u-0126 and Diabetic-Retinopathy

Topics: Animals; Animals, Newborn; Butadienes; CCAAT-Enhancer-Binding Protein-beta; CCAAT-Enhancer-Binding Protein-delta; Cobalt; Diabetic Retinopathy; Gene Expression Regulation; Humans; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; MAP Kinase Signaling System; Neovascularization, Pathologic; Nitriles; Proto-Oncogene Proteins c-fos; Rats; Receptors, G-Protein-Coupled; Retina; Vascular Endothelial Growth Factor A

Diabetic retinopathy (DR) is one of the most serious complications of diabetes and has become a major blinding eye disease, but its treatment remains unsatisfactory. The ERK1/2 signaling pathway has been shown to participate in regulating secretion of VEGF in DR from our previous studies. The role of VEGF in the development of DR provides a target for treatment. Our present research focuses on Müller cells, a major source of VEGF secretion, to investigate the role of ERK1/2 signaling pathway on regulation of VEGF release in diabetes.. Immunofluorescence was used to observe the ERK1/2 phosphorylation activity on early diabetic rat retinal Müller cells. Müller cells were stimulated by high glucose in vitro. Western blot and immunohistochemistry were used to determine ERK1/2 signaling pathway expression and phosphorylation. AP-1 DNA binding activity status was monitored by electrophoretic mobility shift assay (EMSA). ELISA and PCR monitored VEGF secretion. Inhibition of ERK1/2 phosphorylation with U0126 was observed for changes in VEGF secretion.. Phos-ERK1/2 was expressed on Müller cells early in diabetes. In vitro high glucose stimulation of Müller cells increased VEGF secretion with a peak at 24 hours. An ERK1/2 specific inhibitor, U0126, stopped the phosphorylation of ERK1/2, lowered AP-1 DNA binding activity, and reduced Müller cells secretion of VEGF under high glucose conditions.. ERK1/2 signaling pathway has some role in regulating Müller cells secretion of VEGF in DR. Targeting the ERK1/2 signaling pathway in Müller cells through intervention of the upstream signaling pathway or nuclear transcription factors of VEGF secretion could be a type of anti-VEGF treatment for DR.

Topics: Animals; Blotting, Western; Butadienes; Diabetes Mellitus, Experimental; Diabetic Retinopathy; Electrophoretic Mobility Shift Assay; Enzyme Inhibitors; Enzyme-Linked Immunosorbent Assay; Fluorescent Antibody Technique, Indirect; Glucose; Male; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Neuroglia; Nitriles; Phosphorylation; Rats; Rats, Sprague-Dawley; Real-Time Polymerase Chain Reaction; Transcription Factor AP-1; Vascular Endothelial Growth Factor A

Vascular endothelial growth factor (VEGF) is one of the major factors promoting diabetic retinopathy (DR). A better understanding of the signaling pathway in VEGF regulation is of clinical importance to identify more precise therapeutic targets for diabetic retinopathy. The ERK1/2 signaling pathway has been shown to play a key role in some oncoma and hematologic diseases by mediating VEGF release. This research was conducted to determine whether the ERK1/2 signaling pathway also plays a major role in VEGF release in DR development.. One hundred Sprague-Dawley (SD) rats were induced to diabetes by streptozotocin (STZ) injection and monitored at several time points (1, 2, 3, 4, 8, and 12 weeks) for ERK1/2 phosphorylation, Activator protein (AP)-1 activity and concentration, and VEGF protein and mRNA expression, using immunohistochemical and biochemical methods. RESULTS. The ERK1/2 signaling pathway was rapidly activated 1 week after diabetes was induced. AP-1, the downstream transcription factor of ERK1/2, was also activated, and VEGF became highly regulated in a similar trend. U0126, an inhibitor of ERK1/2, also downregulated VEGF expression, in addition to ERK1/2 and AP-1 activity.. ERK1/2 signaling pathway is involved in VEGF release in diabetic rat retina; therefore, ERK1/2 may be a potential therapeutic target of DR.

Topics: Animals; Blotting, Western; Butadienes; Diabetes Mellitus, Experimental; Diabetic Retinopathy; DNA; Electrophoretic Mobility Shift Assay; Enzyme Inhibitors; Immunoenzyme Techniques; Male; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Nitriles; Phosphorylation; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Transcription Factor AP-1; Vascular Endothelial Growth Factor A; Vitreous Body