Lipid mediators and their signaling in ocular surface inflammation and meibomian gland dysfunction

干眼症（DE）影响五分之一的退伍军人，并影响生活质量。这是一种复杂的疾病， 多种症状（如疼痛、视觉障碍）和体征（如泪液分泌减少、泪液分泌增加） 蒸发）。DE的关键病理生理成分是眼表炎症和睑板腺 功能障碍（MGD）。尽管它的频率和发病率，目前的治疗并不能缓解症状， 患者为了适当地管理DE，有必要了解机制和具体的 疾病亚型的贡献者。本提案的总体目标是通过研究提高这方面的知识， 生物活性鞘脂（SPL）在MGD和DE的其他方面的作用。我们的初步和公布的 研究表明，SPL成分变化（鞘磷脂（SM）/神经酰胺（Cer）比值升高）质量较差 睑板腺分泌物（睑脂）;促炎（ω-6）/促消退（ω-3）脂质的比例较高，以及 DE患者泪液中前列腺素E2水平较高。通过SM产生的生物活性Cer 鞘磷脂酶（SMase）及其衍生物，神经酰胺1-磷酸和鞘氨醇1-磷酸， 炎性脂质，并可诱导形成其他炎性脂质，如洋地黄素。我们检测到 人眼泪中的SMase活性。在这个建议中，我们将测试的假设，在SPL组成的变化， 睑脂导致DE的各种体征，包括眼表炎症、泪膜紊乱和角膜炎。 MGD。在目标1中，我们将分析睑脂和泪液SPL以及促炎和抗炎PUFA和类二十烷酸 通过LC-MS/MS从120个病例（睑脂质量差）和120个对照（睑脂质量正常）中选择， 我们前瞻性地从一个庞大的退伍军人群体中收集样本， (mean年龄62 ± 10岁; 467名男性，46名女性; 230名白色，272名黑人）。相关和回归分析将 以临床数据为因变量，以脂质介质为自变量。在目标2中， 将从对应于样品的240个Schirmer条带中测定SMAse（酸性和中性）的活性 目标1。然后，我们将建立模型，研究SMase活性、SPL和眼表之间的关系 炎症在目标3中，我们将在体外实验中确定SPL和炎症之间的分子联系。 通过使用人永生化睑板腺上皮细胞系（HMGEC）的模型来测试 生物活性SPL在正常和高渗条件下介导炎症、细胞活力、细胞死亡等。 影响：我们的结果将通过MGD状态识别SPL差异，阐明生物活性之间的关系， SPL、炎性脂质介质和SMase活性与眼表炎症和眼表炎症的临床特征 MGD和DE亚型。所产生的知识将通过确定相对的 不同脂质对DE不同表现的贡献，并为开发新的 治疗的分子靶点。

Dry eye (DE) affects 1 in 5 veterans and impacts quality of life. It is a complex disease that manifests with multiple symptoms (e.g. pain, visual disturbance) and signs (e.g. decreased tear production, increased evaporation). Key pathophysiological components of DE are ocular surface inflammation and meibomian gland dysfunction (MGD). Despite its frequency and morbidity, current therapies do not relieve symptoms in all patients. In order to appropriately manage DE, it is necessary to understand mechanisms and specific contributors to disease sub-types. The overall goal of this proposal is to improve such knowledge by studying the role of bioactive sphingolipids (SPL) in MGD and other aspects of DE. Our preliminary and published work suggest SPL composition changes (higher sphingomyelin (SM)/ ceramide (Cer) ratio) in poor quality meibomian gland secretion (meibum); a higher ratio of pro-inflammatory (ω-6) /pro-resolving (ω-3) lipids, and higher levels of prostaglandin E2 in tears from DE patients. Bioactive Cer generated from SM by sphingomyelinases (SMase) and its derivatives, ceramide 1-phosphate and sphingosine 1-phosphate are inflammatory lipids and can induce formation of other inflammatory lipids such as prostaglandins. We detected SMase activity in human tears. In this proposal, we will test the hypothesis that changes in SPL composition in meibum contribute to various signs of DE including ocular surface inflammation, tear film disturbances, and MGD. In Aim 1, we will profile meibum and tear SPL and pro- and anti-inflammatory PUFAs and eicosanoids by LC-MS/MS from 120 cases (poor meibum quality) and 120 controls (normal meibum quality), chosen from our prospectively collected samples from a large veteran population after a comprehensive eye evaluation (mean age 62±10; 467 males, 46 females; 230 white, 272 black). Correlation and regression analysis will be performed with clinical data as dependent variables and lipid mediators as independent variables. In Aim 2, we will determine acitivity of SMAse (acidic and neutral) from 240 Schirmer's strips corresponding to the samples in Aim 1. We will then build models looking at relationships between SMase activity, SPL, and ocular surface inflammation. In Aim 3, we will identify molecular connections between SPL and inflammation in an in vitro model by using human immortalized meibomian gland epithelial cell line (HMGEC) to test the effects of bioactive SPL on mediating inflammation, cell viability, cell death etc. in normal and hyperosmolar conditions. Impact: Our results will identify SPL differences by MGD status, elucidate relationships between bioactive SPL, inflammatory lipid mediators, and SMase activity with ocular surface inflammation and clinical features of MGD and DE sub-types. The knowledge generated will advance the field by determining the relative contributions of different lipids on different manifestations of DE and provide the foundation for developing new molecular targets for therapy.