Sphingolipid signaling in corneal wound healing and fibrosis

 描述(由申请人提供)：人类角膜纤维化和疤痕形成是角膜损伤后的常见结果。尽管在这一领域取得了重大进展，但目前的治疗方法有限，视力恢复很差。其中一个问题是角膜创面愈合是一个复杂的过程，涉及角膜细胞、细胞外基质成分和生长因子。预防角膜纤维化的机制尚不清楚。需要进一步的了解，以及新的治疗方法来治疗角膜疤痕。当我们考虑到全世界总共有2.5亿人视力受损，大约600万人失明，主要是由于角膜纤维化和疤痕形成时，问题的深度是显而易见的。角膜瘢痕的主要特征是存在肌成纤维细胞，通常表现为α-SMA(平滑肌肌动蛋白)的表达，以及过度和不适当的细胞外基质成分，如III型胶原。预防疤痕的形成将是理想的；然而，调查人类角膜非纤维性愈合发展的研究极其有限。这主要有两个原因：1)人体组织的不可获得性使研究变得困难，2)角膜瘢痕形成的机制还没有完全了解。我们建议研究一种新的在角膜纤维化中的作用，称为鞘磷脂。近年来，鞘磷脂被发现与多种组织中的纤维化有关，并被发现与转化生长因子途径密切相关，转化生长因子途径是一种由转化生长因子亚型与细胞表面受体结合后引发的一系列事件，已被证明参与了角膜纤维化的发展。我们将利用我们之前的经验，利用我们的人体3D培养模型来研究鞘磷脂在人类角膜纤维化中的作用。初步数据显示，鞘脂和转化生长因子途径之间存在直接联系。这将是了解鞘脂作用于角膜纤维化的机制的独特工具。我们也有相应的小鼠模型，以便将我们的体外研究结果与体内研究结果相关联。我们提出了两个特定的目标来研究以下问题：第一，鞘磷脂，特别是鞘氨醇-1-磷酸(S1P)及其受体是如何与转化生长因子相关的，以及它们如何在体外改变纤维化级联反应？第二，我们能否使用S1P生物合成的基因敲除小鼠品系来确定鞘磷脂在体内的作用？与公共健康相关--角膜疤痕可导致完全或部分失明。研究一种新的调节非纤维化角膜发育的机制和途径将为解决这一威胁视力的过程提供重要的解决方案。最终，这项研究将导致可以治疗人类角膜疤痕的治疗剂的开发。

 DESCRIPTION (provided by applicant): Human corneal fibrosis and scarring is a usual outcome following corneal injury. Despite significant advancements in the field, current treatments are limited, the visual recovery is poor. One of the problems is that corneal wound healing is a complex process, involving corneal cells, extracellular matrix (ECM) components and growth factors. The mechanism by which corneal fibrosis can be prevented is still unknown. Further understanding is required, along with new treatments to treat corneal scarring. The depth of the problem is clear when we consider that a total of 250 million people worldwide have compromised vision and around 6 million have been blinded, majorly due to corneal fibrosis and scarring. The main characteristic of a corneal scar is the presence of myofibroblasts, often indicated by the expression of α-SMA (smooth muscle actin), and the excessive and improper deposition of ECM components such as type III collagen. Preventing scar formation would be ideal; however, studies investigating the development of non-fibrotic healing in human corneas are extremely limited. There are two main reasons for that: 1) Unavailability of human tissue makes investigations difficult, and 2) Mechanistic pathways of corneal scarring are not fully understood. We propose to investigate a novel player in corneal fibrosis known as sphingolipids. Sphingolipids have been recently linked to fibrosis in a variety of tissues and they are found to be in close connection to the TGF-ß (transforming growth factor) pathway, a cascade of events initiated by TGF-ß isoforms after binding to its receptors on cell surface, which has been shown to be involved in corneal fibrotic development. We will utilize our previous experience using our human 3D culture model in order to investigate the effect of sphingolipids in human corneal fibrosis. Preliminary data shows a direct link between sphingolipids and TGF-ß pathways. This would be a unique tool to understand the mechanism by which sphingolipids act on corneal fibrosis. We also have the corresponding mouse models in order to correlate our in vitro findings to in vivo. We propose two specific aims to examine the following questions: First, how are sphingolipids, specifically sphingosine-1-phosphate (S1P), and their receptors related to TGF-ß and how they alter the fibrotic cascade in vitro? Second, can we use genetic knockout mouse lines of S1P biosynthesis to determine the effect of sphingolipids in vivo? Relevance to Public Health - Corneal scarring can lead to complete or partial loss of vision. Investigation of a novel mechanism and pathway that can regulate non-fibrotic corneal development will provide important solutions to this sight threatening process. Ultimately this study will lead to the development of therapeutic agents that can treat corneal scarring in humans.