IL-17, IL-1beta and TNF-alpha stimulate VEGF production by dedifferentiated chondrocytes.
OBJECTIVE
To verify the involvement of proinflammatory cytokines IL-17, IL-1beta and tumor necrosis factor alpha (TNF-alpha) in cartilage vascularization by stimulating the production of vascular endothelial growth factor (VEGF) by chondrocytes isolated from patients with osteoarthritis (OA), in comparison with patients with rheumatoid arthritis (RA) and patients with femoral or humeral neck fracture (FP).


METHODS
Chondrocytes isolated from patients with OA were maintained in monolayer culture for several passages. Chondrocyte dedifferentiation was monitored by the synthesis of cathepsin B by these cells. Chondrocytes freshly isolated at each subculture (subcultures 1-3) were stimulated with IL-17, IL-1beta or TNF-alpha. Supernatants were collected, immunoassayed for the production of VEGF and cathepsin B and assayed as the source of VEGF on the VEGF sensible ECV304 cell line. The cells were used to quantify intracellular cathepsin B enzymatic activity.


RESULTS
In differentiated conditions IL-1beta and TNF-alpha, but not IL-17, can inhibit the spontaneous secretion of VEGF by human OA, RA and FP chondrocytes, and IL-17 can restore the decrease in VEGF secretion caused by TNF-alpha. IL-17, together with IL-1beta and TNF-alpha, can enhance VEGF secretion to various extents by dedifferentiated OA chondrocytes. This change in effect with respect to primary culture was observable for all cytokines at the beginning of dedifferentiation, when the production of VEGF by chondrocytes had dramatically fallen and the cathepsin B synthesis had increased. The amount of VEGF induced by cytokines on dedifferentiated chondrocytes never reached the amount of VEGF produced by differentiated chondrocytes. VEGF produced by chondrocytes stimulated the ECV304 cell line proliferation.


CONCLUSIONS
These results indicate that dedifferentiated OA chondrocytes secrete VEGF after stimulation with proinflammatory cytokines. This event may be responsible for neovascularization found in OA cartilage.
Honorati
Maria Cristina
MC
Laboratorio di Immunologia e Genetica, Istituto di Ricerca Codivilla Putti, Istituti Ortopedici Rizzoli, Via di Barbiano 1/10, 40136 Bologna, Italy. labimge@alma.unibo.it
Cattini
Luca
L
Facchini
Andrea
A
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001172:Arthritis, Rheumatoid / Q000276:immunology / Q000378:metabolism / Q000601:surgery; D019643:Arthroplasty, Replacement; D002401:Cathepsin B / Q000032:analysis / Q000096:biosynthesis; D002454:Cell Differentiation; D002478:Cells, Cultured; D019902:Chondrocytes / Q000276:immunology / Q000378:metabolism*; D016207:Cytokines / Q000494:pharmacology*; D005260:Female; D050723:Fractures, Bone / Q000276:immunology / Q000378:metabolism / Q000601:surgery; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D020381:Interleukin-17 / Q000494:pharmacology; D008297:Male; D008875:Middle Aged; D010003:Osteoarthritis / Q000276:immunology / Q000378:metabolism* / Q000601:surgery; D018709:Statistics, Nonparametric; D013268:Stimulation, Chemical; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D042461:Vascular Endothelial Growth Factor A / Q000032:analysis / Q000378:metabolism*
D016207:Cytokines; D007375:Interleukin-1; D020381:Interleukin-17; D014409:Tumor Necrosis Factor-alpha; D042461:Vascular Endothelial Growth Factor A; D002401:Cathepsin B