title: Alpha-phenyl-N-tert-butylnitrone (PBN) prevents light-induced degeneration of the retina by inhibiting RPE65 protein isomerohydrolase activity.
abstract: α-Phenyl-N-tert-butylnitrone (PBN), a free radical spin trap, has been shown previously to protect retinas against light-induced neurodegeneration, but the mechanism of protection is not known. Here we report that PBN-mediated retinal protection probably occurs by slowing down the rate of rhodopsin regeneration by inhibiting RPE65 activity. PBN (50 mg/kg) protected albino Sprague-Dawley rat retinas when injected 0.5-12 h before exposure to damaging light at 2,700 lux intensity for 6 h but had no effect when administered after the exposure. PBN injection significantly inhibited in vivo recovery of rod photoresponses and the rate of recovery of functional rhodopsin photopigment. Assays for visual cycle enzyme activities indicated that PBN inhibited one of the key enzymes of the visual cycle, RPE65, with an IC(50) = 0.1 mm. The inhibition type for RPE65 was found to be uncompetitive with K(i) = 53 μm. PBN had no effect on the activity of other visual cycle enzymes, lecithin retinol acyltransferase and retinol dehydrogenases. Interestingly, a more soluble form of PBN, N-tert-butyl-α-(2-sulfophenyl) nitrone, which has similar free radical trapping activity, did not protect the retina or inhibit RPE65 activity, providing some insight into the mechanism of PBN specificity and action. Slowing down the visual cycle is considered a treatment strategy for retinal diseases, such as Stargardt disease and dry age-related macular degeneration, in which toxic byproducts of the visual cycle accumulate in retinal cells. Thus, PBN inhibition of RPE65 catalytic action may provide therapeutic benefit for such retinal diseases.
authors.affiliation: Department of Ophthalmology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA. mmandal@ouhsc.edu
authors.forename: Md Nawajes A
authors.initials: MN
authors.lastname: Mandal
authors.forename: Gennadiy P
authors.initials: GP
authors.lastname: Moiseyev
authors.forename: Michael H
authors.initials: MH
authors.lastname: Elliott
authors.forename: Anne
authors.initials: A
authors.lastname: Kasus-Jacobi
authors.forename: Xiaoman
authors.initials: X
authors.lastname: Li
authors.forename: Hui
authors.initials: H
authors.lastname: Chen
authors.forename: Lixin
authors.initials: L
authors.lastname: Zheng
authors.forename: Olga
authors.initials: O
authors.lastname: Nikolaeva
authors.forename: Robert A
authors.initials: RA
authors.lastname: Floyd
authors.forename: Jian-Xing
authors.initials: JX
authors.lastname: Ma
authors.forename: Robert E
authors.initials: RE
authors.lastname: Anderson
mesh_terms: D000217:Acyltransferases / Q000378:metabolism; D000429:Alcohol Oxidoreductases / Q000378:metabolism; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D003497:Cyclic N-Oxides / Q000494:pharmacology*; D005136:Eye Proteins / Q000378:metabolism*; D008027:Light / Q000009:adverse effects*; D018696:Neuroprotective Agents / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012162:Retinal Degeneration / Q000201:enzymology / Q000517:prevention & control; D017948:Retinal Rod Photoreceptor Cells / Q000201:enzymology*; D012243:Rhodopsin / Q000378:metabolism*; D019745:cis-trans-Isomerases / Q000378:metabolism*; D000099178:Retinoid Isomerohydrolase
chemical_list: D000217:Acyltransferases; D000429:Alcohol Oxidoreductases; D002352:Carrier Proteins; D003497:Cyclic N-Oxides; D005136:Eye Proteins; D018696:Neuroprotective Agents; D012243:Rhodopsin; D019745:cis-trans-Isomerases; D000099178:Retinoid Isomerohydrolase; C029217:phenyl-N-tert-butylnitrone; C022520:retinol dehydrogenase; C056942:lecithin-retinol acyltransferase