Molecular Analysis of the Human Aqueous Outflow Pathway

我们研究计划的目标是确定与房水流出阻力有关的蛋白质，以及 确定青光眼中的蛋白质是否发生了变化。水的流出阻力可能会受到影响 由房水中的蛋白质和小梁细胞产生的蛋白质和细胞外基质组成。 由于不存在原发性开角型青光眼(POAG)的动物模型，因此对人类房水的研究 在过去的30年里，外流通路使用细胞和组织培养技术来分析小梁 网络。这些技术使用的介质补充剂(血清等)与网络的正常情况不同 房水环境。我们的研究发现，目前的培养条件改变了小梁网络 蛋白质的表达(在细胞和组织中)，当与在房水中培养的网络相比较时。 在房水中培养的小梁网维持着一种更接近于 新鲜的，非培养的网状结构。我们假设房水中发现的蛋白质需要 维持小梁细胞和网状结构接近其正常生理状态。细胞和网络的研究 不在房水中培养导致的发现可能与理解 活体情况。我们进一步假设，水中蛋白质的类型和比例的变化 (生长因子、细胞因子等)可改变小梁内环境平衡，导致房水异常流出 青光眼的抵抗和发展。目前的提案将调查我们在 以下方式： 描述房水对人眼小梁细胞和分子活动的影响 建立细胞网络培养模型并确定哪些水蛋白对小梁细胞有重要作用 动态平衡。 ？建立正常和开角型青光眼房水中蛋白质的全面图谱 病人。我们还将确定在网络中诱导的基因和蛋白质表达的变化 正常房水和POAG房水。 ？开发一种可用于小梁细胞培养的模拟房水的培养液。

The goal of our research program is to identify the proteins involved in aqueous outflow resistance, and characterize whether proteins are altered in the glaucomatouseye. Aqueous outflow resistance can be affected by the proteins in aqueous humor and the proteins and extracellular matrix produced by trabecular cells. Because no animal model of primary open angle glaucoma (POAG) exists, studies of the human aqueous outflow pathway over the past 30 years have used cell and tissue culture techniques to analyze the trabecular meshwork. These techniques use media supplements (serum, etc) that differ from the meshwork's normal aqueous humor environment.Our studies find that current culture conditions alter trabecular meshwork protein expression (in both cell and tissue) when compared with meshworks cultured in aqueous humor. Trabecular meshworks cultured in aqueous humor maintain a protein profile that more closely resembles fresh, non-cultured meshworks. Wehypothesize that proteinsfound in aqueous humor are required to maintain trabecular cells and meshworks near their normal physiologic state. Studies of cells and meshworks not cultured in aqueous humor result in findings that may not be physiologicallyrelevant to understanding the in vivo situation. We further hypothesize that changes in the types and proportions of aqueous proteins (growth factors, cytokines, etc.) can alter trabecular homeostasis resulting in abnormal aqueous outflow resistance and development of glaucoma. The current proposal will investigate our hypothesis in the following manner: ¿ Characterize the effect of aqueous humor on molecular and cellular activities in human trabecular meshwork culture models and determine which aqueous proteins are important for trabecular cell homeostasis. ¿ Create a comprehensive profile of proteins found in aqueous humor obtained from normal and POAG patients. We will also determine gene and protein expression changes in the meshwork induced by normal and POAG aqueous humor. ¿ Develop a medium that can be used to mimic aqueous humor for trabecular meshwork cell culture.