Project 5 - Clearance of TIMP-3 by choroidal epithelial cells and its influence on macular degeneration

Applying for Summer 2025

School/Institute: No rwich Medical School, UEA

Introduction: Age-related macular degeneration (AMD) is a common condition that causes loss of central vision, making everyday tasks like reading and facial recognition increasingly difficult. One form of the disease, called “wet AMD”, is characterised by the formation of new blood vessels in the retina and can be treated by injection of anti-VEGF therapies into the eye. “Dry AMD” is characterised by progressive death of retinal cells, and is currently untreatable. A limitation to developing effective therapies is that the molecular mechanisms underlying AMD are not fully understood, with ageing, genetics, and environmental factors all playing a role in its development.

Supported by “The Macular Society” and “Fight for Sight”, we have been studying a related form of macular degeneration called Sorsby Fundus Dystrophy (SFD). This shares many clinical features with dry AMD, but has a simpler aetiology, as it is caused by mutation of a single gene called Tissue Inhibitor of Metalloproteinase 3 (TIMP3). These mutations cause TIMP-3 protein to accumulate in the retina, forming deposits that damage retinal cells and cause progressive sight loss. We hope that understanding the molecular causes of SFD will help us to develop new therapies to treat both SFD and AMD.

Research in our group recently identified a family of cell surface receptors that mediate clearance of TIMP-3 by healthy retinal pigment epithelium cells. These scavenger receptors bind to extracellular TIMP-3 and endocytose it into lysosomes for intracellular degradation. We found that SFD mutants of TIMP-3 were cleared more slowly, providing an important new insight into the molecular pathogenesis of macular degeneration.

Aims and Objectives: In this project, we will investigate clearance of TIMP-3 by choroidal epithelial cells, which form the blood vessels underlying the retina. This will give us a clearer picture of how different cells in the retina contribute to clearing wild-type and mutated TIMP-3 in healthy and diseases retinas. We will:

Grow a choroidal epithelium cell line in culture.
Use quantitative PCR to measure mRNA expression of various scavenger receptors.
Quantify degradation of purified wild-type and mutant TIMP-3 proteins by choroidal cells using SDS-PAGE and immunoblotting.
Test involvement of specific receptors using antagonists and siRNA (if time allows).

Skills Gained: This project will provide an opportunity to learn about cell culture, RNA isolation, quantitative PCR (RT-PCR), SDS-PAGE and immunoblotting. The student will be hosted by Linda Troeberg at the modern Bob Champion Research and Education Building, attending weekly group meetings and gaining exposure to a vibrant biomedical research environment. The host group has considerable experience in hosting successful 8-week projects for those new to scientific research, with several of these leading to authorship on subsequent publications.

Reference: Betts JHJ and Troeberg L (2024). Mechanisms of TIMP-3 accumulation and pathogenesis in Sorsby fundus dystrophy. Mol Vis 30: 74-91. http://www.molvis.org/molvis/v30/74/