Journal of Pathology J Pathol 2009; 217: 524–533 Published online 4 November 2008 in Wiley InterScience (www.interscience.wiley.com) DOI: 10.1002/path.2483 Original Paper An integrated proteomics approach for studying the molecular pathogenesis of Dupuytren’s disease Sandra Kraljevic Pavelic, 1,2 * Mirela Sedic, 1 Karlo Hock, 1 Srdan Vucinic, 1 Davor Jurisic, 3 Peter Gehrig, 4 Mike Scott, 4 Ralph Schlapbach, 4 Tamara Cacev, 1 Sanja Kapitanovic 1 and Kresimir Pavelic 1,2 * 1 Division of Molecular Medicine, Rudjer Boskovic Institute, Bijenicka Cesta 54, 10000 Zagreb, Croatia 2 Department of Biotechnology, University of Rijeka, 51000 Rijeka, Croatia 3 Clinical Hospital Rijeka, Department of Plastic and Reconstructive Surgery, 51000 Rijeka, Croatia 4 Functional Genomics Center Z ¨ urich, Winterthurerstrasse 190 (UNI Irchel), CH-8057 Z ¨ urich, Switzerland *Correspondence to: Professor Kresimir Pavelic, MD or Sandra Kraljevic Pavelic, PhD, Rudjer Boskovic Institute, Division of Molecular Medicine, Bijenicka Cesta 54, 10000 Zagreb, Croatia. E-mail: pavelic@irb.hr, skraljevic@irb.hr The authors declare that there are no financial/commercial conflicts of interest. Received: 11 June 2008 Revised: 29 September 2008 Accepted: 20 October 2008 Abstract Dupuytren’s disease (DD) is a fibromatosis characterized by non-malignant trans of palmar fascia leading to permanent contraction of one or more fingers. Despite the extensive knowledge of its clinical pathogenesis, the aetiology of thisdisease remains obscure. In the present paper,we report for the firstime on the proteomic profiling of diseased versus unaffected patient-matched palmar fasciae tissues from DD patients using two-dimensional gelelectrophoresis coupled with mass spectrometry analysis. The herein identified proteins were then used to create the protein– protein interactio (interactome). Such an integrated approach revealed the involvement of several different molecular processes related to DD progression, including extra- and intra-cellula oxidative stress, cytoskeletal changes, and alterations in cellular metabolism. In particular, autocrine regulation through ERBB-2 and IGF-1R receptors and the Akt signallin have emerged as novel components of pro-survival signalling in Dupuytren’s fibro thus might provide a basis for a new therapeutic strategy in Dupuytren’s disease. Copyright  2008 Pathological Society of GreatBritain and Ireland. Published by John Wiley & Sons, Ltd. Keywords: Akt; bioinformatics; cytoskeleton; Dupuytren’s disease; proteomics Introduction Dupuytren’s disease (DD) is characterized as a non- malignant transformation of connective tissue beneath the skin ofthe palm. It affects one or more digits, leading to irreversible contracture and loss of hand function [1].A characteristic of DD is abnormal proliferation of fibroblasts and theirdifferentiation into myofibroblasts. The latter generate a contractile forceas a consequence of smooth muscle α-actin overexpression. Recent research into fibrotic diseases highlighted the importance of anomalous expression of growth factors, eg tumour growth factors α and β (TGF-α, TGF-β), found to be elevated in DD patients as well [1].The altered expression of growth factors is accompanied by dense extracellular matrix production [2]. Reactive oxygen species (ROS) may also play a role in DD [3]. Currently, the standard treatment for DD is surgical therapy, which neither curesnor prevents frequent recurrence [2]. In the present study,we performed protein expres- sion profiling of diseased versus unaffected patient- matchedpalmarfasciaetissuesderivedfrom DD patientsby meansof two-dimensional gel elec- trophoresis (2-DE)followed by massspectrometry (MS) identification of theselectedproteinspots, as described previously [4]. Identified proteins were assigned biological contextby creating a protein – proteininteractions map (so-called‘interactome’). Such an integrated approach has disclosed possible new players responsible for DD progression. Materials and methods Clinical specimens Tissue samples from 12 patients (male Caucasians of European ethnicity, aged 59–77 years, who under- wentpartial fasciectomy as treatment of Dupuytren’s disease, diagnosed the last residual diseasephase) were obtained from the Clinical Hospital ‘Dubrava’, Zagreb, Croatia. Informed consent was obtained from all patients. The tissues were immediately exhaustivel washed in ice-cold isotonic buffer, frozen in liquid nitrogen, and stored at −80 ◦ C. Clinicalspecimens were collected in strict compliance with the clinic’s Copyright  2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. www.pathsoc.org.uk