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

Lead Research Organisation: University of Manchester
Department Name: Medical and Human Sciences

Abstract

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.

Technical Summary

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.

Publications

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Eatock Taylor R (2016) A coupled hydrodynamic-structural model of the M4 wave energy converter in Journal of Fluids and Structures

 
Description Clinical Fellowship
Amount £179,398 (GBP)
Organisation Fight for Sight 
Sector Charity/Non Profit
Country United Kingdom
Start 10/2010 
End 09/2013
 
Description Fight for Sight PhD Studentship
Amount £98,768 (GBP)
Funding ID 2033 
Organisation Fight for Sight 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2019 
End 09/2022
 
Description Fight for Sight/National Eye Research Centre Small Grant Award
Amount £14,547 (GBP)
Organisation Fight for Sight 
Sector Charity/Non Profit
Country United Kingdom
Start 11/2014 
End 08/2015
 
Description Fulbright Scholarship
Amount £75,000 (GBP)
Organisation Fight for Sight 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2013 
End 08/2014
 
Description MRC Career Development Award
Amount £664,961 (GBP)
Funding ID MR/K024418/1 
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 09/2013 
End 08/2016
 
Description Macular Society Research Grant
Amount £169,318 (GBP)
Organisation Macular Society 
Sector Charity/Non Profit
Country United Kingdom
Start 01/2018 
End 12/2020
 
Description Project Grant
Amount £110,274 (GBP)
Organisation Fight for Sight 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2016 
End 08/2018
 
Description Project grant
Amount £169,809 (GBP)
Organisation Fight for Sight 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2014 
End 08/2017
 
Description Project grant
Amount £480,344 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 01/2013 
End 12/2016
 
Description Project grant
Amount £217,343 (GBP)
Organisation Macular Society 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2014 
End 08/2017
 
Description Stepping Stones Research Fellowship
Amount £85,502 (GBP)
Organisation University of Manchester 
Sector Academic/University
Country United Kingdom
Start 01/2012 
End 12/2014
 
Description Univerity of Manchester and A*STAR PhD Studentship
Amount £127,983 (GBP)
Organisation University of Manchester 
Sector Academic/University
Country United Kingdom
Start 10/2012 
End 09/2016
 
Title Human eye collection 
Description We are building up a collection of human eye tissue (e.g. from donors of different ages) to support our work on Age-related Macular Degeneration. 
Type Of Material Biological samples 
Provided To Others? No  
Impact 6 primary papers (Clark et al., 2010 J. Biol. Chem.; Clark et al., 2011 IOVS; Keenan et al.., 2012 IOVS; Clark et al., 2013 J. Immunol.;Clark et al., 2014; Keenan et al., 2014 IOVS). Several papers are in preparation. Invited to speak at Complement UK Symposium (Newcastle, UK) 2013; Annual Meeting of Glycoren consortium 2013 (Leiden, The Netherlands); ARVO 2014 (Orlando, USA). 
 
Title Identification of novel disease mechanism for Age-related Macular Degeneration (AMD). 
Description We have identified a novel disease mechanism for Age-related Macular Degeneration (AMD). 
Type Of Material Model of mechanisms or symptoms - human 
Provided To Others? No  
Impact 3 primary papers (Clark et al., 2010 J. Biol. Chem; Clark et al. 2013 J. Immunol.; Keenan et al. 2014 IOVS), 4 review articles (Clark et al., 2010 Biochem. Soc. Trans; Day et al. Expert Reviews Ophthalmology; Clark et al., 2013 Frontiers in Immunology; Langford-Smith et al. 2014 J Innate Immunity) and 2 press releases. 
 
Title License agreement with Echelon Biosciences Inc. for method to make verscian G1 domain 
Description Developed protocol for expression, refolding and purification of human versican G1 domain. 
Type Of Material Technology assay or reagent 
Year Produced 2011 
Provided To Others? Yes  
Impact License agreement (Protocol for expression, refolding and purification of versican G1 domain) between MRCT and University of Manchester (February 2011). MRCT licensed this technology to Echelon Biosciences Inc. (July 2011). This reagent is available from Echelon Biosciences: https://echelon-inc.com/index.php?module=Products&func=view&category_id=10127 
 
Description Development of a human choroidal endothelial cell line 
Organisation Radboud University Nijmegen Medical Center
Country Netherlands 
Sector Academic/University 
PI Contribution We supplied reagents that allowed testing of new cell line providing verification that this has characteristics in common with primary choroidal endothelial cells.
Collaborator Contribution They established the new cell line and performed testing.
Impact A primary publication in IOVS in 2018.
Start Year 2014
 
Description Mapping of glycosaminoglycans in human retina, choroid and sclera. 
Organisation Radboud University Nijmegen
Department Department of Biochemistry
Country Netherlands 
Sector Academic/University 
PI Contribution We have used the reagents supplied to aid our research.
Collaborator Contribution The collaborators have provided reagents to aid our research.
Impact I primary paper in IOVS (2011). Abstract/poster at ARVO 2010 conference, Fort Lauderdale, Florida, USA (2010).
Start Year 2010
 
Description Role of complement factor H in AMD. 
Organisation University Hospital of Wales
Department Department of Medical Biochemistry and Immunology
Country United Kingdom 
Sector Hospitals 
PI Contribution We used reagents provided by these collaborators in our research.
Collaborator Contribution Provision of reagents to support our research.Provision of reagents to support our research.
Impact I primary paper in J. Biol. Chem. (2010). Abstract/poster at BSMB Spring Meeting: "Vascular Matrix in Health & Disease", Manchester, UK (2010). Abstract/poster at ARVO 2010 conference, Fort Lauderdale, Florida, USA (2010). Abstract/poster at BSMB Autumn Meeting "Inflammation meets matrix biology", UEA, Norwich (2010). Abstract/poster at 13th European Meeting on Complement in Human Disease, Leiden (2011). Abstract/poster at Get Connected 2, Wellcome Trust Centre for Cell-Matrix Research, University of Manchester, UK (2011). Abstract/poster at 7th International Conference on Proteoglycans, Sydney, Australia (2011). Abstract/poster/talk presented at ARVO 2012, Fort Lauderdale, USA. Primary papers published in J. Biol. Chem. (2010) and J. Immunol. (2013).
Start Year 2008
 
Description Role of complement factor H in AMD. 
Organisation University of Oxford
Department Department of Pharmacology
Country United Kingdom 
Sector Academic/University 
PI Contribution We used reagents provided by these collaborators in our research.
Collaborator Contribution Provision of reagents to support our research.Provision of reagents to support our research.
Impact I primary paper in J. Biol. Chem. (2010). Abstract/poster at BSMB Spring Meeting: "Vascular Matrix in Health & Disease", Manchester, UK (2010). Abstract/poster at ARVO 2010 conference, Fort Lauderdale, Florida, USA (2010). Abstract/poster at BSMB Autumn Meeting "Inflammation meets matrix biology", UEA, Norwich (2010). Abstract/poster at 13th European Meeting on Complement in Human Disease, Leiden (2011). Abstract/poster at Get Connected 2, Wellcome Trust Centre for Cell-Matrix Research, University of Manchester, UK (2011). Abstract/poster at 7th International Conference on Proteoglycans, Sydney, Australia (2011). Abstract/poster/talk presented at ARVO 2012, Fort Lauderdale, USA. Primary papers published in J. Biol. Chem. (2010) and J. Immunol. (2013).
Start Year 2008
 
Description Role of complement factor H in host recognition. 
Organisation Medical University of Innsbruck
Department Department of Hygiene, Microbiology and Social Medicine
Country Austria 
Sector Academic/University 
PI Contribution We have performed experiments using materials provided by the collaborators.
Collaborator Contribution They have provided research materials to aid the research.
Impact Abstract/poster at13th European Meeting on Complement in Human Disease, Leiden (2011). Abstract/poster at Get Connected 2, Wellcome Trust Centre for Cell-Matrix Research, University of Manchester, UK (2011). Abstract/poster at 7th International Conference on Proteoglycans, Sydney, Australia (2011). Abstract/poster at ARVO 2012, Fort Lauderdale, USA.
Start Year 2010
 
Description The heparin-binding properties of complment factor H. 
Organisation University of Edinburgh
Department School of Chemistry
Country United Kingdom 
Sector Academic/University 
PI Contribution We have used reagents provider by this collaboration to enable our research.
Collaborator Contribution Provision of reagents to enable our research.
Impact Abstract/poster at BSMB Autumn Meeting "Inflammation meets matrix biology", UEA, Norwich (2010). Abstract/poster at 13th European Meeting on Complement in Human Disease, Leiden (2011). Abstract/poster at Get Connected 2, Wellcome Trust Centre for Cell-Matrix Research, University of Manchester, UK (2011). Abstract/poster at 7th International Conference on Proteoglycans, Sydney, Australia (2011). Abstract/poster/talk presented at ARVO 2012, Fort Lauderdale, USA. Paper published in J. Immunol. (2013).
Start Year 2010
 
Description The role of FHL-1 in AMD 
Organisation University of Ulm
Country Germany 
Sector Academic/University 
PI Contribution Used reagent supplied by collaborator.
Collaborator Contribution Collaborator provided research reagent.
Impact Clark et al. (2014) J. Immunol.
Start Year 2012
 
Description "The Body Experience", Manchester Museum 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Type Of Presentation Workshop Facilitator
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Approximately 1900 members of the public visited the one-day event at the Manchester Museum, which highlighted research conducted on different parts of the body (including eyes) at the University of Manchester.

Public feedback revealed strong interest in this, and the possibility of future similar events.
Year(s) Of Engagement Activity 2010,2011
 
Description "Wellcome to the Matrix" open days. 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Type Of Presentation Workshop Facilitator
Geographic Reach Regional
Primary Audience Schools
Results and Impact 30 GCSE pupils (from local Manchester schools) attended each day over 5 or 3 days in 2010 and 2011, respectively. This included a tour of the research facilities and a wide range of participatory activities (e.g. model building) on the research carried out at the Wellcome Trust Centre for Cell-Matrix Research.

Positive feedback on our research environment and increased awareness of the role scientists and opportunities for careers in science.
Year(s) Of Engagement Activity 2010,2011,2013
 
Description A-level study day, Manchester Museum 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Type Of Presentation Workshop Facilitator
Geographic Reach Regional
Primary Audience Schools
Results and Impact 70 A-level students from Manchester schools attended lectures/workshop over two days at the Manchester Museum.

Post event surveys indicated that a large percentage (~80%) were more interested in the possibility of doing future scientific research than before attending. As a direct consequence of this event one student from Manchester Grammar School spent one week's work experience in the laboratory on this specific project.
Year(s) Of Engagement Activity 2011
 
Description Article by Paul Bishop for Macular Society magazine Digest entitled "Establishing a national eye tissue archive" 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Patients, carers and/or patient groups
Results and Impact Article on the importance of donor eye tissue for research into AMD and the establishment of Manchester Eye Tissue Repository
Year(s) Of Engagement Activity 2016
 
Description Elizabeth Thomas Seminar 2016. Presentation by Paul Bishop entitled "Eye Banking/Repository - How does that contribute to Research in Macular Diseases in the UK?" 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Raised awareness of Manchester Eye tissue Repository and subsequently requests have been made for donor eye tissue for research.
Year(s) Of Engagement Activity 2016,2017
 
Description Interview on Insight Radio (RNIB) 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Radio interview following award of Clinical Research Fellowship, which was based on the research carried out in this grant.

This interview was part of AMD awareness week with the aim of increasing public knowledge of this major form of blindness.
Year(s) Of Engagement Activity 2010
 
Description Invited speaker at Eyecare 2017, Glasgow. 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Simon J Clark gave a talk about new treatments being developed for Dry AMD to eye care health professionals and patients. Official feedback was collected and provided (please see URL provided below). Briefly, the talk was the third highest attended talk of the two day conference (80); where 67% of the audience ranked it as "Excellent", and a further 30% as "Good". Dr. Clark also took part in the most attended session of the conference - the 'Ask the Experts' Q & A panel.
Year(s) Of Engagement Activity 2017
URL https://gallery.mailchimp.com/75705ffde3df8421f476475ac/files/94884fc5-64b5-49cb-9a55-ad0630a6b7ca/E...
 
Description Invited speaker at Macular Society Top Doctor event 2015 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Invited speaker at the Macular Society's 'Top Doctor' event 2015. There was much discussion with the lay audience of mainly elderly individuals, some of who suffer from AMD, and interaction both during and after the talks.
Year(s) Of Engagement Activity 2015
 
Description Invited speaker, Womens Institute Science Club, Grazely, Berkshire. 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Simon J Clark was invited to talk about AMD, its affects on people and the economy and new treatments being developed that will make a difference in the future. He also raised money for the charity Fight for Sight.
Year(s) Of Engagement Activity 2016
 
Description Norfolk & Norwich University Hospital Regional Study Day, Bury St. Edmunds, UK. 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach Regional
Primary Audience Health professionals
Results and Impact ~100 Ophthalmology health professionals from Norfolk/Suffolk attended this Regional Study Day.

There was considerable interest in our on-going work on AMD.
Year(s) Of Engagement Activity 2010
 
Description Press release accompanying publication of research paper. 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Type Of Presentation Paper Presentation
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact Press release. Scientists discover new cause of blindness (21st September 2010): http://www.manchester.ac.uk/aboutus/news/display/?id=6123


Story covered by New Scientist (News In Brief: Leading Cause of Blindness Solved, 2nd October 2010):
Year(s) Of Engagement Activity 2010
 
Description Press release on publication in Journal of Immunology 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact The press release was picked up by >10 online news fora, including Science Daily and Science Codex.

None as yet.
Year(s) Of Engagement Activity 2013
URL http://www.manchester.ac.uk/aboutus/news/display/?id=9561
 
Description Royal Society Summer Science Exhibition 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? Yes
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact This event brought the importance of studying sugars to the public, schools, politicians and policymakers; ~10,000 visitors attended this event.

Unclear as yet.
Year(s) Of Engagement Activity 2013
URL http://sse.royalsociety.org/2013/exhibits/sweet-complexity/