Inhibition of fibroblast activation protein and dipeptidylpeptidase 4 increases cartilage invasion by rheumatoid arthritis synovial fibroblasts.
OBJECTIVE
Since fibroblasts in the synovium of patients with rheumatoid arthritis (RA) express the serine proteases fibroblast activation protein (FAP) and dipeptidylpeptidase 4 (DPP-4)/CD26, we undertook the current study to determine the functional role of both enzymes in the invasion of RA synovial fibroblasts (RASFs) into articular cartilage.


METHODS
Expression of FAP and DPP-4/CD26 by RASFs was analyzed using fluorescence-activated cell sorting and immunocytochemistry. Serine protease activity was measured by cleavage of fluorogenic substrates and inhibited upon treatment with L-glutamyl L-boroproline. The induction and expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in RASFs were detected using real-time polymerase chain reaction. Densitometric measurements of MMPs using immunoblotting confirmed our findings on the messenger RNA level. Stromal cell-derived factor 1 (SDF-1 [CXCL12]), MMP-1, and MMP-3 protein levels were measured using enzyme-linked immunosorbent assay. The impact of FAP and DPP-4/CD26 inhibition on the invasiveness of RASFs was analyzed in the SCID mouse coimplantation model of RA using immunohistochemistry.


RESULTS
Inhibition of serine protease activity of FAP and DPP-4/CD26 in vitro led to increased levels of SDF-1 in concert with MMP-1 and MMP-3, which are downstream effectors of SDF-1 signaling. Using the SCID mouse coimplantation model, inhibition of enzymatic activity in vivo significantly promoted invasion of xenotransplanted RASFs into cotransplanted human cartilage. Zones of cartilage resorption were infiltrated by FAP-expressing RASFs and marked by a significantly higher accumulation of MMP-1 and MMP-3, when compared with controls.


CONCLUSIONS
Our results indicate a central role for the serine protease activity of FAP and DPP-4/CD26 in protecting articular cartilage against invasion by synovial fibroblasts in RA.
University Hospital, Zurich, Switzerland.
Caroline
C
Ospelt
Joachim C
JC
Mertens
Astrid
A
Jüngel
Fabia
F
Brentano
Hanna
H
Maciejewska-Rodriguez
Lars C
LC
Huber
Hossein
H
Hemmatazad
Thomas
T
Wüest
Alexander
A
Knuth
Renate E
RE
Gay
Beat A
BA
Michel
Steffen
S
Gay
Christoph
C
Renner
Stefan
S
Bauer
D000818:Animals; D001172:Arthritis, Rheumatoid / Q000378:metabolism* / Q000473:pathology*; D002358:Cartilage, Articular / Q000378:metabolism / Q000473:pathology*; D002478:Cells, Cultured; D054377:Chemokine CXCL12 / Q000378:metabolism; D018819:Dipeptidyl Peptidase 4 / Q000378:metabolism*; D054873:Dipeptidyl-Peptidase IV Inhibitors; D004195:Disease Models, Animal; D010450:Endopeptidases; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects / Q000201:enzymology / Q000473:pathology; D018093:Gelatinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006801:Humans; D020782:Matrix Metalloproteinases / Q000378:metabolism; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051379:Mice; D016513:Mice, SCID; D012697:Serine Endopeptidases / Q000378:metabolism*; D013583:Synovial Membrane / Q000201:enzymology / Q000473:pathology; D000099167:Fibroblast Activation Protein Alpha
D054377:Chemokine CXCL12; D018819:Dipeptidyl Peptidase 4; D054873:Dipeptidyl-Peptidase IV Inhibitors; D010450:Endopeptidases; D004791:Enzyme Inhibitors; D018093:Gelatinases; D020782:Matrix Metalloproteinases; D008565:Membrane Proteins; D012697:Serine Endopeptidases; D000099167:Fibroblast Activation Protein Alpha; C515255:CXCL12 protein, human; C515256:Cxcl12 protein, mouse; C042807:DPP4 protein, human; C545245:Dpp4 protein, mouse