Host tissue recognition by complement factor H in the human eye: mapping changes with age and in AMD

Over half of blindness in the UK is caused by age-related macular degeneration (AMD), with about 50 million people affected worldwide. AMD is a disease that damages the central part of the retina (called the macula) and leads to central vision loss. There is mounting evidence that the risk of developing this disease is strongly influenced by our genes. Recent studies revealed that a common variant in one particular gene (known as Complement Factor H or CFH for short) strongly increases the risk of developing AMD. This variant (called the Y402H polymorphism) is present in about 35% of people of European decent and it results in a small, but significant, change in the CFH protein that alters its functional properties. We have recently made an important discovery that the ?Y? and ?H? forms of CFH (normal and disease-causing, respectively) have different abilities to recognise carbohydrate molecules in the eye. This may influence the localisation of the CFH protein, which is likely to be important to the cause and development of AMD since CFH is part of our immune system that distinguishes healthy from diseased tissues. If the CFH protein is not present in the correct location then a disruption to the function of the immune system would lead to tissue damage.
We have proposed that AMD may result from age-related changes in the carbohydrate molecules within the eye, which we believe would influence CFH location and function. One aim of this research project is to test this idea by analysing eye tissues from donors of different ages (supplied by the Manchester Eye Bank) in a wide range of experiments. In addition to normal eye tissues we will also examine AMD-affected eyes. These studies are possible due to the molecular tools we have developed that allow us to look (using microscopy) at CFH-binding sites in human tissues. Our own expertise in Manchester (for example in AMD, the CFH protein and carbohydrate chemistry) will be complemented by a network of scientific collaborators (in the UK and USA) providing access to specialised biochemicals and cutting-edge technologies.
Overall these studies are likely to lead to a greater understanding of the causes of AMD and may allow the development of new therapeutic strategies for treating this disease.

Age-related macular degeneration (AMD) is the leading cause of blindness in the western world, where the Y402H polymorphism of complement factor H (CFH) is a major risk factor for disease onset/progression. CFH is a regulator of the complement system and there is strong evidence that the dysregulation of complement is an early step in AMD. Previously we reported that the 402H and 402Y variants of CFH interact differentially with sulphated glycosaminoglycans (GAGs). We hypothesised that this functional difference may modify the tissue-binding properties of CFH and could contribute to the pathology of AMD. Recently we have found that these variants (studied in isolation in a recombinant construct and in the context of full-length CFH) recognise different sites within macula tissue of normal human eyes. There are substantially fewer binding sites for the AMD-associated 402H than the 402Y variant within the Bruch?s membrane, i.e. the site where particulate matter (drusen), associated with central vision loss, accumulates. Importantly, our data indicate that binding sites in human macula for CFH are comprised mainly of the GAGs heparan sulphate (HS) and dermatan sulphate (DS), i.e. structurally diverse matrix polysaccharides.
We, therefore, hypothesise that age-related changes in GAG structure within the macula combined with the different GAG-binding properties of the 402H/402Y proteins (i.e. the genetic predisposition) make a major contribution to the pathology of AMD. The aims of this study are to: 1) test this hypothesis by determining how the CFH-binding sites in macula tissue change with age and disease; 2) undertake a detailed analysis of the different GAG structures that mediate binding to the 402H/402Y variants, and 3) characterise the GAG-binding properties of different regions of CFH. Specifically, we will characterise human macula (by immunofluorescence) for 402H- and 402Y-binding sites (and endogenous CFH) from a wide age range of genotyped donors compared to AMD tissues (i.e. containing drusen); the relative contributions of different regions of CFH will be determined. Furthermore, we will identify the HS/DS structures present in macula (e.g. by mass spectrometry), characterise the CFH-GAG interactions using a range of biophysical techniques, and determine the role of GAGs in modulating CFH activity.
These studies will help determine the molecular basis of host tissue recognition in the macula during normal aging and in AMD. We anticipate that this will lead to a greater understanding of the pathology/progression of AMD and may allow the development of new therapeutic strategies for treating this disease.