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Abstracts / Molecular Imm

58

icolin-1 expression reflects FCN1 genotypic variation

.M.F. Munthe-Fog ∗, M.E.M. Møller, C.F.H. Honore, T.H.ummelshoj, P.G. Garred

Rigshospitalet, Copenhagen N, Denmark

Introduction: Ficolin-1 is part of the lectin pathway of comple-ent and mainly expressed by monocytes and granulocytes. A large

roportion of Ficolin-1 is found associated to cell surfaces. The FCN1ene harbors several polymorphisms in the promoter region. So farhe impact of these promoter variations on the FCN1 expressionnd on the levels on circulating Ficolin-1 has not been assessed.

Methods: Blood samples were obtained from 100 blood donors.uantitative PCR analysis of FCN1 expression in isolated monocytesnd granulocytes was performed. The Ficolin-1 levels in plasmaamples and the genotype of SNPs in the promoter region of FCN1ere determined. In addition, Ficolin-1 binding to freshly isolatedonocytes from 15 staff members were analyzed by flow cytome-

ry. Cell surface bound Ficolin-1 was eluted with EDTA buffer andicolin-1 concentration was determined both in isolated monocyticulture supernatants and eluates.

Results: Several of the FCN1 SNPs had a significant impact onoth the expression level of FCN1 and on the circulating levelsf Ficolin-1. The impact of the SNPs on Ficolin-1 levels in cultureupernatant from monocytes was essentially the same as observedn plasma. Moreover, the levels of Ficolin-1 in culture supernatantnd the amount of Ficolin-1 bound to the surface of monocytesere strongly correlated.

Conclusion: We show that different SNPs in the promoter regionf FCN1 are associated with gene expression, release of Ficolin-from cells and Ficolin-1 plasma levels. The results suggest that

enetic variations in FCN1 influence the synthesis of Ficolin-1.

oi:10.1016/j.molimm.2011.06.278

59

isk alleles in complement factor H (CFH) and age-related mac-lopathy susceptibility 2 (ARMS2) are independently associatedith systemic complement activation in age-related macularegeneration (AMD)

. Smailhodzic a,∗, C.W. Klaver b, B.J. Klevering a, C.J.F. Boon a,.M.M. Groenewoud a, B. Kirchhof c, M.R. Daha d, A.I. denollander a, C.B. Hoyng a

UMC St Radboud, Nijmegen, NetherlandsErasmus Medical Centre, Rotterdam, the Netherlands, Rotterdam,etherlandsUniversity of Cologne, Cologne, GermanyLeiden University Medical Centre, Leiden, Netherlands

Introduction: Systemic complement activation is associated withMD and has mainly been attributed to a risk allele in CFH gene.hether other important AMD genes also influence complement

ctivation is unclear. In the present case–control study concen-rations of complement components and their activation productsre measured in AMD patients and in controls and correlated withenetic variants in the CFH, ARMS2, C3, CFI and CFB genes.

Methods: A cohort of 197 AMD patients and 150 unaffectedge-matched controls were recruited for the study. Hemolytic com-lement assays (AP50, CP50 and LP50), complement components

C3, CFB, CFI and CFH) and the activation products (C3d, C5a andC5b-9) were analyzed in serum or plasma. The DNA samples wereenotyped for five single nucleotide polymorphisms (SNPs) pre-

gy 48 (2011) 1666–1733 1685

viously associated with AMD in the CFH, ARMS2, C3, CFB and CFIgenes.

Results: The AMD patients had increased activation of the alter-native complement pathway (p = 0.0003) and elevated levels ofcomplement activation components C3d and C5a (p < 0.0001), CFB(p = 0.002), and an increased C3d/C3 ratio (p < 0.0001) calculatedas a measure of C3 activation. While the CFH risk genotype wassignificantly associated with the elevated C3d/C3 ratios obtained,in the absence of CFH risk alleles the ARMS2 risk genotype alsoshowed increased levels of complement activation (p = 0.013). Fur-thermore, the carriers of the CFB protective allele had lower CFBconcentrations.

Conclusions: The current study found evidence showing that CFHand ARMS2 share a common pathway in the pathogenesis of AMD.Especially the C3d/C3 ratio seems to be a strong marker for AMD.

doi:10.1016/j.molimm.2011.06.279

P60

Exome capture and array comparative genomic hybridization ofa monozygotic twin pair discordant for dense deposit disease

J.C.C. Cruz Corchado ∗, C. Nishimura, C. Meyer, K.F. Frees, M.B.Jones, Y.Z. Zang, A. Weaver, M. Ealy, M.S. Hildebrand, J.H. Smith

University of Iowa, Iowa City, United States

Dense deposit disease (DDD) is a complement-mediated renaldisease that leads to end-organ failure requiring dialysis in 50% ofaffected patients. Its underlying pathophysiology is uncontrolledfluid-phase activation of the alternative pathway (AP) of the com-plement system. The limited number of familial cases suggeststhat DDD is a complex genetic disease with a variety of envi-ronmental triggers. Discordant monozygotic twins could be aninformative resource with which to identify possible genetic vari-ability associated with DDD. Herein we report a monozygotic twinpair discordant for DDD. Comparative genetic analysis did notreveal any genetic variants unique to one twin of the pair in genesof the AP, and using multiplex ligation probe amplification (MLPA),no copy number variation was identified in the CFHR gene fam-ily. An expanded analysis was therefore completed by performinggenome-wide exome capture using the Agilent SureSelect HumanAll Exon 50Mb Kit and array comparative genomic hybridization(aCGH) to screen for coding variants and/or copy number varia-tions, respectively, discordant in these siblings. Herein we reportour findings of this investigation.

doi:10.1016/j.molimm.2011.06.280

P61

Mechanism of the activation of the lectin pathway

P. Gál a,∗, D. Héja b, A. Kocsis a, K. Szilágyi a, J. Dobó a, P.Závodszky a, G. Pál b

a Institute of Enzymology, Hungarian Academy of Sciences, Budapest,Hungaryb Department of Biochemistry, Eötvös Loránd University, Budapest,Hungary

Introduction: The lectin pathway of the complement system pro-vides the first line of defense against pathogens. Activation of thelectin pathway is triggered by carbohydrate arrays on the surfaceof micro-organisms. Multimolecular complexes consisting of pat-

tern recognition molecules (MBL and ficolins) and zymogen serineproteases (MASP-1, -2 and -3) initiate the complement cascade.MASP-2 was shown to generate C3-convertase via cleavage of C2