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Sri Lankan scientist Dinusha Rajapakse in the forefront of a novel discovery

 

A tooth enamel protein has been discovered in eyes with dry age-related macular degeneration. This finding may lead to a novel therapeutic target for blinding disease. We spoke to the principal research scientist of the discovery, Dr. Dinusha Rajapakse (PhD), a visiting research fellow at the National Eye Institute (NEI), which is part of the National Institutes of Health (NIH), in the Section on Molecular Structure and Functional Genomics, regarding this discovery.

Age-related macular degeneration, a major cause of blindness in ageing populations worldwide, is a degenerative disease of the macula. The macula is the central area of the light-sensitive portion in our eyes (retina). Macular degeneration often leads to progressive central vision loss. According to the World Health Organisation (WHO) global eye disease survey report, 14 million people are blind or severely visually impaired due to age-related macular degeneration. This disease has an early and a late stage, with visual impairment occurring during the late stage of the disease. WHO has identified Sri Lanka in particular as an area in need of further investigation regarding this condition in keeping with the aims of the Vision 2020 project.

Age-related macular degeneration falls into two broad categories: Growth of new blood vessels breaking through into the retina, known as choroidal neovascularisation or “wet” age-related macular degeneration and progressive atrophy of the retinal pigment epithelium, and photoreceptor layers, known as geographic atrophy or “dry” age-related macular degeneration. Currently, there are effective therapies for choroidal neovascularisation, but none for geographic atrophy. One of the early signs of age-related macular degeneration is the observation of drusen. Drusen are yellow deposits under the retina and are made up of lipids, a fatty protein, and minerals. An increase in the number and size of these drusen deposits with age are believed to impair delivery of nutrients and removal of waste to the retina cells including light-sensing photoreceptors and eventually lead to age-related macular degeneration.

Types of macular degeneration (Source: iovs.arvojournals.org)

OCT image of eye with dry AMD, showing soft drusen beneath the retinal pigment epithelium (Photo Dr. Dinusha Rajapakse, NEI)

Macular degeneration (Source: iovs.arvojournals.org)

Recently, researchers found a calcium-containing mineral compound called hydroxyapatite in drusen deposits in dry age-related macular degeneration. Hydroxyapatite is a key component of tooth enamel and bone. Small balls of hydroxyapatite filled with cholesterol, called spherules, were found in drusen from people with age-related macular degeneration. When the build-up of hydroxyapatite increases, the drusen further develops and the retinal pigment epithelium and eventually the photoreceptors die, leading to blindness. The photoreceptors cannot grow back, so the blindness is permanent. 

In a new study, researchers have now identified a protein called amelotin, that normally deposits mineralised calcium in tooth enamel, may also be responsible for calcium deposits in the back of the eye in people with dry age-related macular degeneration. According to the study from researchers at the NEI, part of the NIH in the US, this protein may turn out to be a therapeutic target for the blinding disease. The findings were published in the journal Translational Research.

Using a cell culture model of retinal pigment epithelial cells, the researchers were able to show that amelotin expression gets turned on by a certain kind of stress and causes formation of a calcium deposit also seen in bones and teeth. When they looked in human donor eyes with dry age-related macular degeneration, they saw the expression of the amelotin protein. She explained, in the study, they discovered that if they starved retinal pigment epithelial cells grown in transwells, a type of cell culture system, for nine days, the cells began to deposit hydroxyapatite. 

They determined that the protein amelotin, encoded by the gene AMTN, is strongly upregulated after extended starvation and is responsible for the mineralisation of hydroxyapatite in their cell culture model. Blocking this pathway in their retinal pigment epithelial cell line also blocked the production of these drusen-like deposits. They then verified their findings from the cell culture model by examining human cadaver eyes with dry age-related macular degeneration, wet age-related macular degeneration, or without age-related macular degeneration. They found hydroxyapatite and amelotin only in the eyes with dry age-related macular degeneration, and not in the other eyes. While amelotin was found sometimes in areas of dry age-related macular degeneration without drusen, it was primarily present in soft drusen areas with large deposits of hydroxyapatite.

In a press release from the NIH, Dr. Dinusha Rajapakse said: “Prior to this study, nobody really knew how hydroxyapatite was accumulating in dry age-related macular degeneration drusen. Finding this tooth-specific protein in the eye, this protein that’s linked to hydroxyapatite deposition – that was really unexpected.”

Dr. Rajapakse and colleagues say it is unclear why retinal pigment epithelium cells in dry age-related macular degeneration begin depositing these hydroxyapatite spherules, but they think it may be a protective mechanism gone wrong. They speculate that these protein, lipid and mineral deposits may help block blood vessels from growing into the retina, which is a key feature of wet age-related macular degeneration. But when the hydroxyapatite deposits get too extensive, they may also block nutrient flow to the retina cells leading to cell death.

The study shows that amelotin looks like a key player for the formation of these very specific hydroxyapatite spherules. That is amelotin’s main function in tooth enamel formation, and now it is performing a similar function in the back of the eye. As amelotin is not normally present in the eye, the scientists believe they could come up with a drug that specifically blocks the function of amelotin in the eye, and that this might delay progression of the disease.

However, good animal models for testing dry age-related macular degeneration therapeutics are urgently needed. Therefore, based on the findings from this study, Dr. Rajapakse and the team at the NEI are creating a new mouse model for the disease. The future plan of the team is to use their cell culture model, which mimics features of dry age-related macular degeneration, to potentially perform high-throughput drug screening to find molecules that slow or prevent the development of soft drusen and eventually test these drugs on the new animal model they are creating.

These advances will undoubtedly have a large impact on the success of clinical trials in the hope of finding a cure for this blinding disease. While there are many challenges and unmet needs in understanding and treating this condition, this is an exciting time to be working in this area to bring together new technologies to harness therapeutic strategies and better understand retinal physiology.