Hyaluronan in the cornea: Regulation of limbal stem cell fate and lymphangiogenesis

项目总结 角膜缘干细胞(LSCs)是损伤后重建角膜上皮所必需的。伤势严重。 角膜导致LSCs或LSCN(LSCN)的实质性损害可能导致LSC缺乏 (LSCD)是一种严重的疾病，会导致角膜混浊、炎症、血管生成、严重 疼痛和结膜形成，最终可导致完全失明。人类的移植 LSCs体外扩增到受损的角膜上是一种常见的外科手术， 总体成功率为60%-70%。体外培养LSCs的一个主要障碍是 它们很容易分化为中央角膜上皮细胞，阻碍了它们用于治疗应用 并强调了在体外重建LSCN微环境以进行LSC扩增的必要性。 有趣的是，我们最近发表的研究表明LSCN是由透明质酸(HA)组成的。 支持干细胞表型所需的丰富基质。本提案的目标1将描述 LSCN中存在的复杂HA基质的精确组成。目标2将致力于复制 LSCN中存在的基质在体外扩增LSCs的培养条件下并建立 LSCN中的HA基质如何在体内调节LSC的表型。淋巴管是 仅存在于角膜缘(角膜富含透明质酸的区域)，因此角膜是无血管的。眼球 损伤和炎症，如LSCD，通常会导致不可逆转的角膜血管生成和 淋巴管生成，它降低了角膜的透明度，导致视力丧失，限制了手术的成功 LSC移植。淋巴管标志物LYVE-1(淋巴管内皮细胞受体1)是一种 透明粘附素(一种与透明质酸特异结合的蛋白质)，在淋巴系统中表达。Lyve-1 能够结合可溶性和固定化HA；然而，HA/Lyve-1相互作用是否发挥作用 在淋巴管生成中的作用仍然不清楚。有趣的是，我们的初步数据显示HA的角膜 基因敲除的小鼠在角膜缘只显示残留的淋巴管，这表明HA支架可能是 是角膜淋巴管生成所必需的。透明质酸在整个角膜中的上调先于病情加重 淋巴管扩张，角膜基质内注射HA导致淋巴管扩张 船只。因此，我们推测HA在调节角膜淋巴管生成中起重要作用。 目的3探讨角膜缘透明质酸调节角膜缘细胞生长的机制。 淋巴管的发育。

Project summary Limbal stem cells (LSCs) are required for reconstituting the corneal epithelium after injury. Injury to the cornea leading to substantial damage of LSCs or the LSC niche (LSCN) may lead to LSC deficiency (LSCD), a serious medical condition that causes corneal opacification, inflammation, vascularization, severe pain and conjunctivalization, and which can ultimately lead to complete loss of vision. The transplantation of LSCs expanded ex vivo onto the damaged cornea is a common surgical procedure that is carried out all around the world with an overall success rate of 60%- 70%. A major hurdle in culturing LSCs ex vivo is that they readily differentiate into central corneal epithelial cells, hampering their use for therapeutic applications and underscoring the need to regenerate the LSCN microenvironment in vitro for LSC expansion. Interestingly, our recently published work demonstrates that the LSCN is composed of an hyaluronan (HA) rich matrix which is required to support the stem cell phenotype. Aim 1 of this proposal will characterize the precise composition of the complex HA matrix present in the LSCN. Aim 2 will work towards reproducing the matrix present in the LSCN in culture conditions for ex vivo expansion of LSCs and also establishing how the HA matrix present in the LSCN regulates the LSC phenotype in vivo. Lymphatic vessels are present solely in the limbus (the HA rich region of the cornea) and the cornea is therefore avascular. Ocular injuries and inflammation, such as LSCD, often lead to irreversible corneal angiogenesis and lymphangiogenesis, which reduce corneal transparency, leading to vision loss and limiting the success of LSC transplantation. The lymphatic vessel marker Lyve-1 (LYmphatic Vessel Endothelial receptor 1) is a hyaladherin (a protein that specifically binds HA) and is expressed throughout the lymphatic system. Lyve-1 is capable of binding both soluble and immobilized HA; however, whether the HA/Lyve-1 interaction plays a role in lymphangiogenesis remains elusive. Interestingly, our preliminary data reveal that corneas of HA knock-out mice show only vestigial lymphatic vessels in the limbus, suggesting that an HA scaffold may be necessary for corneal lymphangiogenesis. HA up-regulation throughout the cornea precedes exacerbated extension of lymphatic vessels, and injecting HA into the corneal stroma leads to the extension of lymphatic vessels. Therefore, we postulate that HA plays an important role in regulating corneal lymphangiogenesis. Aim 3 proposes to identify the mechanism by which HA within the corneal limbus regulates the development of lymphatic vessels.