Lens Epithelial Cell Response to Biomaterial Interfaces

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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.