Lipogenesis in the meibomian glands and adnexa in the norm and pathology

正常和病理情况下睑板腺和附件的脂肪生成

• 批准号: 9762911
• 负责人: Igor A Butovich
• 金额: $ 40.5万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-30 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9762911
• 关键词: Adhesions; Affect; Aftercare; Age; Aging; Anabolism; Antibodies; Biochemical; Biochemistry; Cell surface; Characteristics; Chemicals; Cholesterol; Complex; Development; Disease; Dose; Dyslipidemias; Enzymes; Equilibrium; Eye diseases; Fatty Acids; Film; Functional disorder; Gender; Gene Expression; Gene Expression Profile; Generations; Genomics; Global Change; Goals; Grant; Health; High Pressure Liquid Chromatography; Human; Immunohistochemistry; Impairment; In Situ; In Situ Hybridization; Incidence; Intake; Intervention; Lead; Link; Lipids; Mammals; Maps; Mass Spectrum Analysis; Medical; Menopausal Status; Metabolic Diseases; Microscopy; Molecular; Morphology; Mus; Nutritional Study; Ocular Pathology; Ocular Physiology; Pathology; Pathway interactions; Patients; Pharmacologic Substance; Phosphatidylcholine-Sterol O-Acyltransferase; Physiology; Polarization Microscopy; Polysaccharides; Property; Protein Analysis; Proteins; Proteomics; Quality of life; RNA analysis; Regulation; Reporting; Research; Role; Rupture; Sampling; Scheme; Severities; Stains; Sterol O-Acyltransferase; Structure; Testing; Tissues; Vision; Western Blotting; Woman; Work; amphiphilicity; base; biophysical properties; biophysical techniques; blood lipid; clinical Diagnosis; experimental study; eye dryness; immunocytochemistry; innovation; lipid biosynthesis; lipid metabolism; meibomian gland; meibomian gland dysfunction; melting; metabolic abnormality assessment; metabolomics; microcalorimetry; ocular surface; public health relevance

Research Abstract The goal of this project is to elucidate the enzymatic pathways that underlie the biosynthesis of lipids that are produced by human meibomian glands (HMG) and are collectively known as meibum. Meibum is a critically important part of the tear film (TF) that, among other functions, protects the ocular surface from desiccating. Meibum is formed predominantly from a unique mixture of very, and extremely, long chain complex lipids. Our previous projects were devoted to biochemical and biophysical characterization of meibomian lipids (ML). Now we propose to investigate the biosynthetic pathways that lead to the formation of meibum in HMG. The following three lines of research are proposed: (I). Despite recent progress in the characterization of the lipidomes of meibum and TF, there is no clear understanding of the biosynthetic path- ways involved in the biosynthesis of meibum. The unusual and, often, unique qualities of ML imply their in situ and/or de novo origination, which, in turn, requires proper molecular mechanisms to be implemented in HMG. We will study the levels of expression, and map, major lipid-metabolizing enzymes and related proteins, which, based on our preliminary studies, are expected to be present in HMG. This will be achieved by using a combination of approaches, such as microarray RNA analyses, immunohistochemistry/immunocytochemistry, in-situ hybridization, proteomics, and lipidomics. (II). Recent reports indicate that high total blood lipid levels, including cholesterol and TAG, are correlated with higher incidence and/or severity of MG dysfunction. Also, a positive impact of ω3 fatty acids (ω3FA) supplements on the ocular surface was reported for humans and mice. Yet, the reasons and molecular mechanisms of these effects are unknown. We will determine the balance between the in situ/de novo biosynthesis of ML and the levels and composition of blood lipids. We will test if 1) enhanced intake of pharmaceutical doses of ω3FA, and (2) use of statins, can have a positive impact on the biosynthesis, composition and properties of meibum. (III). Recently, we reported that: (1) up to 20% of normal meibum is formed of non-lipid components; (2) there are considerable differences between normal and DE meibum: the latter is enriched with non-lipid material, and forms lipid layers that are less structurally stable than those formed of healthy meibum. Based on those findings, we propose to assess the non-lipid content of mei- bum using: (a) metabolomic approaches based on HPLC-MS, GC-MS, and 2D-DESI MS; (b) Western blot with specific antibodies against major meibum and TF proteins; (c) histochemical approaches targeting proteins and polysaccharides; (d) various biophysical methods, such as hot stage polarized light microscopy, microcalori- metry, Brewster angle microscopy, to study rheological and melting characteristics of meibum and tear film lipids. We believe that our work will provide innovative information that may lead to new concepts in medical treatment of MG-related diseases and conditions, and may establish a link between them and dyslipidemia.

研究摘要 该项目的目的是阐明脂质生物合成的酶促途径 由人类梅博姆腺（HMG）产生，统称为meibum。 Meibum是一个 催泪膜（TF）至关重要的部分，除其他功能外，还保护眼表面免受 干燥。 meibum主要由非常且极为长链的独特混合物形成 复杂的脂质。我们以前的项目致力于生化和生物物理特征 Meibomian脂质（ML）。现在，我们建议研究导致形成的生物合成途径 Meibum在HMG中。提出了以下三项研究：（i）。尽管最近取得了进展 Meibum和TF的脂质组的表征，对生物合成途径没有明确的了解 Meibum的生物合成涉及的方式。 ML的不寻常且通常是独特的特质暗示着它们的原位 和/或从头起源，这又需要在HMG中实施适当的分子机制。 我们将研究表达水平和地图，主要脂质代谢酶和相关蛋白质的水平，这些蛋白质是 根据我们的初步研究，预计将存在于HMG中。这将通过使用 方法的组合，例如微阵列RNA分析，免疫组织化学/免疫细胞化学， 原位杂交，蛋白质组学和脂肪组学。 （ii）。最近的报告表明，总血脂水平高 包括胆固醇和TAG在内，与MG功能障碍的更高入射和/或严重程度相关。另外， 据报道，ω3脂肪酸（ω3FA）补充剂在人类和小鼠中报道了眼表面。 然而，这些作用的原因和分子机制尚不清楚。我们将确定余额 ML的原位/DE从头生物合成与血脂的水平和组成之间。我们将测试1） ω3FA的药物剂量的摄入量增强，（2）汀类药物的使用可以对 Meibum的生物合成，组成和特性。 （iii）。最近，我们报告了：（1）多达20％的正常 Meibum由非脂质成分组成； （2）正常和DE之间存在很大差异 meibum：后者富含非脂质材料，形成脂质层，在结构上不如结构稳定 那些由健康的meibum组成。根据这些发现，我们建议评估MEI的非脂质含量 BUM使用：（a）基于HPLC-MS，GC-MS和2D-DESI MS的代谢组方法； （b）Western印迹与 针对主要的Meibum和TF蛋白的特异性抗体； （c）针对蛋白质和的组织化学方法 多糖； （d）各种生物物理方法，例如热阶段的极化光学显微镜，微钙化 公制，Brewster角显微镜，研究Meibum和Tear膜的流变学和融化特征 脂质。我们认为，我们的工作将提供可能导致医学新概念的创新信息 治疗与MG相关的疾病和状况，并可能在它们与血脂异常之间建立联系。