Eye spy: Proteins involved in diabetic macular edema

Macular moments

One of the leading causes of blindness in many countries around the world derives from diabetes and is known as diabetic macular edema (DME). This disease is a severe threat. People with type 2 diabetes have a 10% risk of developing it with the threat of vision loss to come. And the number of cases is high. In the USA alone, about one million people are existing with DME with another 300,000 cases diagnosed annually.

The disease is a direct consequence of high blood sugar levels, which cause the blood vessels in the retina to leak into the macula as a result of a breakdown in the blood-retinal barrier. The macula is the part of the eye that gives detailed central vision. The blood leakage causes the macula to swell, adopting a convex shape and distorting or blocking vision. It can bring about severe loss of sight and, eventually blindness.

A group of scientists in Spain has been working together to try and find new ways to treat DME. As senior reporter Rafael Simó from the Vall d' Hebron Research Institute in Barcelona pointed out in their new investigation, current treatments are not entirely satisfactory. Laser treatment of the eye or existing drug treatments can produce strong side effects like impairment of colour vision or glaucoma.

So, this research team decided to examine the proteins involved in DME to try and identify new potential drug targets. They reasoned that a close examination of the vitreous fluids from patients with and without DME would pinpoint those proteins affected by DME and shed light on the way the condition develops.

Vitreous humour tests

They accessed their human vitreous humour bank for appropriate samples, selecting four type 2 diabetic patients who had DME but no proliferative diabetic retinopathy (PDR) and four with PDR and no DME. PDR is another kind of retinal damage that can occur in diabetic patients. As controls, the samples from patients without diabetes but who had idiopathic macular hole were selected.

Albumin and immunoglobulin G are the two most abundant proteins in vitreous humour and their presence can mask other, less abundant proteins. To prevent their influence, they were removed by ultrafiltration. Together, they make up about 80% of the total proteins in each of the three sample groups, so taking them out would not significantly affect the abundance ratios of the other proteins.

The protein pellets were purified before being subjected to 2D difference gel electrophoresis using fluorescent labels. After measuring the abundances of the proteins on the gels, those that had markedly different abundances in the DME samples were identified by mass spectrometry.

This comparison reduced the number of protein spots from around 1300 to just four, none of them having been associated before with DME. The amended abundances of three of this group were confirmed by ELISA on different sets of samples from the vitreous bank.

Therapeutic targets

Only one of the four proteins was more abundant in the DME samples and that was hemopexin. It is thought to have several functions in the body, including acting as a protectant against heme-based oxidative stress and nitric oxide toxicity. It has also been associated with the response to proinflammatory cytokines, so could be increased as a direct result of the blood-retinal barrier being damaged.

The remaining three proteins were all reduced in abundance in the DME vitreous fluid. Clusterin, also known as apolipoprotein J, and transthyretin are also known to have different functions whereas crystallin S is part of the well-known group of crystallins that have important functions in the lens.

Despite the discovery of the altered proteins, their role in DME is unclear. However, they all have potential as therapeutic targets for this disease and might prove to be more effective routes to treating DME than current therapies.

A number of proteins associated with PDR were also identified but they have not been confirmed by ELISA and their roles in the condition remain to be clarified.

Although these results are promising, there are some limitations to the investigation, principally the low amount of samples that were tested. A total of four from patients suffering with DME is far too low a number on which to base any new drug treatments. However, it is a promising start, especially if the proteins can be confirmed on a larger set of samples.