Isolation and characterization of rheumatoid arthritis synovial fibroblasts from primary culture--primary culture cells markedly differ from fourth-passage cells.
To reduce culture artifacts by conventional repeated passaging and long-term culture in vitro, the isolation of synovial fibroblasts (SFB) was attempted from rheumatoid arthritis (RA) synovial membranes by trypsin/collagenase digest, short-term in vitro adherence (7 days), and negative isolation using magnetobead-coupled anti-CD14 monoclonal antibodies. This method yielded highly enriched SFB (85% prolyl-4-hydroxylase+/74% Thy-1/CD90+ cells; <2% contaminating macrophages; <1% leukocytes/endothelial cells) that, in comparison with conventional fourth-passage RA-SFB, showed a markedly different phenotype and significantly lower proliferation rates upon stimulation with platelet-derived growth factor and IL-1beta. This isolation method is simple and reliable, and may yield cells with features closer to the in vivo configuration of RA-SFB by avoiding extended in vitro culture.
Zimmermann
T
T
Experimental Rheumatology Unit, Friedrich Schiller University Jena, Jena, Germany.
Kunisch
E
E
Pfeiffer
R
R
Hirth
A
A
Stahl
H D
HD
Sack
U
U
Laube
A
A
Liesaus
E
E
Roth
A
A
Palombo-Kinne
E
E
Emmrich
F
F
Kinne
R W
RW
D001172:Arthritis, Rheumatoid / Q000473:pathology*; D018929:Cell Culture Techniques / Q000379:methods*; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D005347:Fibroblasts / Q000473:pathology*; D006801:Humans; D010003:Osteoarthritis / Q000473:pathology; D013583:Synovial Membrane / Q000473:pathology*