Presentation: Leptin Stimulation Increases Expression Of Frizzled-1, Discoidin Domain Receptor-2 And Receptor For Advanced Glycation End Products In Articular Chondrocytes (2008)

1303 Leptin Stimulation Increases Expression Of Frizzled-1, Discoidin Domain Receptor-2 And Receptor For Advanced Glycation End Products In Articular Chondrocytes

Purpose: Knee osteoarthritis (OA) is the most common cause of musculoskeletal disability in the elderly and currently no medical treatments are available that alter the progressive natural history of the disease. Numerous epidemiological studies have linked obesity to an increased risk of knee OA, however the molecular mechanisms that define the relationship between them is unknown. Clinically significant OA is characterized by the loss of articular cartilage and proliferation of subchondral/periarticular bone which implicates altered matrix metabolism and turnover in the pathological process. We hypothesized that leptin, a major adipokine present in synovial fluid, alters the expression of cell surface receptors on articular chondrocytes that may be linked to pathophysiology of OA.
Methods: The human chondrocyte cell line (C28/I2) was grown in DMEM with 10% FBS. All experiments were performed using at passage 5. Cells were grown to confluence in DMEM with 10% FBS, then exposed to low serum media, for a period of 24 hours. The cells were then treated with recombinant human leptin and grown for an additional 48 hours. Replicate aliquots of cells were exposed to cyclohexamide for 30 minutes at 37oC before the addition of leptin. At various time points following leptin stimulation cells were harvested and total cell lysates were used to perform immunoblots for the detection of ERK1/2, STAT3, Frizzled-1 (Fzd1), discoidin domain receptor-2 (DDR-2) and receptor for advanced glycation endproducts (RAGE). Immunoblot bands specific for the respective proteins were visualized after exposure to film.
Results: Leptin stimulation of the C28/I2 was associated with activation of previously reported signal transduction pathways including ERK1/2 and STAT3 from 3-100 minutes following leptin stimulation. Fzd1 and RAGE increased 3 fold while DDR-2 increased 2 fold 24 hours following leptin stimulation. In every case upregulated expression was also observed at 48 hours following leptin stimulation. The upregulated expression of the respective cell surface receptors was blocked by pre-treatment with cyclohexamide indicating that the increased levels of protein was a result of de novo gene expression.
Conclusions: These experiments suggest that leptin can directly increase the expression of several cell surface receptors in articular chondrocytes Fzd1, DDR2 and RAGE that may play a role in the pathophysiology of OA. Ligand stimulation of all three of these receptors has been associated with the upregulated expression of matrix metalloproteinases, specifically MMP-3 and MMP-13, both of which have been implicated in the loss of cartilage and remodeling of bone that occurs in progressive OA. Additional studies to define specifically the leptin signaling events that are linked to the increased expression of the specific proteins will provide us with an improved understanding of OA and may lead to the identification of novel treatments.

 S. Ohba, None; T.M. Lanigan, None; B.J. Roessler, None.