Transition of healthy to diseased synovial tissue in rheumatoid arthritis is associated with gain of mesenchymal/fibrotic characteristics

Marjan Mc Steenvoorden,Clemens Löwik,Gabri Van Der Pluijm,Adriana C Gittenberger-Degroot,Cornelis Pj Visser,Tom Wj Huizinga,Marco C Deruiter,Bert J Wisse,Jeroen Degroot,René Em Toes,Tanja Ca Tolboom

doi:10.1186/ar2073

Marjan Mc Steenvoorden, Clemens Löwik + Show 9 more

Open Access

https://doi.org/10.1186/ar2073

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Abstract

The healthy synovial lining layer consists of a single cell layer that regulates the transport between the joint cavity and the surrounding tissue. It has been suggested that abnormalities such as somatic mutations in the p53 tumor-suppressor gene contribute to synovial hyperplasia and invasion in rheumatoid arthritis (RA). In this study, expression of epithelial markers on healthy and diseased synovial lining tissue was examined. In addition, we investigated whether a regulated process, resembling epithelial to mesenchymal transition (EMT)/fibrosis, could be responsible for the altered phenotype of the synovial lining layer in RA. Synovial tissue from healthy subjects and RA patients was obtained during arthroscopy. To detect signs of EMT, expression of E-cadherin (epithelial marker), collagen type IV (indicator of the presence of a basement membrane) and α-smooth muscle actin (α-sma; a myofibroblast marker) was investigated on frozen tissue sections using immunohistochemistry. Fibroblast-like synoviocytes (FLSs) from healthy subjects were isolated and subjected to stimulation with synovial fluid (SF) from two RA patients and to transforming growth factor (TGF)-β. To detect whether EMT/fibrotic markers were increased, expression of collagen type I, α-sma and telopeptide lysylhydroxylase (TLH) was measured by real time PCR. Expression of E-cadherin and collagen type IV was found in healthy and arthritic synovial tissue. Expression of α-sma was only found in the synovial lining layer of RA patients. Stimulation of healthy FLSs with SF resulted in an upregulation of α-sma and TLH mRNA. Collagen type I and TLH mRNA were upregulated after stimulation with TGF-β. Addition of bone morphogenetic protein (BMP)-7 to healthy FLS stimulated with SF inhibited the expression of α-sma mRNA. The finding that E-cadherin and collagen type IV are expressed in the lining layer of healthy and arthritic synovium indicates that these lining cells display an epithelial-like phenotype. In addition, the presence of α-sma in the synovial lining layer of RA patients and induction of fibrotic markers in healthy FLSs by SF from RA patients indicate that a regulated process comparable to EMT might cause the alteration in phenotype of RA FLSs. Therefore, BMP-7 may represent a promising agent to counteract the transition imposed on synoviocytes in the RA joint.

Highlights

The synovial lining has been described as a mesenchymal tissue because it lacks several epithelial properties, such as tight junctions and desmosomes [1], its function and morphology resemble that of epithelial tissues
Expression of collagen type IV mRNA was found in cultured fibroblast-like synoviocytes (FLSs) from both rheumatoid arthritis (RA) patients and healthy controls (Figure 3i), indicating that FLSs are able to produce collagen type IV
Arrows indicate collagen IV, CD55 or double staining in the lining layer. (i) PCR results: top, collagen type IV mRNA expression in fibroblast-like synoviocytes from RA patients and healthy controls; bottom, β2microglobulin (β2M) mRNA expression in the same samples

Summary

Introduction

The synovial lining has been described as a mesenchymal tissue because it lacks several epithelial properties, such as tight junctions and desmosomes [1], its function and morphology resemble that of epithelial tissues. The β2M = β2-microglobulin; AGE = advanced glycation endproducts; BCIP = 5-bromo-4-chloro-3-indolyl-phosphatase; BMP = bone morphogenetic protein; ED-A = extra domain A; EMT = epithelial to mesenchymal transition; FLS = fibroblast-like synoviocyte; HRP = horse radish peroxidase; NBT = 4-nitro blue tetrazolium chloride; PBS = phosphate-buffered saline; RA = rheumatoid arthritis; RAGE = receptor for advanced glycation endproducts; SF = synovial fluid; sma = smooth muscle actin; TGF = transforming growth factor; TLH = telopeptide lysylhydroxylase. Function of epithelium is to form a barrier between the external and internal environment and to regulate transport between the cavity it encloses and the adjacent tissue by facilitating transport and secretion [2]. In rheumatoid arthritis (RA) synovial hyperplasia and inflammation play a prominent role. While influx of inflammatory cells such as macrophages are important in the inflammation of the tissue [4], proliferation of fibroblast-like synoviocytes (FLSs) seems to be a major cause of the hyperplasia of the synovial tissue [5,6]. The data indicate that FLSs could play a role in cartilage degradation, as they are found at sites of cartilage degradation in RA and are able to degrade and invade cartilage when co-implanted into a SCID mouse [7]

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