Intra-articular injection of tumor necrosis factor-alpha in the rat: an acute and reversible in vivo model of cartilage proteoglycan degradation.
OBJECTIVE
To develop an in vivo model for rapid assessment of cartilage aggrecan degradation and its pharmacological modulation.


METHODS
Tumor necrosis factor-alpha (TNFalpha) was injected intra-articularly (IA) in rat knees and aggrecan degradation was monitored at various times following challenge. Articular cartilage was assessed for aggrecan content by Safranin O staining and by immunohistochemistry for the NITEGE epitope. Synovial fluids (SFs) were analyzed for sulfated glycosaminoglycans (GAGs) using the dimethylmethylene blue dye assay and for aggrecan fragments generated by specific cleavage at aggrecanase-sensitive sites by Western blot analysis with neoepitope antibodies. Indomethacin, dexamethasone, and an aggrecanase inhibitor were evaluated for their ability to modulate TNFalpha-induced proteoglycan degradation in vivo.


RESULTS
(1) IA injection of TNFalpha in the knee joint of rats resulted in transient aggrecan degradation and release of aggrecanase-generated aggrecan fragments from the articular cartilage into the SF; (2) a correlation was observed between histologically assessed depletion of aggrecan from the articular cartilage and the appearance of specific neoepitopes in the SF; (3) aggrecan degradation was inhibited by an aggrecanase inhibitor as well as by dexamethasone, but not by the non-steroidal anti-inflammatory drug (NSAID), indomethacin.


CONCLUSIONS
TNFalpha injection in the knee joints of rats results in rapid transient cartilage proteoglycan degradation, mediated by cleavage at the aggrecanase sites. Biomarker read-out of specific neoepitopes in the SF enables the use of this mechanism-based model for rapid evaluation of aggrecanase-mediated aggrecan degradation in vivo.
Malfait
A M
AM
Pfizer Global Research and Development, St Louis Laboratories, St Louis, MO 63017, USA. anne-marie.malfait@pfizer.com
Tortorella
M
M
Thompson
J
J
Hills
R
R
Meyer
D M
DM
Jaffee
B D
BD
Chinn
K
K
Ghoreishi-Haack
N
N
Markosyan
S
S
Arner
E C
EC
D053674:Aggrecans / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D001169:Arthritis, Experimental / Q000188:drug therapy / Q000473:pathology*; D015153:Blotting, Western; D002358:Cartilage, Articular / Q000187:drug effects / Q000473:pathology*; D007150:Immunohistochemistry; D007270:Injections, Intra-Articular; D007719:Knee Joint / Q000187:drug effects / Q000473:pathology*; D008297:Male; D010003:Osteoarthritis / Q000188:drug therapy / Q000473:pathology*; D011509:Proteoglycans / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism* / Q000494:pharmacology
D053674:Aggrecans; D011509:Proteoglycans; D014409:Tumor Necrosis Factor-alpha