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S.-W. Chang, S.-F. Chou, J.-L. Chuang; Mitomycin C Upregulates IL-8 and MCP-1 Chemokine Expression via Mitogen-Activated Protein Kinases in Corneal Fibroblasts. Invest. Ophthalmol. Vis. Sci. 2007;48(13):2750.
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To investigate the expression of chemokines and their signaling pathways following application of mitomycin C (MMC) to corneal fibroblasts.
Primary porcine and human corneal fibroblasts from passages 3-6 were treated with MMC at concentrations of 0.05, 0.1, or 0.2mg/mL for 1, 2, 5, or 10 minutes. The relative expressions of interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) were investigated with reverse transcription and quantitative real-time polymerase chain reaction (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA). The effects of MMC on the activation of kinases were analyzed by Western blot analysis with specific anti-phosphokinase antibodies. The signaling pathways by which MMC regulates the expression of IL-8 and MCP-1 were evaluated by pharmacological kinase-specific inhibitors.
The expressions of IL-8 and MCP-1 were upregulated after MMC treatment in both treatment time- and concentration-dependent manners. Furthermore, the upregulated expressions of IL-8 and MCP-1 increased with longer incubation time. MMC treatment enhanced the phosphorylation of p38, JNK, and ERK at different time points. The MMC-related IL-8 and MCP-1 expressions were inhibited by both a p38-specific inhibitor (SB203580) and an ERK-specific inhibitor (PD98059). A JNK inhibitor (SP600125) reduced the expression of MMC-induced MCP-1 but not of IL-8.
MMC treatment upregulated the expression of IL-8 and MCP-1 mRNA and protein secretion by the activation of MAPKs in corneal fibroblasts.
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