Invasive properties of fibroblast-like synoviocytes: correlation with growth characteristics and expression of MMP-1, MMP-3, and MMP-10

Abstract

Background: Matrix metalloproteinases (MMPs) have a pivotal role in the destruction of cartilage in rheumatoid arthritis (RA), which is mediated by the fibroblast-like synoviocytes (FLS). Objective: To examine the in vitro invasiveness of synoviocytes obtained from inflamed joints of patients with arthritis in relation to the expression of MMP 1-14, 17, 19, cathepsin-K, the tissue inhibitors of matrix metalloproteinases TIMP-1 and TIMP-2 by FLS. Methods: FLS were derived from 56 patients (30 with RA, 17 with osteoarthritis (OA), and nine with avascular necrosis (AVN)). Invasive growth of FLS through an artificial matrix (Matrigel) was measured in a transwell system. The number of cells that migrated through the matrix were counted. Proliferation rate was determined by counting the FLS after seven days of culturing. Expression of MMPs, cathepsin-K and TIMPs was investigated with reverse transcriptase-polymerase chain reaction and related to the expression of a household gene, β-actin. Results: FLS from RA showed greater invasive growth than FLS from OA and AVN. The median number of cells that grew through the matrix membrane was 4788 for RA, significantly higher than the number for OA, 1875 (p<0.001) and for AVN, 1530 (p=0.014). The median rate of proliferation of RA FLS was 0.27 per day compared with OA 0.22 per day (p= 0.012) and AVN 0.25 per day, but there was no correlation between the rate of proliferation and invasive growth in vitro. FLS from RA and OA that expressed MMP-1, MMP-3, or MMP-10 were significantly more invasive (median number of invasive cells: 3835, 4248, 4990, respectively) than cells that did not express these MMPs (1605, p=0.03; 1970, p=0.004; 2360, p=0.012, respectively). There was also a significant relationship between the expression of MMP-1 and MMP-9 and the diagnosis RA (both p=0.013). The expression levels of mRNA for MMP-1 and MMP-2 correlated with the protein levels produced by the synoviocytes as measured by an enzyme linked immunosorbent assay (ELISA). Conclusion: FLS of RA invade more aggressively in a Matrigel matrix than OA and AVN FLS; this is not because of a higher rate of proliferation of RA FLS. The significant correlation between the expression of MMP-1, MMP-3, and MMP-10 and invasive growth in a Matrigel transwell system suggests that these MMPs play a part in the invasive growth of FLS obtained from patients with RA.

Invasiveness of fibroblast‐like synoviocytes is an individual patient characteristic associated with the rate of joint destruction in patients with rheumatoid arthritis

“…RA FLS were significantly more invasive than OA FLS in this study (mean Ϯ SD 2,884 Ϯ 2,326 and 4,573 Ϯ 2,502 for OA and RA FLS, respectively; P Ͻ 0.001) (Figure 1). These results are consistent with those of previous studies (1,4).…”

“…Invasiveness of FLS was measured as previously described (4). Briefly, transwells (6.5 mm diameter, 8.0 m pore width; Costar, Cambridge, MA) were coated with paraffin to prevent meniscus formation.…”

Association of the –2849 interleukin‐10 promoter polymorphism with autoantibody production and joint destruction in rheumatoid arthritis

“…Synovial tissue was obtained from RA and OA patients at the time of joint replacement or synovectomy. From the synovial tissue, FLS were isolated and their invasiveness was tested in vitro, using a Matrigel transwell system as previously described (19). Briefly, cells were separated from tissue by collagenase treatment and grown to confluence in Iscove's modified Dulbecco's medium (IMDM; BioWhittaker, Verviers, Belgium).…”

Altered expression of MicroRNA in synovial fibroblasts and synovial tissue in rheumatoid arthritis

“…Considering the stimulatory effects of TNF␣ and IL-1␤ on the expression of miR-155 and the high abundance of these cytokines in rheumatoid joints, we analyzed the expression of miR-155 in the RA synovium. The samples of synovial tissue from RA patients proved to be highly enriched in miR-155, with levels of expression being 8-fold higher than those found in noninflamed synovia from patients with OA (P Ͻ 0.02) ( Figure 1B In this context, we chose to investigate the expression of MMPs 1, 3, 9, and 13 and the cytokine IL-6 as markers of the destructive and inflammatory properties of RASFs (27,28). We found that enforced expression of miR-155 in RASFs resulted in a substantial reduction in the constitutive expression of MMP-3 as compared with that in cells transfected with scrambled control.…”