Development of an in-vitro animal model to evaluate novel pharmaceutical approaches to DED management

Abstract

In the ophthalmic field, multifactorial pathologies such as Dry Eye Disease (DED) andcataract are largely studied in living animal models that can fail to precisely mirror thecomplexity of these conditions in humans. Recent advances in biomedical technologieshave improved the reliability of in-vitro/ex-vivo animal alternatives, and to date,the corneal and crystalline lens tissue have been independently maintained physiologicallystable for 10 days. This thesis details the development of a novel and complete ex-vivo anterior eye model,which is capable of sustaining both the cornea and crystalline lens in a physiologicallystable state in loco for 7 days. The platform is based on porcine eyes, which representa high quality and reliable human tissue source substitute, and being slaughterhousewaste, also perfectly align the project with the 3Rs principle of replacing, refining andreducing living animal experimentation. The model is modular and scalable, allowingfor the maximisation of experimental reliability, and the minimisation of wasteand energy use. In addition, the whole system is designed to be fitted in a laminarflow cabinet, avoiding external biological contamination, and is easily transportablebetween tissue engineering laboratories, maximising accessibility. The model was validated estimating cell viability over time. Stromal fibroblasts werefound to be viable up to the seventh day of culture, and corneal and crystalline lenstissue maintained their transparency over the culturing period. Dry Eye Disease wassuccessfully induced in the model by irrigating the ocular surface every 40s, and validatedusing impression cytology technique. Moreover, due to the unique presence inloco of the crystalline lens, the model was also used as a platform to perfect cataractsurgery and successfully implant intraocular lenses (IOLs). The novel and complete ex-vivo anterior eye model developed in this thesis providesfurther insights into pre-clinical anterior segment investigations in ophthalmology,taking a step forward toward bridging the existing gap between in-vitro and in-vivobiomedical technologies.