Protein Kinase C-Mediated Regulation of Matrix Metalloproteinase and Tissue Inhibitor of Metalloproteinase Production in a Human Retinal Muller Cells

Purpose: Matrix metalloproteinases (MMPs) play an important role in the degradation of extracellular matrix (ECM) proteins in the retina. Breakdown of ECM proteins results in neovascularization and tractional retinal detachment, which eventually lead to the symptoms of proliferative diabetic retinopathy. Muller cells are reported to be one of the MMP-producing cells in the retina. However, the molecular mechanism of MMP production derived from Muller cells remains to be fully elucidated. Materials and Methods: The human retinal Muller cell line (MIO-M1) was continuously-subcultured in high glucose (25 mM) condition. After the cells reached confluence, they were treated for 24 h with phorbol ester and/or a protein kinase C (PKC) inhibitor, GF109203X in high (25 mM) or low (5 mM) glucose condition. Gelatinase activities in conditioned medium were assessed using gelatin zymography. RT-PCR was performed to analyze the mRNA expression level of MMP-9. Western blot analysis used to detect the protein expression of tissue inhibitors of metalloproteinases (TIMPs). Electrophoresis mobility shift assay was conducted to examine transcription factors involved in MMP-9 transcription. Results: We demonstrated the protein kinase C (PKC)-mediated regulation of proMMP-9 transcription and protein expression through the action of phorbol ester, an activator of PKC. Moreover, we demonstrated the expression of TIMPs, known as natural inhibitors of MMPs at the protein level in a human retinal Muller cell line for the first time, and report that production of these proteins was also regulated in a PKC-dependent manner. Conclusion: Our results suggest that imbalance between MMP and TIMP proteins may promote neovascularization by PKC activation in retinal Muller cells. In addition, the development of novel compounds with regulatory action on MMP and TIMP production through inhibiting PKC activity in retinal Muller cells may lead to new therapeutic approaches for the treatment and prevention of diabetic retinopathy.