TY - JOUR
T1 - Short-term gene expression changes in cartilage explants stimulated with interleukin 1β plus glucosamine and chondroitin sulfate
AU - Chan, Pooi See
AU - Caron, John P.
AU - Orth, Michael W.
PY - 2006/7
Y1 - 2006/7
N2 - Objective. To determine the short-term effects of glucosamine (GLN) and chondroitin sulfate (CS) on expression of genes encoding inflammatory mediators and matrix enzymes in bovine cartilage explants stimulated with interleukin 1 (IL-1). Methods. Dose-response experiments were conducted for IL-1, GLN, and CS to select concentrations of each optimized for detecting treatment effects on cartilage explants. Based on the dose-response experiments, treatments included fetal bovine serum (FBS) control, 15 ng/ml IL-1, and 15 ng/ml IL-1 with the addition of 10 μg/ml GLN and 20 μg/ml CS. Media were measured for nitric oxide (NO) and prostaglandin E 2 (PGE 2) while explants were frozen for RNA extraction at 8, 16, and 24 hours. Gene expression relative to FBS control for inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), microsomal PGE synthase-1 (mPGEs1), nuclear factor-κB p65 subunit (NF-κB), matrix metalloproteinase (MMP)-3 and 13, aggrecanase (Agg)-1 and 2, and tissue inhibitor of metalloproteinase-3 (TIMP-3) were assessed by quantitative real-time polymerase chain reaction (RT-PCR). In a separate study using incubation of explants with the same treatments for 48 hours, proteoglycan release was measured with dimethylmethylene blue assay and TIMP-3 protein was evaluated with Western blots. Results. The GLN and CS combination abrogated IL-1-induced gene expression of iNOS, COX-2, mPGEs1, and NF-κB at all timepoints. NO, PGE 2, and proteoglycan release were reduced with the combination. The abundance of stimulated MMP-13, Agg-1, and Agg-2 mRNA was repressed, whereas TIMP-3 was upregulated by the combination at all timepoints. The abundance of TIMP-3 protein was increased by the combination relative to IL-1 at 48 hours. Conclusion. GLN and CS in combination suppress synthesis and expression of genes encoding inflammatory mediators and proteolytic enzymes while upregulating TIMP-3. This provides a plausible mechanism for the purported mild antiinflammatory and chondroprotective properties of GLN and CS.
AB - Objective. To determine the short-term effects of glucosamine (GLN) and chondroitin sulfate (CS) on expression of genes encoding inflammatory mediators and matrix enzymes in bovine cartilage explants stimulated with interleukin 1 (IL-1). Methods. Dose-response experiments were conducted for IL-1, GLN, and CS to select concentrations of each optimized for detecting treatment effects on cartilage explants. Based on the dose-response experiments, treatments included fetal bovine serum (FBS) control, 15 ng/ml IL-1, and 15 ng/ml IL-1 with the addition of 10 μg/ml GLN and 20 μg/ml CS. Media were measured for nitric oxide (NO) and prostaglandin E 2 (PGE 2) while explants were frozen for RNA extraction at 8, 16, and 24 hours. Gene expression relative to FBS control for inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), microsomal PGE synthase-1 (mPGEs1), nuclear factor-κB p65 subunit (NF-κB), matrix metalloproteinase (MMP)-3 and 13, aggrecanase (Agg)-1 and 2, and tissue inhibitor of metalloproteinase-3 (TIMP-3) were assessed by quantitative real-time polymerase chain reaction (RT-PCR). In a separate study using incubation of explants with the same treatments for 48 hours, proteoglycan release was measured with dimethylmethylene blue assay and TIMP-3 protein was evaluated with Western blots. Results. The GLN and CS combination abrogated IL-1-induced gene expression of iNOS, COX-2, mPGEs1, and NF-κB at all timepoints. NO, PGE 2, and proteoglycan release were reduced with the combination. The abundance of stimulated MMP-13, Agg-1, and Agg-2 mRNA was repressed, whereas TIMP-3 was upregulated by the combination at all timepoints. The abundance of TIMP-3 protein was increased by the combination relative to IL-1 at 48 hours. Conclusion. GLN and CS in combination suppress synthesis and expression of genes encoding inflammatory mediators and proteolytic enzymes while upregulating TIMP-3. This provides a plausible mechanism for the purported mild antiinflammatory and chondroprotective properties of GLN and CS.
KW - Arthritis
KW - Chondroitin sulfate
KW - Gene expression
KW - Glucosamine
KW - Inflammatory mediators
KW - Matrix metalloproteinase
UR - http://www.scopus.com/inward/record.url?scp=33745798175&partnerID=8YFLogxK
M3 - Article
C2 - 16821268
AN - SCOPUS:33745798175
SN - 0315-162X
VL - 33
SP - 1329
EP - 1340
JO - Journal of Rheumatology
JF - Journal of Rheumatology
IS - 7
ER -