Inhibition of the Extracellular Signal-regulated Kinase Pathway Restores the Discogenic Phenotype of Inflammatory Intervertebral Disc Cells

Background: Intervertebral disc (IVD) degeneration is a spinal disease caused by trauma and/or repetitive mechanical overloading of the spine which triggers inflammatory response pathways. Long-term disc inflammation may lead to development of spinal pseudoarthrosis. The aim of the present study was to elucidate the role of the extracellular signal-regulated kinase (ERK) pathway in inflammation-induced IVD cells. Methods: Inflammatory human nucleus pulposus cells (NPC) were stimulated using tumor necrosis factor alpha (TNFα) and the ERK pathway was blocked using a selective molecule-based inhibitor U0126. Gene expression of catabolic and anabolic events, pro-inflammatory, and NPC markers were investigated. The enzymatic activity of matrix metalloproteinases (MMP)2/9 were determined by gelatin zymography. The cytoxicity of U0126 concentrations on NPC was quantified using resazurin assay, and the specificity of U0126 on ERK1/2 signaling was determined.Results: The pro-inflammatory cytokines like MMP3/13 and interleukin 6 in nucleus pulposus (NP) inflammatory conditions were down-regulated by U0126 and a trend towards an increase of the NP-specific collagen type 2, aggrecan and keratin 19 was observed suggesting a recovery of the NP phenotype. U0126 does not seem to have effect on prostaglandin production, aggrecanases and some anabolic genes. We confirmed that U0126 selectively blocks the ERK phosphorylation and U0126 affects the cells metabolic activity only for high concentrations. Conclusions: Inhibition of ERK signaling down-regulates important metalloproteinase, pro-inflammatory cytokines, and up-regulates NP markers in order to restore the discogenic phenotype of inflammatory NPC.