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Shin Yoshitake, Tomoaki Murakami, Yoko Dodo, Kiyoshi Suzuma, Akihito Uji, Masahiro Fujimoto, Nagahisa Yoshimura; . Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):5172.
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© ARVO (1962-2015); The Authors (2016-present)
Optical coherence tomography (OCT) delineates inner segment ellipsoid zone (ISe) of photoreceptor cells, in which mitochondria are highly accumulated. In eyes with diabetic macular edema (DME), the disruption in the ISe line is related to visual impairments, although the molecular mechanisms regarding the photoreceptor damage remains ill-defined. We here identified an autoantigen and analyzed the functions of the autoantibody in DME.
Retinal images and serum specimens were obtained from diabetic patients, who were divided into three groups, i.e., diabetic patients without diabetic retinopathy (DR) (DM group), DR patients without DME (DR group), and DME patients (DME group). In order to show the presence of autoantibodies, Western blot and immunostaining using serum IgG were performed. After crosslinking between serum IgG and protein G, cell lysates of porcine photoreceptors were immunoprecipitated with serum IgG, followed by mass spectrometry analyses to identify the autoantigen. The titer of serum IgG was evaluated using enzyme-linked immunosorbent assay (ELISA). Serum specimens were injection into the subretinal spaces of C57BL/6 mice for the functional analyses of the autoantibody.
Immnoblotting and immunohistochemistry showed that DME serum contained autoantibodies for proteins in photoreceptor cells. The combined methods of immunoprecipitation and mass spectrometry identified a novel autoantigen, fumarase, which is one of the enzymes in the citric acid cycle within the mitochondrial matrix. The titer of anti-fumarase antibodies in DME serum was significantly higher than that in DM serum or DR serum. In animal experiments, histological analyses and OCT images demonstrated that serum IgG from DME patients was adherent to photoreceptor inner and outer segments and colocalized to C5b-9 which represents complement activation, 24 hours after the injection. Photoreceptor cells were damaged in eyes injected with DME serum, which was restored in eyes administrated with the serum in which anti-fumarase antibodies were depleted 7 days after the treatment.
We identified a novel autoantigen, fumarase, and demonstrated the contribution of anti-fumarase antibody to the pathogenesis in DME.
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