Increased oxidative stress in diabetes regulates activation of a small molecular weight G-protein, H-Ras, in the retina.
OBJECTIVE
Increased superoxide levels are implicated in the pathogenesis of diabetic retinopathy. We have shown that functional activation of a small molecular weight G-protein, H-Ras, is one of the signaling steps involved in glucose-induced apoptosis of retinal capillary cells. The goal of this study was to elucidate the mechanism(s) by which oxidative stress could result in the activation of H-Ras in diabetes.


METHODS
Experiments were performed in isolated retinal endothelial cells that were treated with H(2)O(2), or the cells in which glucose-induced superoxide accumulation was inhibited either by superoxide dismutase mimetic (MnTBAP) or by overexpressing mitochondrial superoxide dismutase (MnSOD). The in vitro experiments were complemented with in vivo experiments using the retina from mice overexpressing MnSOD.


RESULTS
H(2)O(2) activated H-Ras and its downstream signaling pathway, including Raf-1 and phosphorylation of p38 (p-p38) MAP kinase. Inhibition of superoxide significantly attenuated glucose-induced activation of H-Ras, Raf-1 and p-p38 MAP kinase. Overexpression of MnSOD in mice prevented diabetes-induced activation of both H-Ras and p-p38 MAP kinase.


CONCLUSIONS
Our results clearly indicate that the activation of H-Ras and its downstream signaling pathway in the retina and its vasculature could be under the control of superoxide, and H-Ras activation in diabetes can be prevented by inhibiting superoxide accumulation.
Kowluru
Vibhuti
V
Department of Ophthalmology, Kresge Eye Institute, Wayne State University, Detroit, MI 48201, USA.
Kowluru
Renu A
RA
D000818:Animals; D017209:Apoptosis; D002417:Cattle; D002478:Cells, Cultured; D003921:Diabetes Mellitus, Experimental / Q000235:genetics* / Q000378:metabolism* / Q000503:physiopathology; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D016166:Free Radical Scavengers / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D011905:Genes, ras; D005947:Glucose / Q000494:pharmacology; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D012160:Retina / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism; D013997:Time Factors
D016166:Free Radical Scavengers; D016877:Oxidants; D013481:Superoxides; D006861:Hydrogen Peroxide; D013482:Superoxide Dismutase; D019908:Proto-Oncogene Proteins c-raf; D020928:Mitogen-Activated Protein Kinases; D005947:Glucose