Microarray Analysis of Dupuytren’s Disease Cells: The Profibrogenic Role of the TGF-� Inducible p38 MAPK Pathway

Abstract

Background: Dupuytren’s disease (DD) is a nodular palmar fibromatosis that causes irreversible permanent contracture of fingers and results in the loss of hand function. Surgery still remains the only available solution for DD patients but cannot permanently cure the disease nor reduce high recurrence rates. With this rationale, we designed a study aimed at an improved understanding of the molecular mechanisms underlying DD. Our major focus was an analysis of the global gene expression profile and signalling pathways in DD cells with the aim of identifying novel biomarkers and/or therapeutic targets. Methods: Primary cells were cultured from surgically removed diseased and healthy tissue. Microarray expression analysis (HG-U133A array, Affymetrix) and qPCR was performed with total RNA isolated from primary DD cells. Mechanistic studies involving inhibition of p38 phosphorylation were performed on normal human fibroblasts’ and primary DD cells’ in vitro models. Expression of stem cell markers in primary fibroblasts/myofibroblasts was assessed as well. Results: We identified 3 p38MAPK signalling pathway regulatory genes, THBS1, GADD45α and NUAK1, all involved in cellular proliferation and production of the extracellular matrix proteins. Inhibition of the p38MAPK signalling pathway induced down-regulation of myofibroblast markers, α-smooth muscle actin and palladin. A stem-cell like subpopulation positive for CD90 marker was identified among primary DD cells. Conclusion: The study reveals involvement of the p38 MAPK pathway as a possible signalling cascade in the pathogenesis of Dupuytren’s disease. Moreover, a particular stem cell-like CD90⁺ subpopulation was identified that might contribute to DD development.

Identification of Histological Patterns in Clinically Affected and Unaffected Palm Regions in Dupuytren's Disease

“…This study confirmed that DDC tissues had increased extracellular matrix (ECM) deposition as compared to NPF and NFF, as previously suggested by Rehman and cols [30] and by Ratkaj and cols [28]. In this sense, one of the most important ECM components is the fibrillar component, which typically becomes very abundant in Dupuytren's disease [11], [21], [31], and a major biochemical abnormality found in Dupuytren's tissue is an increase in total collagen associated with an increase in the ratio of type III to type I collagen [21].…”

“…Moreover, collagens types V and XIV play a role in regulating fibrillogenesis by controlling the initiation of collagen fibril assembly [35], [36]. The significant alteration of both collagens that we found in DDC tissues points to the idea that the complex process of collagen fibrils formation is disregulated in Dupuytren's disease, as previously suggested [28], [30]. Beside this, our analysis of non-fibrillar collagens such as collagens types IV, VI, VII, VIII and X revealed a significant alteration of these collagens as well, suggesting that the diffuse tissue collagen network of normal connective tissues could also be altered in DDC and NPF.…”

“…These results are in agreement with our idea that NPF cells could be pathological, although at lower extent than diseased cells of the fibrous cord, at least at this stage. Previous reports already demonstrated that cells cultured from tissues affected by Dupuytren's disease may have higher proliferation rate than control tissues [25], [26], [27], and several authors found a significant overexpression of several genes involved in cell proliferation in these cells [28]. From a translational standpoint, these results suggest that tissues that are apparently unaffected by DD such as NPF could be also affected by the disease.…”

TRPA1 is required for TGF-β signaling and its loss blocks inflammatory fibrosis in mouse corneal stroma

“…This change in gene expression patterns may be mediated by alterations of TGF- β 1-dependent signaling cascades; fibrogenic gene expression and myofibroblast conversion in fibroblasts are reportedly mediated by Smad and p38 MAPK cascades activated by TGF- β 1. 9,33,35–37 The expression of TRPA1 is upregulated in the cells with exposure to TGF- β 1, suggesting the presence of positive feedback through a TGF- β 1-induced activation of signaling cascades mediated by TRPA1. A previous report indicated that the TRPA1 receptor participates in tissue inflammation.…”

Genome-Wide Analysis Using Exon Arrays Demonstrates an Important Role for Expression of Extra-Cellular Matrix, Fibrotic Control and Tissue Remodelling Genes in Dupuytren's Disease

“…Few previous studies have compared expression levels in primary fibroblasts derived from the affected and unaffected regions of DD patients [49] or from the two patient types [37], [43]. One compared the tissue between DD patients and hand trauma patients [30].…”