Elucidation on Predominant Pathways Involved in the Differentiation and Mineralization of Odontoblast-Like Cells by Selective Blockade of Mitogen-Activated Protein Kinases.

Aim: To analyze the effect of three mitogen-activated protein kinase (MAPK) inhibitors, namely, SB202190 (p38 inhibitor), SP600125 (JNK inhibitor), and PD98059 (ERK inhibitor) in Dex-stimulated MDPC-23 cell differentiation and mineralization.

Methods: Experiment was divided into five groups, control (cells without Dex and inhibitors treatment), Dex (cells with Dex treatment but without inhibitors), Dex + SB202190, Dex + SP600125, and Dex + PD98059. Cell differentiation was assessed by alkaline phosphatase (ALP) activity assay and real time RT-PCR. Cell mineralization was investigated by alizarin red staining.

Results: Exposure to SB202190 (20  μ M) significantly decreased the mineral deposition in Dex-treated cells as demonstrated by alizarin red staining. Treatment of SP600125 (20  μ M) attenuated the mineralization as well, albeit at a lower degree as compared to SB202190 (20  μ M). Similarly, SB202190 (20  μ M) completely abrogated the ALP activity stimulated by Dex at six days in culture, while no changes were observed with regard to ALP activity in SP600125 (20  μ M) and PD98059 (20  μ M) treated cells. The upregulation of bone sialoprotein (BSP), ALP, and osteopontin (OPN) in Dex challenged cells was completely inhibited by SB202190.

Conclusion: Blockade of p38-MAPK signaling pathway resulted in significant inhibition of ALP activity, mineralization, and downregulation of osteogenic markers. The data implicated that p38 signaling pathway plays a critical role in the regulation of MDPC-23 cells differentiation and mineralization.