Abstract
This thesis focuses on the development of an advanced in vitro model of the outer blood-retina barrier (oBRB), a structure implicated in several ocular diseases. These diseases can lead to severe vision loss and blindness, and despite their prevalence, current treatment options are limited and invasive. The goal of this research was to create a cost-effective, reproducible, and physiologically relevant model of the oBRB to enhance our understanding of disease onset and progression, while also providing a potential platform for drug screening.A central focus of this work was the engineering of a biomimetic Bruch’s membrane (BrM), the extracellular matrix (ECM) of the oBRB, which plays a crucial role in supporting retinal pigment epithelial (RPE) cells and maintaining retinal homeostasis. Several approaches were explored to recreate the structural and biochemical complexity of the native BrM. Electrospun nanofibres were evaluated for their ability to form porous scaffolds that mimic the collagen-rich composition of the native BrM. Commercial membranes were also tested, alongside various surface coatings including collagen I and IV, to further enhance cell attachment and functionality.
To add biological complexity, decellularised pig skin and cellular ECM scaffolds were incorporated into the model, providing a more physiologically relevant environment due to their proteomic similarity to human BrM. These scaffolds were validated for their ability to support the attachment and growth of differentiated ARPE-19 cells, a widely used RPE cell line.
The optimised BrM mimetic was then integrated into a microfluidic organ-on-a-chip device to replicate the dynamic in vivo conditions of the oBRB. This device was carefully designed and fabricated to support the formation of a confluent RPE monolayer, while allowing for continuous nutrient flow and waste removal. Following the success of the single channel device, a double-channel chip was designed, featuring a decellularised scaffold that mimicked the BrM's architecture, further enhancing the physiological relevance of the model. By optimising flow rates and scaffold integration, the device successfully recreated the natural oBRB environment.
This research presents a novel oBRB-on-a-chip model, incorporating features not currently available in existing models, such as the use of decellularised scaffolds and differentiated ARPE-19 cells in a microfluidic system. The resulting platform provides a more accurate representation of the oBRB’s structure and function, offering new opportunities for retinal disease research, drug discovery, and therapeutic testing.
| Date of Award | 11 Dec 2024 |
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| Original language | English |
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| Supervisor | Amanda Mackenzie (Supervisor) & Paul De Bank (Supervisor) |