The apoptosis inhibitor of macrophage/CD5 molecule-like protein (AIM/CD5L) is expressed in the human retina and is a target of autoreactivity in age-related macular degeneration (AMD)

Abstract

Purpose: To report on the identity of a novel antigen targeted by auto-antibodies (AAbs) found in the serum of 131 AMD patients [previously identified via comparison of Western blots (WB) vs. 245 unaffected subjects, (Lenchik et al. ARVO 2013, Abs. 4103; Iannaccone et al. ARVO 2012, Abs. 2272)] and to show its localization at the human retinal level.

Methods: Protein lysates obtained from human macular full-thickness retina/RPE/Bruch’s membrane/choroid punches were immunoprecipitated with a representative selection of AMD sera positive to WB testing to identify antigens targeted by AAbs using 2D gel electrophoresis (GE) and mass spectrometry (MS). MS methods have been previously reported (Lenchik et al. ARVO 2013, Abs. 4103). A 38-kDa candidate protein was confirmed by immunoprecipitation-WB (IP-WB) and by direct ELISA against recombinant proteins on 18 AMD representative sera vs.16 controls. Immunohistochemistry (IHC) against human macular sections was used to confirm the localization of antigen expression in the human retina.

Results: The 38-kDa candidate protein identified by our screens was confirmed to be the AIM/CD5L protein in multiple independent samples tested by 2DGE and MS. ELISA showed that AMD serum reactivity against AIM/CD5L was 2.5-fold higher than in control samples (student’s t-test, p=0.000007). We show, to the best of our knowledge for the first time, that AIM/CD5L, previously known to be expressed in the mouse retina from EST (NEIBank) and RNA in situ (Jackson Labs) studies, is expressed throughout the human retina, vitread to the outer limiting membrane (OLM), and more intensely at the level of the retinal ganglion cell layer (GCL), outer nuclear layer (ONL) and outer plexiform layer (OPL). Sera of AMD patients stain human retinal sections at locations consistent with its observed expression pattern.

Conclusions: The AIM/CD5L protein is a novel target involved in autoreactivity in patients with AMD. Consistent with our observations in the context of autoreactivity in AMD and with our working hypothesis, a role for AIM/CD5L as a potential immunomodulatory molecular has recently emerged. We suggest that impaired AIM/CD5L retinal function may contribute to AMD in situ. Continued identification of more AAb targets should lead to further insight into the role of autoimmunity in the pathogenesis of AMD.

Keywords: 412 age-related macular degeneration • 555 immunomodulation/immunoregulation • 557 inflammation