Effects of proinflammatory cytokines on canine articular chondrocytes in a three-dimensional culture.
OBJECTIVE
To determine the effects of interleukin (IL)-1 and tumor necrosis factor (TNF)-alpha on canine chondrocytes cultured in an agarose-based 3-dimensional (3-D) system.


METHODS
Humeral head articular cartilage chondrocytes obtained from 6 adult dogs.


METHODS
Chondrocytes were cultured in a 3-D system for < or = 12 days in serum-free medium with IL 1alpha, IL-1beta, or TNF-alpha at concentrations of 20, 50, or 100 ng/mL. After 1, 3, 6, and 12 days, glycosaminoglycan (GAG) concentrations in 3-D constructs; nitric oxide and prostaglandin E2 (PGE2) concentrations in media samples; and relative expressions of selected genes, including metalloproteinase (MMP)-13 and tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2, were evaluated. Control specimens were comprised of chondrocytes cultured without proinflammatory cytokines.


RESULTS
In control 3-D constructs, GAG content was significantly higher than for all other constructs. Compared with control values, relative expressions of MMP-13, TIMP-1, and TIMP-2 genes in the IL-1beta (50 ng/mL) group were significantly higher at day 1; at all evaluations, media concentrations of nitric oxide were significantly higher in all TNF-alpha-treated cultures; and concentrations of PGE2 in media samples were significantly higher in the IL-1beta (50 ng/mL) and IL-1beta (100 ng/mL) groups at days 1 and 3, in the IL-1beta (100 ng/mL) group at day 6, and in all TNF-alpha groups at days 1, 3, and 6.


CONCLUSIONS
Results suggested that TNF-alpha more readily induces production of nitric oxide and PGE2 by canine chondrocytes, compared with IL-1beta. In vitro, IL-1alpha appeared to have a minimal effect on canine chondrocytes.
Kuroki
Keiichi
K
Comparative Orthopaedic Laboratory, University of Missouri, Columbia, MO 65211, USA.
Stoker
Aaron M
AM
Cook
James L
JL
D053674:Aggrecans; D000818:Animals; D002358:Cartilage, Articular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D018929:Cell Culture Techniques; D019902:Chondrocytes / Q000187:drug effects / Q000378:metabolism*; D024043:Collagen Type II / Q000378:metabolism; D017364:Collagenases / Q000378:metabolism; D004285:Dogs / Q000378:metabolism*; D005109:Extracellular Matrix / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D007375:Interleukin-1 / Q000494:pharmacology / Q000502:physiology*; D037181:Lectins, C-Type; D020781:Matrix Metalloproteinase 1 / Q000378:metabolism; D053509:Matrix Metalloproteinase 13; D011509:Proteoglycans / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D013997:Time Factors; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000378:metabolism; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology / Q000502:physiology*
D053674:Aggrecans; D024043:Collagen Type II; D016326:Extracellular Matrix Proteins; D007375:Interleukin-1; D037181:Lectins, C-Type; D011509:Proteoglycans; D011994:Recombinant Proteins; D019715:Tissue Inhibitor of Metalloproteinase-1; D014409:Tumor Necrosis Factor-alpha; D019716:Tissue Inhibitor of Metalloproteinase-2; D017364:Collagenases; D053509:Matrix Metalloproteinase 13; D020781:Matrix Metalloproteinase 1