The Association of Physicians of Great Britain and Ireland 2008
One Hundred and Second Annual General MeetingThe One Hundred and Second Annual General Meeting was held in the Rattray Lecture Theatre at the University of Leicester, on 10th and 11th April, 2008. The attendance book was signed by ordinary members and senior members.
The President, Professor Edwin Gale, took the chair.
(1) Minutes
The minutes of the last Annual General Meeting, having been published in the QJM, were taken as read, confirmed and signed.
(2) Matters arising
Professor Gale advised that the Future editorship of the QJM was an outstanding matter from 2007. He welcomed Professor Michael Bannon, the new QJM Editor to the meeting.
(3) Election of Officers and Committee
Prof Gale advised it was his pleasure to step down as President and invited Prof. H. Thurston to lead the meeting.
The following Officers and Executive Officers were elected:
Executive Committee
President:Professor H. Thurston
President-elect:Professor M. Sheppard
Honorary Treasurer:Professor J. Iredale
Honorary Secretary:Professor Dermot Kelleher
Members for England and Wales
Professor A. FinlayProfessor P. Woo
Professor K. MooreProfessor I. Hall
Professor W. RosenbergProfessor F. Karet
Members for Scotland
Dr A. Jardine
Professor D. Newby
Dr J. Petrie
Members for Ireland
Professor S. Donnelly
Professor J. Nolan
Professor P. Bell
(4) Election of Honorary members
The following Honorary members were elected:
Honorary Members
Professor Peter BarnesProfessor Sir Ravinder Maini
Professor Sir John BellProfessor Sir Alex Markham
Professor Edwin GaleProfessor Sir John Savill
Professor Andrew HattersleyProfessor Tony Weetman
(5) Election of Senior Members
The following senior members were elected:
Professor A.B. AtkinsonProfessor J.H. McKillop
Dr N.D. BaxDr P.R. Mills
Professor D.J. BetteridgeProfessor K.G. Nicholson
Professor P. CiclitiraProfessor C.D. Pusey
Professor S.M. CobbeDr P.A. Routledge
Dr C.S. CockramProfessor P. Rubin
Professor D.A. CompstonProfessor G.I. Sandle
Professor T.M. CoxProfessor M.F. Scanlon
Dr C.C. DohertyProfessor D.J. Shale
Dr G.M. DusheikoProfessor D.J. Sheridan
Dr M.E. EdmondsProfessor D.G. Thompson
Professor M.J. FarthingProfessor N.C. Thomson
Professor A.P. GreeningProfessor L.T. Weaver
Professor A. GrossmanDr J. Webster
Professor J.M. HopkinProfessor C.M. Wiles
Professor D.G. JohnstonProfessor J.D. Williams
Professor R.T. JungProfessor K.L. Woods
Dr L. KennedyProfessor P. Woo
Professor S. LightmanProfessor A.Y. Finlay
Dr J.T. MacfarlaneDr Ann Dornhorst
Professor A.M. McGregorProfessor D.G. Scott
(6) Ordinary Members
There were 20 nominated from 27 proposals received. The following ordinary members were elected:
Arlt, Wiebke, BA, MBChB, CCST, DSc, FRCP. Professor of Medicine, MRC Senior Clinical Fellow, University of Birmingham, Division of Medical Sciences, Institute of Miomedican Sciences Rm225, Birmingham, B15 2TT, UK.
Bannon, Michael, MB, MSc (MIS), DCH, FRCPI, FRCPCH. Postgraduate Dean/Director, Department of Postgraduate Medical & Dental Education, University of Oxford, The Triangle, Roosevelt Drive, Headington, Oxford, OX3 7XP, UK.
Bassett, John Howard Duncan, BA, BM BCh, MRCP, PhD, FRCP. Clinical Senior Lecturer and MRC Clinician Scientist, Imperial College Honorary Consultant Physician, Hammersmith Hospital, 5th Floor MRC Clinical Sciences Centre, Molecular Edocinology, Hammersmith Hospital, Du Cane Road, London, W12 0NN, UK.
Ceriello, Antonio HD. Professor of Diabetes and Endocrinology, Clinical Sciences Building, Clinical Sciences Research Institute, Clifford Bridge Road, Coventry, CV2 2DX, UK.
Cruickshank, John Kennedy, BSc, MB ChB, MD, MRCP, FRCP. Professor of Cardiovascular Medicine and Clinical Epidemiology, Hon Consultant Physician, Core Technology Facility (3rd Floor), Division of Cardiovascular & Endocrine Sciences, University of Manchester, 46 Grafton Street, Manchester, M13 9NT, UK.
Evans, Thomas Ronald, J MB BS, MRCP, MD, FRCP, FRCP. Professor of Translational Cancer Research, Centre for Oncology and Applied Pharmacology, Cancer Research UK Beatson Laboratories, University of Glasgow, Garscube Estate, Switchback Road, Glasgow, G61 1BD, UK.
Farooqi, Ismaa Sadaf, MB ChB, MRCP, PhD. Wellcome Trust Senior Clinical fellow, University of Cambridge and Honorary Consultant Physician in Diabetes and Endocrinology, Cambridge University NHS Foundation Trust, Level 4, Institute of Metabolic Science, University of Cambridge, Metabolic Research Studies, Box 289, Addenbrooke's; Hospital, Cambridge, CB2 0QQ, UK.
Gribben, John G, BSc, MBChB, MRCP, MRCPath, MD, FRCPath, FRCP, DSc. Professor of Experimental Cancer Medicine, Queen Mary, University of London, Charterhouse Square, Barts and The London School of Medicine, London, EC1M 6BQ, UK.
Irvine, Alan, MB BCh BAO, MRCP, MD, CCST. Consultant Paediatric Dermatologist, Our Lady's; Hospital for Sick Children, Dublin, 12, Ireland.
Kalra, Lalit, MBBS, MD, MRCP, PhD, MD, FRCP. Professor of Stroke Medicine, Kings College School of Medicine, Department of Diabetes, Endocinology & Internal Medicine, Bessemer Road, London, SE5 9PJ, UK.
Keane, Joseph, MD. Consultant Physician in Respiratory Medicine and Director of Research, School of Medicine, Trinity College, Trinity Centre for Health Sciences, St James's; Hospital, Dublin 8, Ireland.
Kearney, Mark, MBChB, MRCP, DM. Professor of Cardiology, University of Leeds, The LIGHT Laboratories, Clarendon Way, Leeds, LS2 9JT, UK.
Kee, Frank, BSc, MB BcH BAO, QUB MRCP, MSc, MFPHM, MD, FFPHM, FRCP, BSc. Director, Centre for Clinical and Population Sciences, The Queen's; University of Belfast, Mulhouse Building, Department of Epidemiology & Public Health, Royal Victoria Hospital, Grosvenor Road, Belfast, BT12 6BJ, UK.
Lachmann, Helen J, BA, Bchir, MA MB, MRCP, MD, CCST 9 in Renal Medicine, Senior Lecturer and Honorary Consultant in Amyloidosis and Renal Medicine, Royal Free and University College Medical School, National Amyloidosis Centre, Department of Medicine, Hampstead Campus, Rowland Hill Street, London, NW3 2PF, UK.
Marshall, Sara, MB BCh, BAO, LRCPI and LRCSI, MRCP, PhD, MRCPath, CCST, FRCPath. Professor of Clinical Immunology and Honorary Consultant Ninewells Hospital & Medical School, Dundee, DD1 9SY, UK.
Sayer, John Andrew, MB ChB, MRCP, PhD. Senior Lecturer in Nephrology, Newcastle University, Institute of Human Genetics, Central Parkway, Newcastle upon Tyne, NE1 3BZ, UK.
Sheerin, Neil, BSc, MBBS, MRCP, PhD, Professor of Nephrology, Newcastle University, School of Clinical Medical Sciences, Framlington Place, Newcastle upon Tyne, NE2 4HH, UK.
Stebbing, Justin, BA, BM BCh, MA, MRCP, PhD, MRC. Senior Lecturer and Honorary Consultant in Medical Oncology, Hammersmith Hospital, Department of Clinical Oncology, Du Cane Road, London, W12 0HS, UK
Stott, David J, MB ChB, MRCP, MD, FRCP. David Cargill Professor of Geriatric Medicine (Honorary Consultant in General and Geriatric Medicine), 3rd Floor QEB, Cardiovascular and Medical Division, Glasgow Royal Infirmary, Glasgow, G31 2ER, UK.
Walters, Matthew, MBChB, MRCP, MSc, MD, FRCP. Senior Lecturer in Medicine and Honorary Consultant Physician, Western Infirmary, Division of Cardiovascular & Medical Sciences, Glasgow, G11 6NT, UK.
The new members were welcomed to the Association by the President.
(7) Treasurer's Business
The President then invited the Honorary Treasurer, Professor J. Iredale, to report to the membership. Professor Iredale reported that the accounts of the Association were in good health. He presented income and expenditure for the months to the end of December 2007. He highlighted the key points of the accounts.
Income from the Oxford University Press has increased, further to the renegotiation of the contract for the QJM. Returns on investment income have increased and have been unaffected by recent stock market volatility.
Expenditure on charitable spending has increased due to the high quality of applications.
Professor Iredale highlighted the plan to remove the £30 000 ceiling for the Links with Developing Countries charitable grants in the years when there are sufficient funds available, and advised there are no plans to alter the membership subscription fee.
He invited people to speak to him during the meeting if they had any questions about the accounts.
(8) Links with Developing Countries/Tropical Health & Education (THET)
Prof Iredale advised the meeting that the three applications received were all funded in full. These totalled £35 000.
(9) Report of the Editor of the Quarterly Journal of Medicine
Professor M. Bannon addressed the meeting. He thanked the Association for inviting him to edit the QJM. He advised that he would now take a strategic approach to the journal. He fed back that the journal is vibrant, relevant, in good shape financially and has a good impact factor. But he noted that the publication exists in a competitive and changing world and it is important the journal evolves.
He advised that he is restructuring the Editorial Board. Board members will help shape content and direction of the QJM, through representing specialist interests and regions. They will also review papers, recommend hot topics and write commentaries and editorials. An advertisement for new board members was placed in the current edition of the QJM with a deadline of the end of April.
Professor Bannon advised he is considering the benefits of having an Associate Editor whose role would be to support the Editor.
(10) Future programme
Professor M. Sheppard, President Elect, welcomed all members to attend the 2009 Meeting in Birmingham, 2–3rd April. He advised he is delighted to host the event at Birmingham University, where there is a new lecture theatre.
Professor Thurston advised that the 2010 meeting will be held in Dundee and further details will be announced in 2009.
(11) Any other business
Professor Kelleher, Honorary Secretary, thanked Concorde Services Limited, Carol Patrick Mary Kavanagh and Carolynn Walthew for their help in organizing the meeting.
Opening Reception and Annual Dinner
The welcome reception on Thursday evening was held in the New Walk Museum and Art Gallery. The Annual Dinner was held in the Garendon Restaurant, University of Leicester. Members enjoyed an excellent dinner and the occasion was enjoyed by all. After dinner, members and guests were addressed by Professor B. Burgess, Vice Chancellor, University of Leicester.
Scientific Business
Thursday 10 April 2008 2.00 p.m.
The Osler lecture—Genetic fingerprinting and beyond
Sir Alec Jeffreys
3.00 p.m.
(1) Endothelin receptor antagonism offers patients with chronic kidney disease potential for cardiovascular and renal protection
N. Dhaun, J. Goddard, D.J. Webb, V. Melville, P. Lilitkarntakul, N. Johnston. Queen's Medical Research Institute, Edinburgh
Background: Endothelin 1 (ET-1) is implicated in the development and progression of chronic kidney disease (CKD). Work in our department has suggested beneficial effects of selective antagonism of the ET-A receptor with preservation of ET-B receptor function.
Aim: We therefore studied the effects of selective ETA receptor antagonism with BQ123 on systemic and renal haemodynamics, arterial stiffness, endothelial function and proteinuria in patients with CKD.
Methods: We conducted two randomized double-blind studies comparing BQ123 with placebo in 22 patients with proteinuric CKD. All patients were treated with ACE inhibitors and/or angiotensin receptor blockers.
Results: In the first study, BQ123 reduced BP (mean arterial pressure: –11 ± 2%, P < 0.01 vs. placebo), and arterial stiffness (pulse wave velocity: –9 ± 1%, P < 0.01 vs. placebo), but had little effect on endothelial function. Despite the reduction in BP, BQ123 significantly increased renal blood flow (26 ± 6%, P < 0.01 vs. placebo), and reduced renal vascular resistance (–30 ± 7%, P < 0.01 vs. placebo). Importantly, BQ123 produced a sustained reduction in proteinuria (–28 ± 4%, P < 0.01 vs. placebo). Subjects with higher baseline proteinuria had a greater absolute (but similar%) reduction in proteinuria after BQ123. As the reduction in PWV and proteinuria might, in part, be explained by reduction in BP, 12 of these patients undertook a sub-study using nifedipine as an active control for the BP reduction seen with BQ123. Despite a similar reduction in BP with both agents, BQ123 caused a greater reduction in PWV and proteinuria than nifedipine.
Conclusion: Selective ET-A receptor antagonism is effective at reducing BP, proteinuria and arterial stiffness in patients with CKD currently treated for hypertension. Furthermore, these acute studies suggest a reduction in proteinuria independent of BP. If maintained longer term, selective ET-A receptor antagonism would confer both cardiovascular and renal protective effects in patients with CKD on top of standard therapy.
3.25 p.m.
(2) Aberrant inflammatory responses and seasonal deficit in regulatory T-cell activity to Epstein-Barr virus cause immune activation in multiple sclerosis
M.A. Vickers, J. Scattergood, N. Marshall, L.E. Christie, K.C. Thomas, W.J. Pickford, C. Linington, R. Johnston, C.E. Counsell, R.N. Barker. University of Aberdeen
Background: It is now accepted that latent Epstein-Barr virus (EBV) infection is associated with multiple sclerosis (MS). MS is also linked with certain class II HLA alleles, which interact with CD4+ T cells. Latent membrane protein 1 (LMP1) is rich in high affinity class II HLA epitopes that stimulate immunosuppressive IL-10 secreting regulatory T cells in healthy individuals.
Aims: To characterize peripheral blood CD4+ helper cell responses to peptides spanning LMP1 in patients with MS.
Methods: Serology, thymidine incorporation, enzyme-linked immunosorbent assays, flow cytometry.
Results: All 252 patients with MS had been infected with EBV (P < 10–10). Less IL-10 was induced by LMP1 peptides in 73 patients vs. 32 healthy volunteers (P < 0.001) and 23 relatives or partners (P = 0.089). In contrast, 
-IFN (P = 0.02 and 0.01), proliferative (P = 0.12 and 0.004) and, especially, IL-17 responses (P < 0.001) were over-represented in patients. As the prevalence of MS is correlated with latitude, and vitamin D is required for IL-10 secretion by Treg cells, we looked for evidence of seasonal variation. Less IL-10 was induced in winter vs. summer (P = 0.044, 0.023, 0.065, respectively). Cultured dendritic cells transfected with recombinant LMP1 stimulated tolerance to other antigens in healthy controls, but augmented Th1 responses to such antigens in patients.
Conclusion: In patients with MS, latent EBV infection causes a chronic inflammatory state that is transferrable to bystander antigens. These data suggest how genetic background, infection, diet and sunlight all interact to cause human autoimmunity in a way that suggests several therapeutic strategies.
4.15 p.m.
(3) Helicobacter pylori virulence: an evolving problem in the stomachs of individuals and families
J. Atherton1, R. Argent1, E. El-Omar2. 1Wolfson Digestive Diseases Centre, University of Nottingham, Nottingham and 2Department of Medicine and Therapeutics, University of Aberdeen
Background: The two major virulence factors of the gastric pathogen H. pylori are the vacuolating cytotoxin, VacA and the cytotoxin-associated gene product A, CagA. Both are polymorphic, and strains with pathogenic forms are more frequently associated with peptic ulcers or gastric adenocarcinoma. Conventional thought is that strains remain pathogenic or non-pathogenic throughout the life of their human host, and are transmitted within families with unchanged pathogenicity.
Aim: We hypothesized that strains may adapt by evolution of their virulence factors.
Results: We studied 44 adult relatives of gastric cancer patients from western Scotland from 13 family groups. Culture of multiple H. pylori isolates from gastric biopsy specimens showed that 41 relatives had single strain infections, as assessed by identical genetic fingerprinting by RAPD-PCR and partial nucleotide sequence analysis of housekeeping genes yphC and mutY. However, in six cases, single colony isolates from these strains showed duplications, deletions or recombination events within the cagA gene resulting in different numbers of CagA tyrosine phosphorylation sites and different CagA phenotypes (changed CagA phosphorylation level during co-culture with epithelial cells and changed effects on the cytoskeleton). Evolution of vacA had occurred in 2 individuals (by recombination) and resulted in changed vacuolating phenotype. Analysis of family groups showed 17 examples of strain passage between family members, and in eight (47%) of these cagA or vacA had evolved to change strain virulence.
Conclusion/Discussion: H. pylori strains evolve in individual stomachs to change virulence and such evolved strains are commonly passed within families. Thus studies associating strain virulence with disease should be interpreted with caution, family members should not be assumed to have strains of equivalent virulence, and vaccination strategies based on virulence factors may be unsound.
4.40 p.m.
(4) A reduction in the number of mitochondrial DNA molecules in single cells during early embryogenesis leads to the rapid segregation of genotypes, and explains the diverse range of clinical phenotypes in patients with mitochondrial disease
L.M. Cree, D.C. Samuels, S. Chuva De Sousa Lopes, H. Karur Rajasimha, P. Wonnapinij, J.R. Mann, H.H.M. Dahl, P.F. Chinnery. Mitochondrial Research Group, Newcastle University
Mitochondrial DNA (mtDNA) mutations affect at least 1 in 5000 of the UK population and usually present with neurological disease. Many patients harbour varying proportions of mutant and wild-type (normal) mtDNA (called heteroplasmy), and the percentage of mutant mtDNA correlates with the clinical phenotype.
MtDNA is exclusively inherited from the mother, but different siblings inherit markedly different amounts of mutant mtDNA, leading to striking differences in disease severity. The cellular and molecular mechanisms responsible for this variability are not known, limiting our ability to give accurate genetic counselling to families.
Previous work in mice showed that the variation in heteroplasmy is also present in unfertilized primary oocytes, indicating that the mechanism must occur during early embryonic development, either before or during the formation and maturation of the female germ line.
To address this issue we initially measured the amount of mtDNA within single cells in pre-implantation mouse blastocysts in vitro, and subsequently we studied single primordial germ cells (PGCs) within the developing germ line and mature gonad in stella-GFP mice. Our observations show that the partitioning of mtDNA molecules into different cells pre- and post-implantation, followed by the segregation mtDNA between proliferating PGCs, is responsible for the different levels ofheteroplasmy seen in the offspring.
We have defined the mitochondrial ‘genetic bottleneck’. This has direct implications for our understanding of the transmission of pathogenic mtDNA mutations in humans, and the recurrence risks of mtDNA disease.
5.05 p.m.
(5) T3 Acts via thyroid hormone receptor 
to regulate skeletal development and adult bone mass
J.H. Duncan Bassett, Graham R. Williams. Hammersmith Hospital, Imperial College London
Background: Hypothyroidism delays bone development whereas thyrotoxicosis causes osteoporosis. Understanding of how bone responds to thyroid disease has been challenged by studies proposing thyroid stimulating hormone (TSH) as a negative regulator of bone and suggesting bone loss in hyperthyroidism results from TSH deficiency.
Aim: To investigate this issue we characterized mutant mice harbouring dominant negative mutations or deletions of the genes encoding TR and TRβ.
Results: Ossification, linear growth and bone mineralization were retarded in juvenile TR mutants whereas these parameters were advanced in TRβ mutants. In contrast, adult TR mutants had increased cortical and trabecular bone whereas TRβ mutants were osteoporotic. Downstream signalling analyses revealed skeletal hypothyroidism in TR mutants but skeletal thyrotoxicosis in TRβ mutants. TR is expressed at 15-fold higher levels in bone than TRβ, whereas TRβ controls negative feedback regulation of TSH. Thus, TR is functionally predominant in bone and skeletal responses to disruption of TRβ are secondary to effects on systemic thyroid status. To determine the skeletal role of TSH we compared two mouse strains with congenital hypothyroidism. Pax8-/- mice have thyroid agenesis, 1900-fold increased TSH levels and a normal TSH receptor (TSHR), whereas hyt/hyt mice have 2300-fold elevated TSH but a non-functional TSHR. We reasoned these mice must display opposing phenotypes if TSH has a major role in bone. However, Pax8-/- and hyt/hyt mice both having delayed ossification, reduced cortical bone, a trabecular bone remodelling defect and reduced mineralization.
Conclusion: These data exclude a role for TSH in the skeleton in vivo and indicate the pituitary-thyroid axis regulates bone via TR-mediated T3 actions. Increased bone mass in TR mutant mice identifies TR as a potential therapeutic target in the prevention and treatment of osteoporosis.
9.00 a.m.
(6) The burden of disease associated with filaggrin mutations: a population based, longitudinal birth cohort study
J. Henderson1, K. Northstone1, S.P. Lee2, H. Liao2, Y. Zhao2, S. Mukhopadhyay2, G. Davey Smith1, C.N.A. Palmer2; W.H.I. Mclean2, A.D. Irvine3. 1University of Bristol, 2University of Dundee and 3Trinity College Dublin
Background: Atopic disease is a major health problem. We have demonstrated that mutations in the filaggrin gene (FLG) confer major susceptibility to eczema and related asthma. Using a large population-based birth cohort study we determined the natural history and burden of atopic disease conferred by the two commonest FLG mutations.
Methods: We analyzed the effect of the commonest null alleles (R501X and 2282del4) on several atopic phenotypes, at various ages, in a cohort of 
7000 English children born in 1990–1991.
Results: FLG null alleles associated strongly with both atopic and non-atopic eczema; eczema associated with these mutations presents in early life and is more persistent. FLG mutations confer a risk of both early wheeze and childhood asthma in the context of prior eczema. Strong associations were identified with sensitization to grass, house dust mite and cat and, in particular, sensitization to multiple allergens.
Conclusions: FLG mutations are strong genetic determinants of eczema, early wheeze and asthma in the context of eczema, and atopic sensitization. They also confer risk of a particular trajectory for children with eczema, with increased duration of disease and greater risk of asthma and multiple allergic sensitizations. FLG alleles help define the risk profile of children with eczema and help define the ‘eczema plus early wheeze’ and ‘eczema plus asthma’ phenotypes.
9.25 a.m.
(7) Improved endothelial function in coronary artery disease
C. Delles1, J.A. Dymott1, C.A. Hamilton1, J.P. Rocchiccioli1, G.J. Bryce2, G.A. Berg3, A.F. Dominiczak1. 1BHF Glasgow Cardiovascular Research Centre, University of Glasgow, Glasgow, 2Department of Vascular Surgery, Gartnavel General Hospital, Glasgow and 3Department of Cardiothoracic Surgery, Western Infirmary, Glasgow
We have previously shown that LDL cholesterol levels determine endothelium-dependent vasodilation in coronary artery disease (CAD). Here we examine if a change in clinical practice leading to more intensive cholesterol lowering resulted in improved endothelial function.We studied saphenous vein specimens of patients with CAD undergoing surgical revascularisation and controls in 2006/7 and compared results with those of age matched patients and controls examined in 2002/3. Endothelium dependent vasodilation was assessed by maximum relaxation to calcium ionophore A23187 [GenBank] (10 µM) in organ baths and vascular superoxide (O2–) production by lucigenin enhanced chemiluminescence. Pre-incubation of rings with rotenone (3 µM) and allopurinol (100 µM) was used to assess the amount of mitochondria and xanthine oxidase derived O2–, respectively.
In patients with CAD total (4.0 ± 0.2 vs. 5.1 ± 0.2 mmol/l) and LDL cholesterol levels (2.0 ± 0.1 vs. 3.3 ± 0.2 mmol/l) were significantly (P < 0.001) lower in 2006–7 (n = 38) compared to 2002/3 (n = 26) and remained unchanged in controls (P = n.s.). Endothelium dependent vasodilation was significantly (P < 0.001) greater in recent (47 ± 4%) compared to previous patients with CAD (23 ± 3%; n = 32 each) and remained unchanged in controls (64 ± 4 vs. 66 ± 3%; P = n.s.; n = 14 each). In contrast, vascular O2– generation was similar in 2006/7 compared to 2002/3 both in patients with CAD (0.78 ± 0.09 vs. 0.74 ± 0.07 nmol/mg/min; P = n.s.; n = 28 each) and in controls (0.37 ± 0.06 vs. 0.42 ± 0.07 nmol/mg/min; P = n.s.; n = 10 each). Rotenone (
O2–, –0.22 ± 0.07 vs. –0.02 ± 0.03 nmol/mg/min; P = 0.013) and allopurinol (O2–, –0.12 ± 0.05 vs. 0.04 ± 0.04 nmol/mg/min; P = 0.015) inhibited O2– production significantly more in patients with CAD than in controls. Intensive lipid lowering is associated with improved endothelial function in patients with severe CAD, whereas vascular oxidative stress remains unchanged. This directs us to new drug gable targets with mitochondrial and orate pathways as attractive candidates.
9.50 a.m.
(8) Evidence that mutations in the SQSTM1 gene are necessary and sufficient to Paget's disease of bone
A. Daroszewska1, J. Rojas1, R. van't Hof1, M. Helfrich1, R. Layfield2, S.H. Ralston1. 1University of Edinburgh and 2University of Nottingham
Introduction: Paget's disease of bone (PDB) is a common disease with a strong genetic component characterized by focal increases in bone turnover. Mutations affecting the ubiquitin associated domain of the SQSTM1 gene predispose to PDB but it has been argued that these mutations alone are insufficient to cause the disease and that an additional environmental trigger is required. Here we report upon the generation and characterization of two novel animal models for PDB which carry different mutations in the SQSTM1 gene.
Methods: We conducted skeletal phenotyping of mice with a ‘knock-in’ of the P392L mutation of SQSTM1 which is the most common cause of familial PDB and of mice with a truncating mutation at codon 409 (S409X) gene which deletes most of the UBA domain and mimics the truncating mutations that have been described in humans.
Results: Mice which carried the S409X mutation developed focal lytic lesions at the lower limbs with a penetrance of 80% by 12 months of age in heterozygotes and 91% in homozygotes. Histological analysis showed that these lesions were caused by focal increases in bone turnover with increased osteoclast and osteoblast activity as in human PDB. Mice with the P392L mutation also developed lytic lesions with a penetrance of 100% by 12 months in homozygotes and 66% in heterozygotes. Studies in vitro in both mouse strains showed a significant increase in RANKL-induced osteoclast formation in SQSTM1 mutants compared with wild type.
Conclusions: Mice carrying mutations of the SQSTM1 gene exhibit several phenotypic features of human PDB including focal osteolytic lesions and increased osteoclast formation in vitro. We conclude that SQSTM1 mutations are sufficient to cause a PDB-like disorder in mice confirming that these mutations can cause PDB in the absence of any additional trigger. The mutant mice represent the first animal model of PDB to be developed and will be a valuable resource to investigate disease mechanisms and to explore new treatment strategies.
10.15 a.m.
(9) Peroxisome proliferator activated receptor- (PPAR-) in Colon cancer
A. Shonde, K. Mccartney, D. Bell, A. Bennett, C.J. Hawkey. Wolfson Digestive Diseases Centre, University Hospital, Nottingham
Introduction: Peroxisome proliferator activated receptor (PPAR) is a ligand activated nuclear hormone affecting transcription of many genes potentially involved in malignancy. We reported marked depression in human colorectal cancers (Gut52:1317). We therefore investigated its role in development of colonic polyps in APCmin/+ mice using ligand activation and gene deletion.
Methods and Results: Time course of development of polyps and potentially precursor aberrant crypt foci (ACF) in 20 APCmin/+ PPAR–/– vs. 20 APCmin/+ mice was established by sacrificing mice at 12, 18 and 24 weeks or when anaemia/loss of weight indicated significant tumour burden (term). Only one ACF was detected in APCmin/+ mice (at term), but evident at 12 weeks, increasing to five at term in APCmin/+ PPAR–/– mice. Polyps also developed earlier (12 weeks) and were more numerous in APCmin/+ PPAR–/– mice (2.4 ± 0.68 SEM at term vs. 1.0 ± 0.55 in APCmin/+ mice P > 0.0004). Twenty APCmin/+ PPAR–/– and 20 APCmin/+ mice were then dosed with methylclofenapate (MCP, PPAR ligand) 25 mg/kg or safflower oil (control). MCP reduced mean polyp numbers from 2.26 (±0.34) to 1(±0.25) [P > 0.014] and area from 78.72 µm2 (± 13.41) to 34.54 µm2 (± 8.57) [P > 0.008] in APCmin/+ mice but had no effect in APCmin/+ PPAR–/– mice. Polyps 2.65 (±0.40) vs. 2.60 (±0.43), area (159 µm2 [±31.26] vs. 215 µm2 [±35.57]). We used immunohistochemistry and expression profiling to investigate associated changes in polyp gene expression. APCmin/+ PPAR–/– mice showed enhanced expression of cyclo-oxygenase-2 (COX-2) and β catenin at mRNA and protein level, localized to surface epithelium and (for COX-2) stromal cells. Because COX inhibition by non-steroidal anti-inflammatory drugs (NSAIDs) can potentially enhance expression of the natural PPAR ligand leukotriene (LT) B4, by substrate diversion we used cell and organ culture to show indomethacin increased LTB4 by 3.1-fold in colonic and 8.3-fold in oesophageal adenocarcinomas. In colonocytes transfected with PPAR- and a PPRE response element, LTB4 enhanced PPAR- expression (5-fold) and reduced thymidine incorporation (14% [P > 0.0009]) but had no effect on apoptosis (Annexin 5 staining).
Conclusions: PPAR-, which has reduced expression in human colorectal cancer, inhibits cellular proliferation and experts a negative effect on ACF and polyp development in APCmin/+ mice, affecting expression of genes associated with malignancy. Substrate diversion of arachidonic acid metabolism with increased synthesis of the PPAR- ligand LTB4 could mediate chemo preventative actions of NSAIDs.
11.10 a.m.
(10) COPD is associated with a selective defect in phagocytosis of bacterial pathogens
L.E. Donnelly, A.E. Taylor, T.K. Finney-Hayward, P.J. Barnes. Airway Disease, National Heart and Lung Institute, Imperial College London, London
Background: Alveolar macrophages (AM) act to remove inhaled particles and pathogens from the lungs. Patients with chronic obstructive pulmonary disease (COPD) suffer acute exacerbations due to bacterial infections and these are implicated in the progression of the disease.
Aim: To investigate whether AM are defective in pathogen clearance in COPD, leading to colonization and exacerbations.
Methods: Macrophage phagocytosis was investigated using both AM and monocyte-derived macrophages (MDM) from non-smokers (NS), smokers (S) and COPD subjects. Phagocytosis was measured by exposing cells to fluorescently labelled inert beads or bacteria (Haemophilus influenzae, Streptococcus pneumoniae or Escherichia coli 1 mg/ml). Internalized particles were detected fluorimetrically and receptor expression by FACS.
Results: AM and MDM phagocytosed comparable numbers of inert beads suggesting that MDM reflect AM. MDM from the three groups phagocytosed beads to a similar extent. However, phagocytosis of H. influenzae, S. pneumoniae and E. coli was reduced in both AM and MDM from COPD patients compared with NS. This suggested that defective phagocytosis in COPD was restricted to bacterial clearance and recognition. However, cell surface scavenger receptor expression (MARCO, mannose receptor, CD163, CD36, TLR2, TLR4) was similar in all groups. Phagocytosis of E. coli by COPD MDM was more susceptible to colchicine than MDM from NS or S (COPD: 28.3 ± 2.8, NS: 39.4 ± 3.0, S: 39.2 ± 2.9 AFU x103, n = 6–8, P < 0.05), suggesting that COPD cells are more susceptible to microtubule disruption. Microtubules are stabilized by acetylation, but COPD MDM expressed significantly less acetylated tubulin than cells from NS [0.48 ± 0.06 vs. 0.23 ± 0.04 (expression relative to actin; n = 6–10, P < 0.05)], however, expression of the relevant deacetylases, HDAC6 or SIRT2 was similar.
Conclusion: Reduced bacterial phagocytosis in both AM and MDM indicates an intrinsic defect in COPD patients and is a possible susceptibility marker for airway obstruction in smokers.
11.35 a.m.
(11) Kisspeptin potently increases reproductive hormone release in women with hypothalamic amenorrhea – a potential novel therapy for infertility
W. Dhillo1,*, C. Jayasena1,*, O. Chaudhri1, G. Nihjer1, K. Murphy1, A. Ranger1, A. Lim2, D. Patel2, A. Mehta2, C. Todd2, R. Ramachandaran1, V. Salem1, G. Stamp1, M. Donaldson1, M. Ghatei1, S. Bloom1. 1Imperial College London, Hammersmith Campus and 2Imperial College Healthcare NHS Trust, Charing Cross Hospital, London
Background: Hypothalamic amenorrhea is defined as the cessation of menstruation due to abnormal signalling between the hypothalamus and the pituitary gland. It accounts for over 30% of cases of amenorrhea in women of reproductive age. Current treatments for hypothalamic amenorrhea have limited success rates and significant side effects. Kisspeptin, the endogenous ligand of the GPR54 receptor, has recently been shown to be a key regulator of the hypothalamo-pituitary-gonadal (HPG) axis. Kisspeptin or GPR54-null mice exhibit reproductive dysfunction. Exogenous administration of kisspeptin can effectively treat hypothalamic amenorrhea in animal models. We have recently shown that subcutaneous (sc) injection of kisspeptin (6.4 nmol/kg) potently stimulates reproductive hormone release in human females with normal menstrual cycles. The effects of kisspeptin administration to human females with hypothalamic amenorrhea are not known.
Aim: To investigate the effects of sc injection of kisspeptin on reproductive hormone in with hypothalamic amenorrhea.
Methods: Volunteers with hypothalamic amenorrhea received a sc bolus injection of either kisspeptin (6.4 nmol/kg) or 0.9% saline (n = 6 per group). Blood was sampled prior to injection and then every 30 min for 4 h post-injection for measurement of plasma reproductive hormones. The local ethics committee approved this study.
Results: Kisspeptin potently increased mean luteinizing hormone (LH) and follicle stimulating hormone (FSH) when compared to saline injection (mean change 4 h post-injection compared to baseline: LH (IU/l); saline –0.5 ± 0.6, kisspeptin 24.0 ± 3.5, P < 0.0001 vs. saline. FSH (IU/l); saline –0.6 ± 0.7, kisspeptin 10.2 ± 3.5, P < 0.001 vs. saline).
Conclusion: This is the first report demonstrating that elevation of plasma kisspeptin in women with hypothalamic amenorrhea potently stimulates LH and FSH release. Thus kisspeptin may form the basis of a novel therapy for infertility in these patients.
*Joint first authors.
12.00 noon
(12) Drug target discovery using proteomics in a clinically relevant model of sepsis: identification of cyclophilin as a novel mediator of organ injury
J.W. Dear1, A. Leelahavanichkul2, S.L. Constant2, P.S.T. Yuen2, D.J. Webb1, R.A. Star2. 1Clinical Pharmacology Unit, Centre for Cardiovascular Science, Edinburgh University and 2Renal Diagnostics & Therapeutics Unit, National Institutes of Health, USA
Sepsis-induced multi-organ failure continues to have a high mortality. The liver is an organ central to the disease pathogenesis. The objective of this study was to identify the liver proteins that change in abundance with sepsis and, therefore, identify new drug targets.
We used a mouse model of sepsis based on cecalligation and puncture (CLP) but with fluid and antibiotic resuscitation. Liver proteins that changed in abundance were identified by difference in-gel electrophoresis (DIGE). We compared liver proteins from 6 h post-CLP to sham-operated mice (‘early proteins’) and 24 h post-CLP with 6 h post-CLP (‘late proteins’). Proteins that changed in abundance were identified by tandem mass spectrometry. We then inhibited the receptor for one protein and determined the effect on sepsis-induced organ dysfunction. The liver proteins that changed in abundance after sepsis had a range of functions such as acute phase proteins, coagulation, ER stress, oxidative stress, apoptosis, mitochondrial proteins and nitric oxide metabolism. We found that cyclophilin increased in abundance after CLP. When the receptor for this protein, CD 147, was inhibited, sepsis-induced renal dysfunction was reduced (serum creatinine 24 h post-CLP: sham antibody 0.59 ± 0.07 mg/dl; anti-CD147 antibody 0.39 ± 0.05 mg/dl, n = 20, P = 0.04). There was also a significant reduction in serum cytokine production when CD147 was inhibited (serum TNF-a 24 h post-CLP: sham antibody 225 ± 40 pg/ml; anti-CD147 antibody 105 ± 25 pg/ml, n = 8, P = 0.03. Serum IL-6 24 h post-CLP: sham antibody 82 ± 14 ng/ml; anti-CD147 antibody 16 ± 7 ng/ml, n = 8, P = 0.01. Serum IL-10 24 h post-CLP: sham antibody 1370 ± 146 pg/ml; anti-CD147 antibody 667 ± 265 pg/ml, n = 8, P = 0.03).
By applying proteomics to a clinically relevant mouse model of sepsis we identified a number of novel proteins that changed in abundance. The inhibition of the receptor for one of these proteins, cyclophilin, attenuated sepsis-induced acute renal failure. The application of proteomics to sepsis research can facilitate the discovery of new therapeutic targets.
12.25 p.m.
(13) The VEGF receptor neuropilin-1 is expressed in a soluble form in human platelets and plays a novel role in thrombin-induced platelet aggregation
J. Martin, I. Zachary, D. Sanz, P. Frankel. University College London, London
Background: The vascular endothelial growth factor (VEGF) receptor Neuropilin-1 (NRP-1) mediates VEGF signalling and functions in endothelial cells, but its role in platelet biology is unknown.
Aim: To investigate the role of NRP-1 in platelet aggregation.
Methods: Platelet aggregation was determined in an aggregometer, NRP-1 expression by western blot and immunofluorescent confocal microscopy.
Results: A highly specific antagonist of ligand binding to the NRP-1 extracellular domain (EG00086) caused a striking inhibition of aggregation of fresh human platelets in response to increasing concentrations of thrombin. In addition, we also found that treatment with VEGF similarly inhibited thrombin-induced platelet aggregation. The effect of EG00086 was concentration-dependent with a maximum inhibition at 100 µM, in agreement for the concentration-dependence for inhibition by EG00086 of VEGF binding to NRP-1. Western blot analysis showed that NRP-1 was expressed in human platelets both as the normal full-length protein of 130 kDa also expressed in endothelial cells, and additionally as a smaller 70 kDa species, that was not readily detectable in endothelial cells. This smaller NRP-1 form was strongly expressed in platelets and was detectable using antibody specific for the NRP-1 extracellular domain indicating that it represents a soluble NRP-1 species. Immunofluorescent staining of fixed, permeabilized platelets showed that NRP-1 was expressed in structures similar to alpha-granules, but did not show strong co-localization with von Willebrand Factor.
Conclusion: Our data reveal a novel role for NRP-1 in platelet aggregation and indicate that platelets express a soluble form of NRP-1 comprising the extracellular domain. We propose that release of soluble NRP-1 by activated platelets may play an important role in regulation of platelet function, and that NRP-1 antagonists may have therapeutic potential for combating human thromboembolic disease.
This work was funded by the British Heart Foundation (Grants RG/02/2001 and CH/92022).
3.00 p.m.
(14) Molecular mimicry between lysosomal membrane protein 2 and the bacterial adhesin fimH identies a pathogenic epitope in crescentic glomerulonephritis
A. Rees1, D. Cunningham2, O. Ashour1, C. Alderson2, D. Kerjaschki1, R. Kain1. 1Institute of Clinical Pathology, Medical University Vienna, Austria and 2Immunology Program, School of Medicine, University of Aberdeen, Aberdeen
Background: Pauci-immune crescentic glomerulonephritis (FNGN) is a severe inflammatory disease and a common cause of renal failure. The patients usually have anti-neutrophil cytoplasmic antigens (ANCA) autoantibodies specific for either myeloperoxidase (MPO) or proteinase 3 (PR3). Previously we showed that some of the patients also have antibodies to LAMP-2 a glycoprotein expressed in lysosomes and on the cell surface that has key roles in cell survival. Their significance is unknown.
Aims: We aim to analyse the significance of anti-LAMP-2 antibodies and their potential role in pathogenesis in FNGN.
Methods: Specific ELISA and western blotting were used to identify anti-LAMP-2 antibodies; epitopes were mapped using recombinant proteins and synthetic peptides. Pathogenicity of anti-LAMP-2 antibodies was tested in vivo in WKY rats and in vitro using microvascular endothelial cells.
Results: Circulating anti-LAMP-2 antibodies were present in 78 of 84 (95%) of patients with active FNGN. These bound epitopes on the protein backbone of glycosylated LAMP-2 on surface of transfected CHO cells. Antibodies to LAMP-2 can cause injury because intravenously injected polyclonal anti-LAMP-2 antibodies induced FNGN with crescents in 14 WKY rats. A monoclonal anti-LAMP-2 antibody induced apoptosis of microvascular endothelium in vitro whereas control monoclonals to CD4 and PR3 had no effect. Further analysis showed the patients’ auto pg/ml o antibodies recognized two common LAMP-2 epitopes one of which (P41-49) was 100% homolous to the bacterial adhesin fimH. Binding studies demonstrated the patients’ anti-P41-49 auto antibodies cross-reacted with fimH. Furthermore, 10 rats immunized with fimH developed anti-fimH antibodies that cross-reacted with LAMP-2 and induced FNGN. Finally, 9 of 13 prospectively studied patients with FNGN had microbiologically proven infections with fimbriated pathogens shortly before presentation.
Conclusion: Thus, fimH triggered autoimmunity to LAMP-2 provides a novel clinically relevant molecular mechanism for the development of pauci-immune FNGN.
3.25 p.m.
(15) The envelope protein of the hepatitis c virus (E2) inhibits IL-2 secretion from T lymphocytes
D. Kelleher1, D. Petrovic1, L. Golden-Mason2, S. Mitchell1, Y. Volkov1, C. O’Farrelly1, A. Long1. 1Clinical Medicine, Trinity College Dublin, Dublin and 2Education & Research Centre, St Vincent's Hospital, University College Dublin, Dublin
Background: T-cell activation is one of the primary responses of the adaptive immune system to pathogens, such as Hepatitis C virus (HCV). Production of IL-2 is central to this response. We have shown that the PKCβ enzyme is critical for IL-2 secretion but not mRNA production. More recently we have shown that E2, through its interaction with CD81, modulates PKCβ activity by stimulating its sequestration in lipid rafts, preventing T-cell migration.
Aim: The aim of this study was to investigate the effect of E2 on T cell IL-2 production.
Results: Analysis of cytokine levels in extracts of snap-frozen liver biopsies from individuals infected with HCV revealed significantly lower levels of IL-2 in these patients when compared to those with other inflammatory liver diseases, such as ALD and PBC (IL-2 levels in nanograms per 100 mg protein: 5 ± 0.93 (HCV) vs. 25 ± 2.9 (ALD), P < 0.027 vs. 20 ± 2 (PBC), P < 0.003). In a HuT 78 T-cell model, pre-treatment of cells with E2 but not C22 or NS3 attenuated PMA stimulated IL-2 production [IL-2 levels in pg/ml: PMA stimulated cells 889.3 ± 19.1 vs. E2/PMA stimulation 105.9 ± 0.2, P < 0.016, (resting cells 20 ± 0.9)]. This was also observed in mitogen stimulated PBMCs (healthy donors). Inhibition of IL-2 was at the level of secretion and not transcription (demonstrated by Real Time PCR). Intracellular IL-2 was detectable by immunofluorescence in PMA activated cells pretreated with E2. Multiplex analysis revealed that E2 inhibited TNF, IFN and IL-4 suggesting a common secretory pathway defect. Treatment of HuT 78 cells with the lipid raft disruptor methyl-β-cyclodextrin (MCD) reversed the ability of E2 to inhibit IL-2 secretion.
Discussion: These data suggest that targeting of PKCβ to the lipid raft microdomains in the T-cell membrane, following E2-CD81 interaction, disrupts the association of PKCβ with the microtubule cytoskeleton, thereby inhibiting export of IL-2. E2-mediated inhibition of IL-2 secretion represents a mechanism whereby HCV can undermine the human immune response to establish persistent infection and chronic disease.
3.50 p.m.
(16) Obesity therapy: bariatric surgery without the surgeon
B.C.T. Field, A.M. Wren, V. Peters, K.C.R. Baynes, S. Alsaraf, V. Amber, K.J. Wynne, N.M. Martin, M.A. Ghatei, S.R. Bloom. Department of Metabolic Medicine, Imperial College London
Background: Bypass surgery is the only really effective treatment for obesity but is associated with significant mortality and morbidity. It is thought to work by permanently elevating circulating gut hormones. Peptide YY (PYY) and oxyntomodulin (OXM) are secreted post-prandially from intestinal L-cells and act as key satiety signals. We have previously shown that administration of either PYY or OXM alone reduces food intake in man. However, supraphysiological levels are required and may cause nausea.
Aim: To investigate whether low dose PYY and OXM administered in combination have additive effects on food intake compared to either gut hormone alone.
Methods: We performed a randomized, double-blind, placebo-controlled, crossover study in nine overweight volunteers (REC no. 06/Q0406/50). Subjects participated in six study days, receiving intravenous infusion of PYY 2 nmol/m2/90 min (established dose), PYY 1 nmol/m2/90 min (low dose), OXM 3.0 pmol/kg/min (established dose), OXM 1.5 pmol/kg/min (low dose), PYY 1 nmol/m2/90 min + OXM 1.5 pmol/kg/min (combined low dose) or saline. Food intake was compared to the saline control day.
Results: Low dose PYY did not reduce food intake. Established dose PYY resulted in significant nausea in the first four subjects and was not administered thereafter. Neither established dose nor low dose OXM altered food intake significantly in this study. However, combined low dose PYY + OXM significantly reduced food intake by 35.5% (P < 0.01). This was achieved without adverse effects. There was no difference in ascertained food palatability between treatments.
Conclusion: PYY and OXM at low doses have additive effects in reducing food intake. The combination is free of adverse effects. Such combination treatment mimics physiology and allows use of lower doses of gut hormones to achieve effective reduction in appetite, while minimising side effects. This represents an exciting new approach in the development of a safe and effective obesity therapy.
4.45 p.m.
(17) Peripheral blood mononuclear cells isolated from Charcot patients differentiate into osteoclasts which exhibit enhanced RANKL mediated bone resorption
M. Edmonds1, N. Petrova1, G. Mabillleau2, A. Sabokbar2. 1Kings College Hospital, London and 2Botnar Research Centre, Oxford University
Background: Charcot osteoarthropathy is characterized by bone and joint destruction but the cause is unknown.
Aim: The aim was to investigate the cellular mechanisms of osteoclastic bone resorption in Charcot osteoarthropathy and the effect of receptor activator of nuclear factor 
β ligand (RANKL).
Method: Peripheral blood mononuclear cells (PBMCs), which can act as osteoclast precursors, were isolated from nine diabetic patients with acute Charcot osteoarthropathy, eight diabetic control patients and eight healthy subjects and were cultured on dentine slices in the presence of macrophage-colony stimulating factor (M-CSF), which is required for the survival of osteoclast precursors. Functional evidence of osteoclastic resorption was determined after 21 days and was expressed as the percentage of eroded surface by the number of newly formed osteoclasts.
Results: The mean area of resorption was significantly increased in the Charcot patients (0.264 ± 0.06%) compared with diabetic patients (0.000 ± 0%) and healthy controls (0.004 ± 0.0038%), (P < 0.001). The addition of RANKL led to a marked increase in the extent of resorption in the Charcot patients from 0.264 ± 0.06% to 41.6 ± 8.1%, (P = 0.008). Diabetic patients also increased from 0.000 ± 0% to 14.2 ± 6.5%, P = 0.012 as did healthy controls (from 0.004 ± 0.0038% to 10.4 ± 1.95% P = 0.012), but the extent of resorption after RANKL was significantly greater in the Charcot patients compared with diabetic and healthy controls, (P = 0.003). The addition of osteoprotegerin, an antagonist of RANKL, to the cultures with M-CSF and RANKL led to suppression of bone resorption in the Charcot patients from 41.6 ± 8.1% to 5.9 ± 2.41% (P = 0.001) and also from 14.2 ± 16.5% to 0.45 ± 0.31% (P = 0.012) in the diabetic patients and from 10.4 ± 1.9% to 0.000 ± 0.000% (0.012) in the healthy controls.
Conclusion: These results indicate, that PBMCs isolated from acute Charcot patients, are capable of differentiating into mature osteoclasts and exhibit enhanced RANKL mediated bone resorption. This may be an important factor in the pathogenesis of the Charcot foot.
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