Lens Epithelial Cell Response to Biomaterial Interfaces

项目总结 白内障仍然是全球致盲的主要原因，每年有300多万例白内障摘除。 仅在美国。在白内障手术中，晶状体囊内的内容物通过一个 在晶状体前囊上开一个孔，并在囊内放置聚合物人工晶状体(IOL)。领先者 当晶状体上皮细胞残留时，会发生威胁视力的并发症，后囊混浊(PCO)。 晶状体上皮细胞从晶状体前囊到晶状体后囊膜或植入人工晶状体，并进行上皮到人工晶状体植入。 间充质转化(EMT)。而影响机械生物学和上皮细胞反应的几个因素 此前已有研究，对粘弹性的影响还没有清楚的认识 LEC行为的曲率。该项目的总体目标是使用聚合物和水凝胶衬底， 分别模拟植入物和晶状体微环境，更好地分析其对力学性能的影响 属性对LEC响应和EMT有影响。假设材料的物理和力学性能 晶状体组织摘除及人工晶状体植入术后，晶状体微环境发生改变，有利于晶状体上皮细胞的EMT。 在目标1中，将使用可调聚合物衬底和水凝胶来研究刚性和 粘弹性对LEC响应和EMT的影响。假设衬底比透镜胶囊更硬，并且 具有较低损耗正切的衬底将驱动LEC中的EMT。在目标2中，衬底曲率的影响将是 使用相同的聚合物和水凝胶基质进行了研究。主导假说是LEC迁徙 和EMT是由于人工晶状体植入后晶状体囊变平而导致的较大曲率半径所致。 曲率效应将使用具有不同曲率半径的聚合物微图案化进行评估。玻璃杯 不同大小的微珠将被嵌入水凝胶配方中，模拟晶状体中的变化 手术后的包膜形状。在这两个目标中，都将使用相关的体外和体外模型。LEC增殖， 将对EMT的迁移和标记进行评估。转化生长因子-β和雷帕霉素将作为阳性和阴性 分别为EMT的诱导剂。RT-PCR将量化基因表达，蛋白质表达的变化将是 用免疫印迹和免疫荧光方法进行评价。将进行评估的特定基因和蛋白质 包括SMAD信号蛋白、α-SMA、Slug、Snail、纤维连接蛋白、E-钙粘蛋白和YAP。这个项目的目标是 就是确定衬底的机械性能，即粘弹性和曲率对LEC的影响 EMT的行为和诱导。这将大大提高我们对LEC机械生物学和 这些因素在EMT中的作用，提出了预防病理性EMT的策略。结果将通向未来 预防EMT和植入后纤维化的材料设计研究，特别是预防PCO的材料设计。

PROJECT SUMMARY Cataract remains the leading cause of blindness worldwide with over 3 million extractions performed each year in the United States alone. During cataract surgery, the contents inside the lens capsule are removed through a hole in the anterior lens capsule and a polymeric intraocular lens (IOL) is placed in the capsule. The leading vision-threatening complication, posterior capsule opacification (PCO), occurs when residual lens epithelial cells (LEC) migrate from the anterior to the posterior lens capsule or onto the IOL and undergo epithelial-to- mesenchymal transition (EMT). While several factors impacting mechanobiology and epithelial cell response have been previously investigated, there is not a clear understanding of the impact of viscoelasticity and curvature on LEC behavior. The overall objective of this project is to use polymer and hydrogel substrates that mimic the implants and lens microenvironment, respectively, to better analyze the influence mechanical properties have on LEC response and EMT. It is hypothesized that the physical and mechanical properties of the microenvironment are altered after the removal of the lens tissue and IOL placement, facilitating EMT in LEC. In Aim 1, tunable polymer substrates and hydrogels will be used to investigate the impact of stiffness and viscoelasticity on LEC response and EMT. It is hypothesized that substrates stiffer than the lens capsule, and substrates with lower loss tangent will drive EMT in LEC. In Aim 2, the effect of substrate curvature will be investigated using the same polymer and hydrogel substrates. The governing hypothesis is that LEC migration and EMT are driven by larger radius of curvature caused by flattening of the lens capsule after IOL implantation. Curvature effects will be evaluated using polymers micropatterned with different radii of curvature. Glass microbeads of various sizes will be embedded in hydrogel formulations, mimicking the changes in the lens capsule shape following surgery. In both aims, relevant in vitro and ex vivo models will be used. LEC proliferation, migration, and markers for EMT will be assessed. TGF-β and rapamycin will be used as positive and negative inducers of EMT, respectively. RT-PCR will quantify gene expression, and changes in protein expression will be evaluated using Western blot and immunofluorescence. Specific genes and proteins that will be evaluated include SMAD signaling proteins, α-SMA, Slug, Snail, fibronectin, E-cadherin, and YAP. The goal of this project is to determine how substrate mechanical properties, namely viscoelasticity and curvature, contribute to LEC behavior and induction of EMT. This will significantly enhance our knowledge of LEC mechanobiology and the role of these factors in EMT, suggesting strategies to prevent pathological EMT. The results will lead to future research on design of materials to prevent EMT and fibrosis after implantation, particularly for preventing PCO.