higher protein expression in both primary and metastatic tumourscompared to the matched normal tissues (Mann Whitney p< 0.005)which exhibited minimal protein expression. All three metastaticmelanoma cell lines demonstrated constitutive and inducibleCYP1B1 expression.Conclusion: CYP1B1 is consistently and selectively over expressedin primary and metastatic melanoma and may represent a thera-peutic target for novel CYP1 activated pro-drugs that are currentlyunder development.

O24. Antibody-nanoparticle conjugation for hyperthermictreatment of tumoursK. Vigor, D. Farrell, A. Huhalov, B. Tolner, H. Kogelberg,Q.A. Pankhurst, I.P. Parkin, R. Begent, K. ChesterRoyal Free & University College Medical School UCL, OncologyDepartment, Hampstead Campus, Rowland Hill Street, LondonNW3 2PF, UK

Immunoconjugates of antibodies with toxic agents are becominga significant component of anticancer treatments. One ongoingchallenge to the success of this approach is the exposure of healthytissues to the cytotoxic agents employed. We propose thatmagnetic fluid hyperthermia, using pre-targeted antibody-ironoxide magnetic nanoparticles, can address this issue. In magneticfluid hyperthermia an alternating magnetic field is applied tomagnetic nanoparticles in target tissues causing localised heatingand cell death. The externally employed magnetic field can befocused on the tumour deposit. The system has potential to providepotent and selective cancer-targeted hyperthermic therapy.To test the hypothesis we are using the genetically engineeredhumanised high affinity SM3e single chain Fv (scFv) reactive withthe carcinoembryonic antigen (CEA), a glycoprotein associatedwith epithelial cancers. We have covalently coupled SM3e toperiodate-oxidised dextran coated iron oxide magnetic nanopar-ticles (MNP). The MNP-SM3e complex has been purified and isolatedsuccessfully from unbound SM3e by gel filtration chromatography.The complex has been shown to retain binding to the CEA proteinthrough enzyme linked immunoabsorbent assay (ELISA) and spec-ificity to CEA expressing cells by immunofluorescence. Furthercharacterisation of the MNP-SM3e complex will be discussed inrelation to its therapeutic potential.

O25. VEGF165 transfection of ex vivo expanded keratinocytespromotes healing and neovascularization in porcine fullthickness skin woundsS. Dickens, P. Vermeulen, J.J. VranckxLab of Plastic Surgery and Tissue Engineering Research KULeuven, Belgium

Background: A trigger to enhance Full Thickness Wound (FTW) re-pair is delivery of oxygen and nutritients to the wound site. Afterthe initial inflammatory phase, platelets and keratinocytes (KC)start to secrete Vascular Endothelial Growth Factor (VEGF) toattract Endothelial Cells into the wound granulation tissue. Wepresent a method of ex vivo expansion of autologous KC and lipidmediated gene transfer of a regulable VEGF165 plasmid for thetreatment of FTW.Methods: Autologous KC were cultured to P3 and transfected withan inducible hVEGF165 plasmid and pcDNA6/TR repression mole-cule. This system allows for inducible expression by addition of Tet-racyclin (TC). In a porcine model 12 a 16 Full Thickness Skin Wounds(FTW) were created on the back. We designed 3 groups: The firstreceived 1.2 million transfected KC with addition of 4ug TC (KC/VEGF-group). The second group received 1,2 million transfectedKC without addition of 4 ug TC (KC-group). The third group receivedno KC and was treated with 2 ml saline (S-group). Wounds werecovered with polyvinyl wound chambers and wound fluid (WF)

was collected daily. Biopsies were taken at day 8 and 10 for H&Estaining and immunohistochemistry.Results: At day 10, reepithelialization was significantly enhanced inthe KC/VEGF- and the KC-group compared the S- group (p< 0.01).Moreover, the KC/VEGF- group showed enhanced revascularization(p< 0.0001), compared to the KC- group, as measured with lectinantibodies. However, no difference in reepithelialization wasseen between KC/VEGF- and KC- group.Conclusions: Ex vivo cultured autologous KC enhance reepithelializa-tion in a FTW in a pig model. Overexpression of hVEGF165 by these KCin the early wound phase enhances neovascularization in the woundgranulation tissue. Ex vivo culturing of autologous KC and transfec-tion with VEGF can be useful in reconstructing large skin wounds.

O26. The influence of mmp activity on force generation bydupuytren’s fibroblastsW.A. Townley a, B. Klass a, K.J. Rolfe a, A.D. Cambrey b,P.T. Khaw b, A.O. Grobbelaar a

a RAFT Institute of Plastic Surgery, Mount Vernon Hospital,Northwood, Middlesex HA6 2RN, UKb Ocular Repair and Regeneration Unit (ORB), Institute ofOphthalmology - UCL, London, UK

Introduction: Dupuytren’s disease is a common fibrocontractile dis-order affecting the palmar fascia of the hand that can cause severedisability. Despite advances in surgical management, recurrenceremains a common complication. The matrix metalloproteases(MMPs) are a large family of proteolytic enzymes that play an inte-gral role in scarring in vivo and collagen contraction in vitro. Thepurpose of this study was to assess the influence of MMP on matrixcontraction by Dupuytren’s fibroblasts using the broad-spectrumMMP inhibitor, ilomastat.Methods: Nodule and cord-derived fibroblasts were isolated by ex-plant culture from five Dupuytren’s patients, carpal ligament-de-rived fibroblasts acted as the control. A Culture Force Monitor(CFM) was employed as an in vitro kinetic model of fibroblast-me-diated collagen lattice contraction. Fibroblast-seeded latticeswere allowed to contract for 48 h under basal media conditionsor exposure to ilomastat. The maximum force of contraction(dynes) was recorded. The expression and activity of severalMMPs (-1, -2 and MT1-MMP) and TIMPs (-1, -2) was assessed byRT-PCR and ELISA.Results: Cord and nodule-derived fibroblasts exhibited a two-foldincrease in mechanical tension (150� 15, 180� 20, respectively)under basal conditions compared with carpal ligament (90� 12,p< 0.01). Ilomastat significantly (p< 0.01) inhibited developmentof mechanical tension by cord and nodule but not carpal-ligamentderived fibroblasts. Treatment with ilomastat suppressed MMP-1and -2 activity but did not affect MMP or TIMP gene expression.Conclusions: Dupuytren’s-derived fibroblasts developed greatermechanical tension in the CFM model than control cells. Ilomastatsuppressed MMP-1 activity and the ability of both cord and nodule-derived fibroblasts to generate contractile force, suggesting thatMMP activity may be a therapeutic target in preventing diseaserecurrence in patients.

O27. MRI radiological assessment of distance betweenmetacarpal bones and flexor tendons: Significance in screwfixation of metacarpal fracturesK.H. Tan, A. Dawkins, S. Aslam, R. Page, R. CooperNorthern General Hospital, Herries Road, Sheffield, SouthYorkshire S5 7AU, UK

Introduction and Aims: There are 3 types of metacarpal shaft (MS)fractures: transverse, spiral and comminuted. The use of interfrag-mentary compression screw (lag screw) is well indicated for spiralfractures. However, there is a risk of potential injury to flexor ten-don if the distal end of the lag screw is significantly beyond the

ECSAPS 2006 Abstracts S9

border of the margin of the metacarpal cortex on the volar surfaceof the hand. To date, we are unaware of any study that have at-tempted to measure the vertical distance between the volar cortexof the metacarpal bone and flexor tendon (D). Therefore, we areinterested in using Magnetic Resonance Imaging (MRI), as a radiolog-ical tool to provide further information on this.Methods: Retrospective radiological case study of all patients whohad MRI scan on the hand in Sheffield Teaching Hospitals NHS Foun-dation Trust between April 2004- April 2006 was conducted.For each case, all 4 metacarpal bones (excluding first metacarpalbone) were assessed on the eFilm software and D was measured atproximal 1/4, 1/2 and distal 1/4 positions of the MS.Key Results: 11 patients were included in study and a series of 44metacarpal bones was analysed based on axial T1 weighted MRIimages. Slice thickness for each scan varies from 1.0 mm to4.0 mm (mean 3.0 mm� 0.25).

Conclusions: MRI provides an accurate, non-invasive and non-radi-ating assessment of distance between volar cortex of MS and flexortendon. T1 weighted axial MRI images provide the most optimal an-atomical detail for these measurement.Significance: From our study, we would recommend that the distalend of any lag screw should not be more than 2 mm away from thevolar cortex any metacarpal bone in order to avoid injury to theflexor tendon.

O28. The length of the first metacarpal in the treatment oftriphalangeal thumbM. Zuidam a, E.E.C. Dees a, M.H. Lequin b, S.E.R. Hovius a

a Department of Plastic, Reconstructive, and Hand Surgery, TheNetherlandsb Department of Radiology, Erasmus Medical Centre, Dr.Molewaterplein 50, 3015 GE Rotterdam, The Netherlands

Aims: The Triphalangeal Thumb (TPT) is not a normal functioningthumb due to its extra length, altered stability and position andoften hypoplastic thenar musculature. Is the additional lengthonly caused by the extra phalanx or is the first metacarpal of influ-ence as well? Purpose of this study is to establish length of the firstmetacarpal in TPT for the different types of TPT and the possibleinfluence on corrective surgery.Methods: Hands of patients with TPT (N Z 59) were examined ontype of TPT (Delta, Trapezoid and Full type), position of growthplate and relative length of the first metacarpal. Ratios of the mea-surements were calculated (Second Metacarpal/First Metacarpal)and compared to a normal population.Results: First metacarpals in all three types of TPT were signifi-cantly longer compared to a normal population. Full and Trapezoidtype TPT are significantly longer then Delta type TPT. From the dif-ferent positions (distal, proximal and double) the distal growthplate (58%) was the most common.Conclusions: Positions of the growth plate vary in TPT more when com-pared with a normal population. All types of TPT have a longer firstmetacarpal. A reduction osteotomyof the firstmetacarpal can therefore

be a logical step to reduce thumb length in addition to either removal orarthrodesis with reduction at an interphalangeal joint.

O29. Myofibroblast differentiation and function is altered bydifferent levels of hypoxiaA. Modarressi a,b, B. Hinz B b, Pittet B a

a Division of Plastic and Reconstructive Surgery Department,University Hospitals of Geneva, Switzerlandb Laboratory of Cell Biophysics, Ecole Polytechnique Federale deLausanne, Switzerland

Background: Ischemia is a powerful promoter of inflammatory reac-tions and angiogenesis at the onset of dermal wound healing. How-ever, continuing low levels of tissue oxygenation (hypoxia) impairsphysiological healing, leading to the formation of chronic woundswith improper wound contraction. Key players in wound contrac-tion are myofibroblasts, specialized fibroblastic cells that exhibitenhanced contractile activity due to the expression of a-smoothmuscle actin (a-SMA).Aims: We elucidate how different levels of hypoxia influencedifferentiation, proliferation, survival and contractile function ofcultured subcutaneous myofibroblasts.Methods and material: Rat subcutaneous fibroblasts (SCF) were cul-tured for 5d in normoxic conditions (21% O2), in mild (5% O2) and insevere hypoxia (2.5% O2) with and without of myofibroblast-inducingTGFb1. Cell proliferation, myofibroblast differentiation and apopto-sis were evaluated by staining for DNA, a-SMA and caspase-3, respec-tively. Levels of total and active TGFb1 were measured with a TGFb1cell receptor system. Contraction was assessed by measuring diam-eter reduction of myofibroblast-populated attached collagen and bygrowing cells on deformable (wrinkling) silicone substrates.Results: Compared with normoxic conditions, cell proliferation in-creased significantly in hypoxia. Both, mild and severe hypoxia de-creased myofibroblast contraction, correlating with decreasedlevels of myofibroblast differentiation. All these effects were inde-pendent from the absence or presence of exogenous TGFb1. More-over, levels of active TGFb1 measured in the supernatant of cellscultured in 5% and 2.5% oxygen were higher compared with mediumconditioned in normoxia. However, expression levels of TGFb1 re-ceptor type II (TGFb1-RII) were down-regulated in hypoxia.Conclusion: Our results indicate that hypoxia is an important stim-ulus for subcutaneous fibroblast proliferation and autocrine pro-duction of active TGFb1. However, despite high levels of activeTGFb1 in hypoxia, 5% and 2.5% oxygen levels impair myofibroblastdifferentiation and contractile function. We are further exploringthe molecular mechanisms how hypoxia desensitizes fibroblasticcells against TGFb1 with the aim to develop new therapeutic strat-egies that target altered healing in hypoxic conditions.

O30. Collagen degradation in normal and pathological scarring:Aetiological implicationsC. Linge, C. Vigor, K. Rolfe, A. Al-Hilli, E. Clayton,J. RichardsonThe Raft Institute, Leopold Muller Building, Mount VernonHospital, Northwood, HA6 2RN, UK

Introduction: Apoptosis of fibroblasts is a key wound healing eventthat limits scar tissue production, failure of which may be the basisof scarring pathology. The trigger for this process is unclear butseems to involve contraction of fibrillar collagen. We have previouslyshown that unlike dermal and normal scar fibroblasts, pathologicalscar cells fail to undergo collagen-contraction-induced apoptosis.Aims:To investigate the roleofcollagen remodellingandmodificationin determining the ability of cells to undergo collagen-contraction-induced apoptosis.Methods: Scar fibroblast behaviour in 3D-collagen gels was exam-ined. Vital staining and TUNEL detected apoptosis Pharmacological

Total no of patient, N Z 11

Metacarpalbone

MedianMC (mm)

Mean D atproximal 0.25position (mm)

Mean Dat 0.5position(mm)

D at distal0.25 position(mm)

Second 63� 6 8� 1 13� 2 9� 2Third 62� 6 6� 1 10� 1 8� 2Fourth 55� 5 5� 2 10� 1 7� 2Fifth 50� 4 7� 3 12� 1 8� 2

S10 ECSAPS 2006 Abstracts