Expression and Localization of Cathepsins B, D, and G in Dupuytren’s Disease

Background: The pathogenesis of Dupuytren's disease (DD) remains unclear. An embryonic stem cell (ESC)–like population in the endothelium of the microvessels around tissues that expresses components of the renin-angiotensin system (RAS) has been reported. This study investigated if this primitive population expresses cathepsins B, D, and G, that contribute to RAS bypass loops. Methods: 3,3-Diaminobenzidine immunohistochemical (IHC) staining for cathepsins B, D, and G was performed on sections of formalin-fixed paraffin-embedded DD cords (n = 10) and nodules (n = 10). Immunofluorescence IHC staining was utilized to demonstrate co-expression of these cathepsins with ESC markers. Protein and gene expression of these cathepsins was investigated in snap-frozen DD cords (n = 3) and nodules (n = 3) by Western blotting and NanoString analysis, respectively. Enzymatic activity of these cathepsins was investigated by enzymatic activity assays. Results: 3,3-Diaminobenzidine IHC staining demonstrated expression of cathepsins B, D, and G in DD cords and nodules. Gene expression of cathepsins B, D, and G was confirmed by NanoString analysis. Western blotting confirmed expression of cathepsins B and D, but not cathepsin G. Immunofluorescent IHC staining demonstrated high abundance of cathepsins B and D on the OCT4+/angiotensin converting enzyme+ endothelium and the smooth muscle layer of the microvessels. Cathepsin G was localized to trypase+ cells within the stroma in DD cords and nodules with limited expression on the microvessels. Enzyme activity assays demonstrated functional activity of cathepsins B and D. Conclusions: Cathepsins B, D, and G were expressed in the DD tissues, with cathepsins B and D localized to the primitive population in the endothelium of the microvessels, whereas cathepsin G was localized to phenotypic mast cells, suggesting the presence of bypass loops for the RAS. This is an open-access article distributed under the terms of the Creative Commons Attribution-Non Commercial-No Derivatives License 4.0 (CCBY-NC-ND), where it is permissible to download and share the work provided it is properly cited. The work cannot be changed in any way or used commercially without permission from the journal. Published online 14 February 2018. Received for publication August 31, 2017; accepted January 5, 2018. Presented at the New Zealand Association for Plastic Surgeons' and the Australian and New Zealand Society of Oculoplastic Surgeon's Joint Annual Scientific Meeting, Queenstown, New Zealand August 4–6, 2017. Disclosure: Drs. Davis, Tan, and Itinteang are inventors of a patent application Treatment of Fibrotic Conditions (no. PCT/NZ2016/050187). The authors have no financial interest to declare in relation to the content of this article. The Article Processing Charge was paid for by the Gillies McIndoe Research Institute general fund. Supplemental digital content is available for this article. Clickable URL citations appear in the text. Drs. Tan and Itinteang contributed equally to this work. Swee T. Tan, ONZM, MBBS, PhD, FRACS, Gillies McIndoe Research Institute, PO Box 7184, Newtown 6242, Wellington, New Zealand, E-mail: swee.tan@gmri.org.nz Copyright © 2018 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of the American Society of Plastic Surgeons. All rights reserved.

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