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Renu A Kowluru, Jadwiga Flaga, Manish Mishra; Transcriptional Regulation of Matrix Metalloproteinase-9 in Diabetic Retinopathy. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):4271.
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Diabetes upregulates retinal matrix metalloproteinase-9 (MMP-9), and activated MMP-9 damages the mitochondria, initiating the apoptotic machinery. The promoter region of MMP-9 has binding sites for a number of transcriptional factors, including activator protein 1 (AP-1), specificity protein 1 and nuclear factor-kB (NF-kB). AP-1 is predominantly composed of subunits of the protein from the Jun and Fos families, and activation of AP-1 results in its translocation from the cytoplasm to the nucleus. MMP-9 promoter has two binding sites for AP-1, one in the distal region (-538 to -529) and the other in its proximal region (-80 to 71). In diabetes, the MMP-9 promoter is epigenetically modified and acetylated H3K9 levels are elevated, and this facilitates the binding of p65 subunit of NF-kB. The aim of this study is to investigate the role of AP-1 in the regulation of MMP-9 expression in diabetes.
Retinal endothelial cells, incubated in normal (5mM) or high (20mM) glucose for 4 days, were analyzed for the binding of c-Fos and c-Jun subunits of AP-1 at both the distal and proximal sites of the MMP-9 promoter by chromatin immunoprecipitation technique. The effect of high glucose on their nuclear accumulation was determined by immunofluorescence, and was confirmed by western blot technique by quantifying their expressions in the nuclear fraction. The results were validated in the retina from streptozotocin-induced diabetic rats.
High glucose increased the binding of both subunits of Ap1 (c-Fos and c-Jun) at the distal and at the proximal sites of the MMP-9 promoter. However, in the same cell preparations, the nuclear translocation of c-Fos and c-Jun was not altered. Consistent with the in vitro results, diabetes also increased the binding of c-Fos and c-Jun at both, the proximal and the distal sites of the retinal MMP-9 promoter.
Thus, increased binding of AP-1 at the retinal MMP-9 promoter suggests that in diabetes, AP-1 functions as a positive regulator of MMP-9 transcription, and the epigenetic modifications of the MMP-9 promoter could be facilitating the binding of AP-1. Identifying the mechanism responsible for MMP-9 regulation in diabetes could prevent its activation, and protect retinal capillary cells from undergoing accelerated apoptosis.<br />
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