title: Anti-recoverin antibodies induce an increase in intracellular calcium, leading to apoptosis in retinal cells.
abstract: Autoantibodies against recoverin, a Ca2+-binding protein found in patients with cancer-associated retinopathy (CAR syndrome), penetrate retinal cells and induce their apoptosis via a mitochondrial pathway. The goal of this study was to investigate whether the entry of anti-recoverin antibody into E1A.NR3 retinal cells causes a change in intracellular Ca2+. Intracellular Ca2+ was measured using the Ca2+-sensitive fluorescent dye Fura-2 AM in living E1A.NR3 retinal cells treated with anti-recoverin antibody Rec-1, patients' autoantibodies, and control rat and human IgG. The exposure of retinal cells to Rec-1 antibody and to the CAR patients' autoantibodies in vitro caused a significant increase in intracellular Ca2+, while non-specific antibodies did not induce such an effect. Co-treatment of the E1A.NR3 cells with Rec-1 in the presence of nifedipine, a L-type Ca2+ channel blocker, significantly suppressed the increase of Ca2+. Treatment with nifedipine also blocked changes in the anti-apoptotic protein bcl-xL and in expressions of the pro-apoptotic protein bax. Nifedipine-treated cells also showed a decrease in cytosolic cytochrome c release and a decrease in caspase 3 activation, compared to cells treated only with Rec-1 antibody. The increase in the antibody-induced Ca2+ is at least in part dependent on extracellular Ca2+. Nifedipine was found to inhibit the entry of Ca2+ into the cells and to protect them from Rec-1-induced apoptosis. Increased levels of intracellular Ca2+ may lead to retinal dysfunction and degeneration in the CAR syndrome. Our results provide a molecular basis for the use of Ca2+ blockers in the treatment of the CAR syndrome.
authors.affiliation: Neurological Sciences Institute, Oregon Health & Science University, 505 N.W. 185th Avenue, Beaverton, OR 97006, USA. adamusg@ohsu.edu
authors.forename: Grazyna
authors.initials: G
authors.lastname: Adamus
authors.forename: Sarah
authors.initials: S
authors.lastname: Webb
authors.forename: Sharon
authors.initials: S
authors.lastname: Shiraga
authors.forename: Robert M
authors.initials: RM
authors.lastname: Duvoisin
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D001323:Autoantibodies / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology / Q000627:therapeutic use; D002135:Calcium-Binding Proteins / Q000037:antagonists & inhibitors* / Q000276:immunology; D002460:Cell Line; D045304:Cytochromes c / Q000378:metabolism; D006801:Humans; D009369:Neoplasms / Q000150:complications / Q000276:immunology; D009543:Nifedipine / Q000494:pharmacology / Q000627:therapeutic use; D051381:Rats; D051603:Recoverin / Q000037:antagonists & inhibitors* / Q000276:immunology; D012160:Retina / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D012164:Retinal Diseases / Q000188:drug therapy / Q000276:immunology; D013577:Syndrome; D051028:bcl-2-Associated X Protein / Q000378:metabolism; D051020:bcl-X Protein / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D002121:Calcium Channel Blockers; D002135:Calcium-Binding Proteins; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D051603:Recoverin; D045304:Cytochromes c; D009543:Nifedipine; D002118:Calcium