Vascular endothelial growth factor (VEGF) induces matrix metalloproteinase expression in immortalized chondrocytes.
VEGF (vascular endothelial growth factor), an important angiogenesis factor, appears also to be involved in inflammatory processes. Recent studies have shown that VEGF and its receptors (VEGFR) are expressed on osteoarthritic, but not on normal adult, chondrocytes. To elucidate possible functions of VEGF in osteoarthritic cartilage, the effects of VEGF were studied on immortalized human chondrocytes. Activated matrix metalloproteinase (MMP)-1, MMP-3, MMP-13, tissue inhibitor of metalloproteinases (TIMP)-1, TIMP-2, interleukin (IL)-1beta, IL-6, and tumour necrosis factor-alpha (TNF-alpha) were measured in culture supernatants by enzyme-linked immunosorbent assays, nitric oxide with the Griess reagent, and cell proliferation by [3H]thymidine incorporation. VEGFR-2 mRNA was quantified by real-time reverse transcription-polymerase chain reaction and the protein was identified by immuno-gold electron microscopy. Intracellular signal transduction effects were determined by western blots and electrophoretic mobility shift assays. The chondrocyte cell lines C28/I2, C20/A4, and T/C28a2/a4 expressed functionally active VEGFR-2. VEGF stimulation induced receptor phosphorylation, activation of the mitogen-activated protein kinases ERK 1/2, and long-lasting activation of the transcription factor AP-1 (activator protein-1). VEGF increased secreted MMP-1, MMP-3, and especially MMP-13, which could be effectively reduced by an inhibitor of VEGFR-2 kinase activity. Interestingly, VEGF diminished the expression of TIMP-1 and especially TIMP-2. Under hypoxic conditions, as occur in cartilage, the reduction in TIMP levels was even greater. Furthermore, VEGF induced IL-1beta, IL-6, TNF-alpha, and nitric oxide expression to a small extent and stimulated the proliferation of immortalized chondrocytes. These findings indicate that VEGF is an autocrine stimulator of immortalized chondrocytes that mediates mainly destructive processes in osteoarthritis.
Pufe
Thomas
T
Department of Anatomy, University of Kiel, D-24098 Kiel, Germany.
Harde
Viola
V
Petersen
Wolf
W
Goldring
Mary B
MB
Tillmann
Bernhard
B
Mentlein
Rolf
R
D019898:Autocrine Communication / Q000502:physiology*; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D019902:Chondrocytes / Q000187:drug effects / Q000201:enzymology* / Q000276:immunology; D016207:Cytokines / Q000032:analysis; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D020782:Matrix Metalloproteinases / Q000378:metabolism*; D016253:Microscopy, Immunoelectron; D009569:Nitric Oxide / Q000032:analysis; D010003:Osteoarthritis / Q000201:enzymology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*; D013268:Stimulation, Chemical; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000032:analysis; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000032:analysis; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000032:analysis
D016207:Cytokines; D019715:Tissue Inhibitor of Metalloproteinase-1; D042461:Vascular Endothelial Growth Factor A; D019716:Tissue Inhibitor of Metalloproteinase-2; D009569:Nitric Oxide; D040301:Vascular Endothelial Growth Factor Receptor-2; D020782:Matrix Metalloproteinases