Glucoasamine interferes with mitogen-activated protein kinase pathway by inhibiting JNK and p38 phosphorylation in human chondrocytes.

Previous studies demonstrated the ability of Glucosamine (GlcN) to inhibit mRNA transcription level of Interleukin-1β (IL-1β)- stimulated genes. These genes are under the control of two transcription factors Activator Protein (AP)-1 and NF-kB. The aim of this study was to determine the effects of GlcN on mitogen-activated protein (MAP) kinase phosphorylation and on activation of AP-1, in human chondrocytes. Human immortalized cell line, lbpva55, and human chondrocytes, obtained from healthy donors, were challenged with 10 ng/ml IL-1β cytokine after pre-treatment with 2.5 or 10 mM GlcN. mRNA expression levels of some matrix metalloprotease (MMP) genes were evaluated by Quantitative-Real Time PCR (Q-RT-PCR), protein production levels were evaluated in the culture supernatant by Enzyme Linked ImmunoSorbent Assay (ELISA). MAP kinase phosphorylation was evaluated by Western Blotting. AP-1 transcription factor activation was evaluated by measuring AP-1 component DNA binding activity (TransAM AP-1 family kit). After IL-1β stimulation, MMP-1, -3 and -13 productions were strongly increased both at mRNA and protein level. Treatment with GlcN reduced the expression of these metalloproteases. MMP-1, -3 and -13 expression is regulated by transcription factors such as AP-1, which is activated by phosphorylated MAP kinases. IL-1β stimulated phosphorylation of c-jun N terminal kinase (JNK), p38 MAP kinase and extracellular-signal regulated kinase (ERK)-1/2. GlcN inhibited JNK and p38 phosphorylation and consequently c-jun and to a lower degree junD DNA binding activity. Moreover, we found also down-regulation of c-jun mRNA transcription level. These results demonstrated for the first time, in human chondrocytes, that GlcN inhibits cytokine-stimulated MMP production, by affecting MAP kinase phosphorylation and consequently AP-1 transcription factor.