Protocol for Studying TM Cell Function in a 3-D Matrix

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This chapter describes the protocols to bioengineer a 3D human trabecular meshwork tissue model using Glauconix Biosciences' patented 3D-HTM™ system. These protocols combine species-relevant tissue with nanoengineering techniques in order to produce a 3D matrix to mimic the conventional outflow pathway, which is responsible for the majority of the aqueous humor drainage and intraocular pressure regulation in the eye. Currently, there is a lack of translation between glaucoma in the human eye and animal models, which fails to provide a sufficient platform for compound screening and intraocular pressure (IOP) assessment. To address this unmet need, a 3D-HTM™, which can be co-cultured with human Schlemm's canal cells and induced to a glaucomatous-like state, was developed to provide a more applicable model for the evaluation of IOP-lowering therapeutics. The included protocols address the engineering of this 3D tissue model, as well as the examination of key biological markers of diseased tissue using traditional biological techniques with a few modifications. Additionally, a methodology for indirectly measuring IOP within the bioengineered 3D tissue is described. Overall, these protocols provide an in vitro alternative for studying glaucoma at the anterior segment, which can propel the development of glaucoma therapeutics.