Aim 1 examines the role of SphK2 in regulating mitochondrial function in hepatocytes under metabolic stress. We will determine if exposure to palmitic acid alters mitochondrial sphingolipid metabolism in hepatocytes and further assess the impact of SphK2 depletion on mitochondrial complex IV function and assembly in hepatocytes. Aim 2 determines the impact of hepatic SphK2 on modulating the immune response under metabolic stress. First, we will measure the ability of SphK2 depleted hepatocytes to induce cytokine synthesis in Kupffer cells. Second, we will use spectral flow cytometry to determine SphK2- dependent variations on the hepatic immune cell profile in the WDSW model. Lastly, we will use an adenoviral vector system to determine whether overexpressed hepatocyte SphK2 exhibits protective effects in NAFLD pathogenesis. To date, these functions of hepatocyte SphK2 have not been defined in the literature. Completing our proposed study will contribute to our understanding of SphK2 in hepatocyte mitochondrial function and highlight how SphK2 influences inflammation in NAFLD. Our novel findings are essential to fundamental hepatocyte biology and may influence future studies targeting sphingolipid signaling in developing innovative pharmacotherapies for NAFLD.

非酒精性脂肪肝（NAFLD）是一种快速发展的疾病，没有FDA批准的治疗方法。许多 在过去的几年中，研究已经将NAFLD相关的肝细胞脂毒性与线粒体功能障碍联系起来， 几十年尽管认为炎症的发作驱动了从NAFL到NASH的进展， 多因子损伤相关分子模式（DAMPs）和活性氧（ROS）的释放， 受损的肝细胞在激活枯否细胞和促进外周免疫细胞浸润中是必需的。 鞘脂是一种生化信号介质，在代谢过程中可以调节线粒体功能， 疾病，包括NAFLD。鞘氨醇-1-磷酸（S1 P）是一种在细胞中具有双重作用的分子， 功能，取决于鞘氨醇激酶（SphK）亚型SphK 1和SphK 2的位置。而SphK 1- 虽然文献中对SphK 2 - 1产生的胞浆S1 P信号转导进行了全面描述，但很少有研究评估SphK 2 - 1的作用。 S1 P的。SphK 2定位于细胞核和线粒体。在NAFLD的背景下，核SphK 2 通过抑制组蛋白脱乙酰酶（HDAC）和阻止 促炎基因相反，SphK 2在线粒体中的作用尚未明确。先前的 体外研究评价了胞质S1 P而不是线粒体S1 P对线粒体功能的影响。 虽然已经在肝细胞中评估了SphK 2的功能，但这些研究仅确定了SphK 2的短期作用。 SphK 2对肝脂肪变性的影响。我们的长期目标是了解鞘脂的调节， 影响NAFLD疾病进展的机制。本申请的中心假设是， 线粒体SphK 2的失调促进NAFLD疾病进展。两个具体目标是 来检验这个假设。目的1研究SphK 2在调节线粒体功能中的作用， 代谢应激下的肝细胞我们将确定是否暴露于棕榈酸改变线粒体 肝细胞中的鞘脂代谢，并进一步评估SphK 2耗竭对线粒体 复合物IV在肝细胞中的功能和组装。目的2确定肝SphK 2对调节 代谢应激下的免疫反应首先，我们将测量SphK 2耗尽的肝细胞的能力， 以诱导枯否细胞中的细胞因子合成。其次，我们将使用光谱流式细胞术来确定SphK 2- WDSW模型中肝脏免疫细胞谱的依赖性变化。最后，我们将使用腺病毒 载体系统，以确定过表达的肝细胞SphK 2是否在NAFLD中表现出保护作用 发病机制迄今为止，肝细胞SphK 2的这些功能尚未在文献中定义。完成 我们提出的研究将有助于我们理解SphK 2在肝细胞线粒体功能中的作用， 强调SphK 2如何影响NAFLD中的炎症。我们的新发现对我们的基础研究至关重要 肝细胞生物学，并可能影响未来的研究，针对鞘脂信号转导，在开发创新 NAFLD的药物治疗。