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M. Kanwar, R. A. Kowluru; G-Protein H-Ras in Retinal Microvessels and Its Role in the Development of Diabetic Retinopathy. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4912.
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© ARVO (1962-2015); The Authors (2016-present)
H-Ras, a small molecular weight G-protein, functions as a molecular switch and regulates cell proliferation, differentiation and apoptosis. It is activated in the retina in diabetes and the therapies that inhibit the development of diabetic retinopathy in rodents also inhibit its activation. The purpose of this study is to investigate the effect of diabetes on the activation of H-Ras in the retinal microvessels, the major site of retinal histopathology in diabetes, and to elucidate the H-Ras mediated signaling steps in the retinal microvessels.
Freshly isolated retina from streptozotocin diabetic rats (2 months duration) and age-matched normal rats was used to prepare microvessels using hypotonic lysis method. Ras activation was quantified in the microvessels by Raf-1 binding assay, and the activation of signaling proteins, including Raf-1, p38MAPK, by quantifying their gene transcripts by real time RTPCR and /or protein expression by western blot.
Two months of diabetes activated H-Ras (by Raf-binding assay) by over 40% in the retinal microvessels. In the same microvessel preparations gene transcripts of H-Ras and its effector protein Raf-1 were elevated by 30% and 135% respectively, and their protein expressions by about 25% compared to the values obtained from normal rat retinal microvessels. The gene expression of Ras-Raf-1 downstream signaling protein MAP kinase was increased by over 2 fold. Two months of diabetes also significantly activated transcriptional factor NF-kB and iNOS in the retinal microvessels.
H-Ras and its signaling pathway is activated in the retinal microvessels in diabetes suggesting that H-Ras mediated signaling pathway, in part, could be contributing to the microvascular pathology characteristic of diabetic retinopathy. Inhibition of H-Ras function presents an important part of the complex approach to inhibit the pathogenesis of diabetic retinopathy.
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