Adamus G, Machnicki M, Seigel G M
R.S. Dow Neurological Sciences Institute, Legacy-Good Samaritan Hospital and Medical Center, Portland, Oregon 97209, USA.
Invest Ophthalmol Vis Sci. 1997 Feb;38(2):283-91.
Recoverin has been identified as a target autoantigen for antirecoverin antibodies found in the sera of some patients with cancer-associated retinopathy. The aim of this study was to investigate the role of antirecoverin antibodies in cancer-associated retinopathy.
Human, rat, and rabbit antirecoverin antibodies were purified using a recoverin-affinity column. Purified biotinylated antibodies were cultured with recoverin-positive rat retinal cells E1A.NR3. Antibody uptake by retinal cells in vitro was analyzed by immunocytochemistry. Cytotoxic effect of antibodies on retinal cells was measured by the MTT colorimetric method. Apoptosis was shown by the ladder DNA fragmentation method and by fluorescent dye chromatin fragmentation analysis.
Antirecoverin antibodies obtained either from sera from five cancer-associated retinopathy patients or from sera of immunized animals were internalized by E1A.NR3 cells. Only specific, antirecoverin antibodies produced destruction of the cells in a dose- and time-dependent manner. Normal immunoglobulin G did not have such effects on retinal cells. No additional cell destruction was observed in the presence of complement as compared with cultures incubated with antirecoverin antibodies alone. Internucleosomal DNA fragmentation and presence of apoptotic cells was observed throughout the culture treated with recoverin specific antibodies but not with normal antibodies. Cells not expressing recoverin (Y79, PC12, and GH3) were not susceptible to cell destruction because of antirecoverin antibody action.
These studies showed that antibodies specific to recoverin are able to enter and cause death of cells expressing recoverin. In humans, autoantibodies originally elicited against recoverin expressed in tumor cells may damage retinal photoreceptors and play a role in the pathogenesis of cancer-associated retinopathy. Results suggest that autoantibody to recoverin, when given access to recoverin in the retina through the blood-retina barrier, could initiate photoreceptor degeneration leading to blindness. Such mechanism may be common for other paraneoplastic disorders or autoimmune diseases where antibodies interfere with the normal cell physiology.
恢复蛋白已被确定为在一些癌症相关性视网膜病变患者血清中发现的抗恢复蛋白抗体的靶自身抗原。本研究的目的是探讨抗恢复蛋白抗体在癌症相关性视网膜病变中的作用。
使用恢复蛋白亲和柱纯化人、大鼠和兔抗恢复蛋白抗体。将纯化的生物素化抗体与表达恢复蛋白的大鼠视网膜细胞E1A.NR3一起培养。通过免疫细胞化学分析体外视网膜细胞对抗体的摄取。用MTT比色法测定抗体对视网膜细胞的细胞毒性作用。通过梯形DNA片段化法和荧光染料染色质片段化分析显示细胞凋亡。
从5例癌症相关性视网膜病变患者血清或免疫动物血清中获得的抗恢复蛋白抗体被E1A.NR3细胞内化。只有特异性抗恢复蛋白抗体以剂量和时间依赖性方式导致细胞破坏。正常免疫球蛋白G对视网膜细胞没有这种作用。与单独用抗恢复蛋白抗体培养的培养物相比，在补体存在下未观察到额外的细胞破坏。在用恢复蛋白特异性抗体处理的整个培养物中观察到核小体间DNA片段化和凋亡细胞的存在，但正常抗体处理的培养物中未观察到。不表达恢复蛋白的细胞（Y79、PC12和GH3）由于抗恢复蛋白抗体的作用而不易受到细胞破坏。
这些研究表明，针对恢复蛋白的特异性抗体能够进入并导致表达恢复蛋白的细胞死亡。在人类中，最初针对肿瘤细胞中表达的恢复蛋白产生的自身抗体可能损害视网膜光感受器，并在癌症相关性视网膜病变的发病机制中起作用。结果表明，当恢复蛋白自身抗体通过血视网膜屏障进入视网膜中的恢复蛋白时，可能引发光感受器变性导致失明。这种机制可能与其他副肿瘤性疾病或自身免疫性疾病相同，在这些疾病中抗体干扰正常细胞生理功能。
