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- Author or Editor: Pooi-See Chan x
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Abstract
Objective—To determine effects of glucosamine (GLN) and chondroitin sulfate (CS) on expression of genes encoding putative mediators of osteoarthritis in bovine cartilage explants cultured for 2 weeks.
Sample Population—Articular cartilage explants harvested from carpal joints of 4 Holstein steers after slaughter.
Procedures—Cartilage disks were treated as follows: fetal bovine serum only (control treatment), human recombinant interleukin (IL)-1β (50 ng/mL; IL-1 treatment), GLN (5 μg/mL) with addition of CS (20 μg/mL; GLN-CS treatment), and human recombinant IL-1β (50 ng/mL) with addition of GLN and CS (IL-1–GLN-CS treatment). Media were analyzed for nitric oxide and prostaglandin E2 (PGE2) release. Explants were subjected to quantitative real-time PCR analysis; expressions of mRNA for inducible nitric oxide synthase, cyclooxygenase-2, microsomal prostaglandin E synthase 1, matrix metalloproteinase (MMP)-3 and -13, aggrecanase-1 and -2, tissue inhibitor of metalloproteinase (TIMP)-3, type II collagen, and aggrecan were assessed.
Results—IL-1–GLN-CS and GLN-CS treatments decreased nitrite release, compared with IL-1 treatment; IL-1–GLN-CS treatment decreased IL-1–induced PGE2 release. Expressions of inducible nitric oxide synthase, cyclooxygenase-2, and microsomal prostaglandin E synthase 1 mRNA were abrogated by GLN-CS and IL-1–GLN-CS treatments. Interleukin-1–induced mRNA expressions of proteolytic enzymes were diminished by IL-1–GLN-CS treatment. Compared with control treatment, GLN-CS treatment decreased MMP-3 and aggrecanase-2 mRNA expression. Transcripts of TIMP-3 were increased by IL-1–GLN-CS treatment, compared with IL-1 treatment. Genes encoding type II collagen and aggrecan on day 14 were upregulated by GLN-CS and IL-1–GLN-CS treatments, compared with control treatment.
Conclusions and Clinical Relevance—Treatment with GLN and CS consistently downregulated mRNA expression for inflammatory mediators and matrix degrading enzymes while increasing TIMP-3 transcripts.
Abstract
Objective—To determine the effects of glucosamine (GLN) and chondroitin sulfate (CS), at concentrations attainable in vivo, on expression of genes encoding proteolytic enzymes, enzyme inhibitors, and macromolecules of articular cartilage in interleukin-1(IL- 1)–challenged bovine cartilage explants.
Sample Population—Articular cartilage explants harvested from 9 steers.
Procedures—Cartilage explants were exposed to media containing 10% fetal bovine serum (FBS) only, IL- 1 (50 ng/mL), IL-1 with GLN (5 µg/mL), IL-1 with CS (20 µg/mL), or IL-1 with GLN and CS for 24 and 48 hours. Cartilage was frozen, and RNA was extracted. Gene expression of matrix metalloproteinases (MMPs)-2, -3, -9, -13, and -14; aggrecanases (Aggs)-1 and -2; tissue inhibitors of metalloproteinases (TIMPs)-1, -2, and -3; and type II collagen and aggrecan were assessed with quantitative real-time polymerase chain reaction.
Results—Upregulated MMP-3, MMP-13, and Agg-1 transcripts at 24 hours were repressed by the GLN and CS combination by at least approximately 6-fold. Glucosamine was effective in suppressing IL-1–induced mRNA expression of MMP-13, Agg-1, and Agg-2, whereas CS was effective in decreasing IL-1–induced MMP-13 transcript at 24 hours. At 48 hours, GLN and CS added separately and in combination significantly abrogated Agg-1 and Agg-2 gene induction. The combination also decreased IL-1–stimulated MMP-13 transcript.
Conclusions and Clinical Relevance—GLN and CS, at concentrations that are within the range measured in synovial fluid and blood after oral administration, may regulate expression of matrix degrading enzymes and their inhibitors at the transcriptional level, providing a plausible mechanism for their purported chondroprotective properties. (Am J Vet Res 2005;66:1870–1876)
Abstract
Objective—To determine whether glucosamine and chondroitin sulfate (CS) at concentrations approximating those achieved in plasma by oral administration would influence gene expression of selected mediators of osteoarthritis in cytokine-stimulated equine articular chondrocytes.
Sample Population—Samples of grossly normal articular cartilage obtained from the metacarpophalangeal joint of 13 horses.
Procedure—Equine chondrocytes in pellet culture were stimulated with a subsaturating dose of recombinant equine interleukin (reIL)-1β. Effects of prior incubation with glucosamine (2.5 to 10.0 µg/mL) and CS (5.0 to 50.0 µg/mL) on gene expression of matrix metalloproteinase (MMP)-1, -2, -3, -9, and -13; aggrecanase 1 and 2; inducible nitric oxide synthase (iNOS); cyclooxygenase (COX)-2; nuclear factor κB; and c-Jun- N-terminal kinase (JNK) were assessed by use of a quantitative real-time polymerase chain reaction assay.
Results—Glucosamine at a concentration of 10 µg/mL significantly reduced reIL-1β–induced mRNA expression of MMP-13, aggrecanase 1, and JNK. Reductions in cytokine-induced expression were also observed for iNOS and COX-2. Chondroitin sulfate had no effect on gene expression at the concentrations tested.
Conclusions and Clinical Relevance—Concentrations of glucosamine similar to those achieved in plasma after oral administration in horses exerted pretranslational regulation of some mediators of osteoarthritis, an effect that may contribute to the cartilage- sparing properties of this aminomonosaccharide. Analysis of results of this study indicated that the influence of CS on pretranslational regulation of these selected genes is limited or lacking. (Am J Vet Res 2005;66:1861–1869)
