APOPTOTIC RETINAL CELL-DEATH INDUCED BY ANTIRECOVERIN AUTOANTIBODIES OF CANCER-ASSOCIATED RETINOPATHY

Purpose. Recoverin has been identified as a target autoantigen for ant irecoverin antibodies found in the sera of some patients with cancer-a ssociated retinopathy. The aim of this study was to investigate the ro le of antirecoverin antibodies in cancer-associated retinopathy. Metho ds. Human, rat, and rabbit antirecoverin antibodies were purified usin g a recoverin-affinity column. Purified biotinylated antibodies were c ultured with recoverin-positive rat retinal cells E1A.NR3. Antibody up take by retinal cells in vitro was analyzed by immunocytochemistry. Cy totoxic effect of antibodies on retinal cells was measured by the MTT colorimetric method. Apoptosis was shown by the ladder DNA fragmentati on method and by fluorescent dye chromatin fragmentation analysis. Res ults. Antirecoverin antibodies obtained either from sera from five can cer-associated retinopathy patients or from sera of immunized animals were internalized by E1A.NR3 cells. Only specific, antirecoverin antib odies produced destruction of the cells in a dose- and time-dependent manner. Normal immunoglobulin G did not have such effects on retinal c ells. No additional cell destruction was observed in the presence of c omplement as compared with cultures incubated with antirecoverin antib odies alone. Internucleosomal DNA fragmentation and presence of apopto tic cells was observed throughout the culture treated with recoverin-s pecific antibodies but not with normal antibodies. Cells not expressin g recoverin (Y79, PC12, and GH(3)) were not susceptible to cell destru ction because of antirecoverin antibody action. Conclusions. These stu dies showed that antibodies specific to recoverin are able to enter an d cause death of cells expressing recoverin. In humans, autoantibodies originally elicited against recoverin expressed in tumor cells may da mage retinal photoreceptors and play a role in the pathogenesis of can cer-associated retinopathy. Results suggest that autoantibody to recov erin, when given access to recoverin in the retina through the blood-r etina barrier, could initiate photoreceptor degeneration leading to bl indness. Such mechanism may be common for other paraneoplastic disorde rs or autoimmune diseases where antibodies interfere with the normal c ell physiology.