title: Diabetes-induced activation of caspase-3 in retina: effect of antioxidant therapy.
abstract: Apoptosis of retinal endothelial cells and pericytes is postulated to contribute to the development of retinopathy in diabetes. The goal of this study is to investigate diabetes-induced activation of retinal caspase-3, an apoptosis executer enzyme, in retina, and examine the effects of antioxidants on the activation. Caspase-3 activation was determined in the retina of alloxan diabetic rats (2-14 months duration) and in the isolated retinal capillary cells (endothelial cells and pericytes) by measuring cleavage of caspase-3 specific fluorescent substrate, and cleavage of caspase-3 holoenzyme and poly (ADP ribosyl) polymerase. Effect of antioxidants on the activation of caspase-3 was determined by feeding a group of diabetic rats diet supplemented with a comprehensive mixture of antioxidants, including Trolox, alpha-tocopherol, N-acetyl cysteine, ascorbic acid, beta-carotene and selenium for 2-14 months, and also under in vitro conditions by incubating isolated retinal capillary cells with antioxidants with wide range of actions. Caspase-3 was activated in the rat retina at 14 months of diabetes (P < 0.05 vs. normal), but not at 2 months of diabetes, and administration of antioxidants for the entire duration inhibited this activation. In the isolated retinal capillary cells incubated in 25 mM glucose medium, caspase-3 activity was increased by 50% compared to the cells incubated in 5 mM glucose (P < 0.02), and antioxidants or caspase-3 inhibitor inhibited this increase. Our results suggest that increased oxidative stress in diabetes is involved in the activation of retinal caspase-3 and apoptosis of endothelial cells and pericytes. Antioxidants might be inhibiting the development of diabetic retinopathy by inhibiting microvascular apoptosis.
authors.affiliation: Kresge Eye Institute, Wayne State University, 4717 St. Antoine, Detroit, MI 48201, USA. rkowluru@med.wayne.edu
authors.forename: Renu A
authors.initials: RA
authors.lastname: Kowluru
authors.forename: Prashant
authors.initials: P
authors.lastname: Koppolu
mesh_terms: D000111:Acetylcysteine / Q000008:administration & dosage; D000818:Animals; D000975:Antioxidants / Q000627:therapeutic use*; D017209:Apoptosis; D001205:Ascorbic Acid / Q000008:administration & dosage; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D003921:Diabetes Mellitus, Experimental / Q000201:enzymology*; D003930:Diabetic Retinopathy / Q000201:enzymology / Q000473:pathology / Q000517:prevention & control*; D004730:Endothelium, Vascular / Q000201:enzymology / Q000473:pathology; D004789:Enzyme Activation; D005947:Glucose / Q000378:metabolism; D066298:In Vitro Techniques; D008297:Male; D018384:Oxidative Stress; D020286:Pericytes / Q000201:enzymology / Q000473:pathology; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D012160:Retina / Q000201:enzymology*; D012643:Selenium / Q000008:administration & dosage; D017392:Thiobarbituric Acid Reactive Substances / Q000378:metabolism; D024502:alpha-Tocopherol / Q000008:administration & dosage; D019207:beta Carotene / Q000008:administration & dosage
chemical_list: D000975:Antioxidants; D017392:Thiobarbituric Acid Reactive Substances; D019207:beta Carotene; D011065:Poly(ADP-ribose) Polymerases; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; D024502:alpha-Tocopherol; D012643:Selenium; D005947:Glucose; D001205:Ascorbic Acid; D000111:Acetylcysteine