Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2016 Nov 18;18(1):270.
doi: 10.1186/s13075-016-1156-1.

Rheumatoid synovial fibroblasts differentiate into distinct subsets in the presence of cytokines and cartilage

Adam P Croft  1 , Amy J Naylor  1 , Jennifer L Marshall  1 , Debbie L Hardie  1 , Birgit Zimmermann  2 , Jason Turner  1 , Guillaume Desanti  1 , Holly Adams  1 , Adrian I Yemm  1 , Ulf Müller-Ladner  2 , Jean-Michel Dayer  3 , Elena Neumann  2 , Andrew Filer  1   4 , Christopher D Buckley  5   6
Affiliations

Rheumatoid synovial fibroblasts differentiate into distinct subsets in the presence of cytokines and cartilage

Adam P Croft et al. Arthritis Res Ther. .

Abstract

Background: We investigated two distinct synovial fibroblast populations that were located preferentially in the lining or sub-lining layers and defined by their expression of either podoplanin (PDPN) or CD248, and explored their ability to undergo self-assembly and transmigration in vivo.

Methods: Synovial fibroblasts (SF) were cultured in vitro and phenotypic changes following stimulation with interleukin (IL)-1β, tumor necrosis factor (TNF)-α, and transforming growth factor (TGF)-β1 were examined. To examine the phenotype of SF in vivo, a severe combined immunodeficiency (SCID) human-mouse model of cartilage destruction was utilised.

Results: SF in the lining layer in rheumatoid arthritis (RA) expressed high levels of PDPN compared to the normal synovium, whereas CD248 expression was restricted to sub-lining layer cells. TNF-α or IL1 stimulation in vitro resulted in an increased expression of PDPN. In contrast, stimulation with TGF-β1 induced CD248 expression. In the SCID human-mouse model, rheumatoid SF recapitulated the expression of PDPN and CD248. Fibroblasts adjacent to cartilage expressed PDPN, and attached to, invaded, and degraded cartilage. PDPN+ CD248- SF preceded the appearance of PDPN- CD248+ cells in contralateral implants.

Conclusions: We have identified two distinct SF populations identified by expression of either PDPN or CD248 which are located within different anatomical compartments of the inflamed synovial membrane. These markers discriminate between SF subsets with distinct biological properties. As PDPN-expressing cells are associated with early fibroblast migration and cartilage erosion in vivo, we propose that PDPN-expressing cells may be an attractive therapeutic target in RA.

PubMed Disclaimer

Figures

Fig. 1
Fig. 1
PDPN and CD248 markers identify specific fibroblast subsets localized to distinct anatomical compartments in the inflammatory synovium. a Representative confocal images of synovial biopsy tissue from healthy and rheumatoid arthritis (RA) synovium. Expression of podoplanin (PDPN) and CD248 is shown in healthy (normal) compared to rheumatoid synovium. b Quantification of marker expression in synovial tissue of patients with RA compared to normal controls. Data are expressed as median and interquartile range for CD248 (p = 0.65 by Mann-Whitney U test; n = 7 normal; n = 19 RA) and for PDPN (*p = 0.0013 by Mann-Whitney U test; n = 7 normal; n = 20 RA). c High magnification confocal images of rheumatoid synovium showing the lining (R1) versus sub-lining (R2) layer interface and expression of cell surface markers. Scale bars = 100 μm. PDI protein disulfide isomerase, UA unit area, VCAM-1 vascular adhesion molecule-1
Fig. 2
Fig. 2
Expression of CD248 and podoplanin (PDPN) in cultured RASF. Three representative confocal images of different RASF lines in culture are shown, ×40 magnification: green, CD248; blue, PDI; red PDPN; and grey, nuclei. PDI protein disulfide isomerase
Fig. 3
Fig. 3
Cytokines stimulate differential expression of CD248 and podoplanin (PDPN) in vitro. Expression of PDPN and CD248 by quantitative RT-PCR expressed as mRNA fold-change from unstimulated cells (mean ± SEM). **P < 0.01 by one-way ANOVA with Bonferroni post hoc analysis. IL interleukin, TGF transforming growth factor, TNF tumor necrosis factor
Fig. 4
Fig. 4
Cytokines stimulate differentiation of RASF towards specific fibroblast subsets in vitro. a Dual color flow cytometry of fibroblasts stimulated with either TNF-α or TGF-β in vitro. Histograms of CD248 versus podoplanin (PDPN) expression following cytokine stimulation. Percentage of positive cells in each gate is displayed. b The percentage of PDPN+ and CD248+ cells measured by flow cytometry in cultured RASF in response to cytokine stimulation (TNF-α, 10 ng/ml; IL-1β, 1 ng/ml; TGF-β1, 10 ng/ml). Data are expressed as mean ± SEM of the number of PDPN- and CD248-expressing fibroblasts expressed as a percentage of total cells at each time from N = 5 donor cell lines. Representative histograms of flow cytometric quantification of marker expression following cytokine stimulation are shown. Arrow indicates removal of cytokine. IL interleukin, TGF transforming growth factor, TNF tumor necrosis factor
Fig. 5
Fig. 5
In vivo cartilage destruction and vascular transmigration following implantation of RASF under the kidney capsule of SCID mice. a Representative confocal images of tissue sections of the fibro/cartilage matrix harvested 60 days following implantation showing expression of CD248 and podoplanin (PDPN). Dashed line shows demarcation of the cartilage interface. Scale bar = 100 μm. b The percentage of cells positive for each marker within 20 μm proximity to cartilage is displayed as the mean ± SEM for each cell marker. Time course analysis of the percentage of total nucleated cells expressing PDPN (c) and CD248 (d) within the primary implant compared to the secondary (cartilage only) implant at 6, 12, 18, 30, and 60 days following implantation. Absolute numbers of total nucleated cells over time in sections of secondary implant are also shown compared to the total number of those cells positive for each of these markers over time (e, f). Data are expressed as the mean ± SEM of the percentage of cells expressing each cell marker at each time point from eight tissue sections per implant (n = 3 mice). **P < 0.001 . C cartilage fragment
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Müller-Ladner U, Ospelt C, Gay S, Distler O, Pap T. Cells of the synovium in rheumatoid arthritis. Synovial fibroblasts. Arthritis Res Ther. 2007;9:223. doi: 10.1186/ar2337. - DOI - PMC - PubMed
    1. Müller-Ladner U, Gay S. MMPs and rheumatoid synovial fibroblasts: Siamese twins in joint destruction? Ann Rheum Dis. 2002;61:957–959. doi: 10.1136/ard.61.11.957. - DOI - PMC - PubMed
    1. Takayanagi H, Iizuka H, Juji T, Nakagawa T, Yamamoto A, Miyazaki T, et al. Involvement of receptor activator of nuclear factor kappaB ligand/osteoclast differentiation factor in osteoclastogenesis from synoviocytes in rheumatoid arthritis. Arthritis Rheum. 2000;43:259–269. doi: 10.1002/1529-0131(200002)43:2<259::AID-ANR4>3.0.CO;2-W. - DOI - PubMed
    1. Filer A. The fibroblast as a therapeutic target in rheumatoid arthritis. Curr Opin Pharmacol. 2013;13:413–419. doi: 10.1016/j.coph.2013.02.006. - DOI - PubMed
    1. Müller-Ladner U, Kriegsmann J, Franklin BN, Matsumoto S, Geiler T, Gay RE, et al. Synovial fibroblasts of patients with rheumatoid arthritis attach to and invade normal human cartilage when engrafted into SCID mice. Am J Pathol. 1996;149:1607–1615. - PMC - PubMed

Publication types

MeSH terms

-