Transcriptional regulation of the mPGES-1 gene in primary cultured articular chondrocytes.

Healthy cartilage is maintained in a state of dynamic equilibrium by matrix synthesis and matrix degradation by the chondrocytes. Any dysregulation with increased degradation and/or inadequate synthesis leads to the loss of tissue structure and function, as in rheumatoid arthritis and osteoarthritis. The proinflammatory cytokine IL-1 plays a major role in this phenomenon. IL-1 acts on chondrocytes in part by stimulating the release of prostaglandin E2 (PGE2) at the sites of inflammation. Recently, a human membrane-associated prostaglandin E2 synthase-1 (mPGES-1) was cloned. This enzyme catalyzes the conversion of prostaglandin H2 to PGE2 in a highly specific manner. We previously demonstrated that mPGES-1 mRNA is induced by IL-1 in chondrocytes in a dose-dependent and time-dependent manner. In order to study the transcriptional regulation of mPGES-1 in primary rabbit articular chondrocytes, we have cloned its promoter upstream of the CAT ORF (vector pCAT3-basic; Promega, Charbonnieresles-Bains, France). We show by transient transfection experiments that the mPGES-1 promoter is stimulated by IL-1. A close examination of putative binding sites has revealed the presence of two CCAAT/Enhancer Binding Protein (C/EBP) binding sequences (TTNNGNAAT) located between -548 to -558 base pairs and -610 to -619 base pairs. Co-transfections of expression vectors encoding the two different isoforms of C/EBP (β and δ) strongly stimulate the promoter activity. To further study the role of C/EBP in mPGES-1 expression, we performed gel shift experiments on wild-type and mutated oligonucleotides derived from the mPGES-1 sequence. These experiments confirm the specific binding of C/EBP on the mPGES-1 promoter. Taken together, our results suggest that C/EBP factors indeed bind and regulate the mPGES promoter in articular chondrocytes.