FLICE-inhibitory protein expression in synovial fibroblasts and at sites of cartilage and bone erosion in rheumatoid arthritis.
OBJECTIVE
Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by a hyperplastic synovial tissue, inflammatory infiltrates, and a progressive destruction of cartilage and bone. FLICE-inhibitory protein (FLIP) prevents the association of caspase 8 with FADD and thus exerts an antiapoptotic effect through inhibition of Fas-mediated apoptosis. We undertook this study to examine the expression of FLIP in RA, osteoarthritic (OA), and normal synovial tissues.


METHODS
We investigated the expression of FLIP (long form) in 5 RA, 2 OA, and 2 normal synovial tissue samples. A 393-bp fragment was amplified from complementary DNA obtained from cultured RA synovial fibroblasts (RASF) by reverse transcription-polymerase chain reaction (RT-PCR). Using in situ hybridization, the expression of FLIP messenger RNA (mRNA) in paraffin-embedded synovial tissue sections was investigated semiquantitatively by analyzing the lining layer, the sublining, and sites of invasion. Immunohistochemistry with anti-CD68 antibodies was performed on serial tissue sections to further characterize the cell types expressing FLIP. In addition, quantitative expression of FLIP was measured by real-time PCR.


RESULTS
RT-PCR revealed the expression of FLIP mRNA in all RA and OA samples tested. Using in situ hybridization in synovial tissue, FLIP was detected in all 5 RA samples and in 1 of 2 OA samples, but in neither of the 2 normal control samples. In RA, FLIP expression could be found in both the lining and sublining layers; most importantly, it could also be identified at sites of cartilage invasion and bone destruction. Moreover, quantitative PCR analysis showed 50% higher FLIP expression in RASF than in OASF.


CONCLUSIONS
The expression of antiapoptotic FLIP in RA synovial tissue and in synovial fibroblasts suggests the idea of a novel pathway in RA that potentially extends the lifespan of cartilage- and bone-degrading synovial cells, thus contributing to the progression of joint destruction.
Schedel
Jörg
J
Center of Experimental Rheumatology, WHO Collaborating Center for Molecular Biology, University Hospital of Zurich, Gloriastrasse 25, CH-8091 Zurich, Switzerland. joerg.schedel@klinik.uni-regensburg.de
Gay
Renate E
RE
Kuenzler
Peter
P
Seemayer
Christian
C
Simmen
Beat
B
Michel
Beat A
BA
Gay
Steffen
S
D001172:Arthritis, Rheumatoid / Q000473:pathology* / Q000503:physiopathology; D001842:Bone and Bones / Q000473:pathology; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; D002352:Carrier Proteins / Q000032:analysis / Q000235:genetics*; D002356:Cartilage / Q000473:pathology; D002478:Cells, Cultured; D005347:Fibroblasts / Q000737:chemistry / Q000473:pathology* / Q000502:physiology; D005786:Gene Expression Regulation / Q000502:physiology; D006801:Humans; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D047908:Intracellular Signaling Peptides and Proteins; D010003:Osteoarthritis / Q000473:pathology / Q000503:physiopathology; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis; D013583:Synovial Membrane / Q000473:pathology*
D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; C506386:CFLAR protein, human; D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D012333:RNA, Messenger