Haru Okamoto

Molecular composition of drusen and possible involvement of anti-retinal autoimmunity in two different forms of macular degeneration in cynomolgus monkey (Macaca fascicularis)

We have previously reported a cynomolgus monkey (Macaca fascicularis) pedigree with early onset macular degeneration that develops drusen at 2 yr after birth (1). In this study, the molecular composition of drusen in monkeys affected with late onset and early onset macular degeneration was both characterized. Involvement of anti‐retinalautoimmunity in the deposition of drusen and the pathogenesis of the disease was also evaluated. Funduscopic and histological examinations were performed on 278 adult monkeys (mean age=16.94 yr) for late onset macular degeneration. The molecular composition of drusen was analyzed by immunohistochemistry and/or direct proteome analysis using liquid chromatography tandem mass spectroscopy (LC‐MS/MS). Anti‐retinal autoantibodies in sera were screened in 20 affected and 10 age‐matched control monkeys by Western blot techniques. Immunogenic molecules were identified by 2D electrophoresis and LC‐MS/MS. Relative antibody titer against each antigen was determined by ELISA in sera from 42 affected (late onset) and 41 normal monkeys. Yellowish‐white spots in the macular region were observed in 90 (32%) of the late onset monkeys that were examined. Histological examination demonstrated that drusen or degenerative retinal pigment epithelium (RPE) cells were associated with the pigmentary abnormalities. Drusen in both late and early onset monkeys showed immunoreactivities for apolipoprotein E, amyloid P component, complement component C5, the terminal C5b‐9 complement complex, vitronectin, and membrane cofactor protein. LC‐MS/MS analyses identified 60 proteins as constituents of drusen, including a number of common components in drusen of human age‐related macular degeneration (AMD), such as annexins, crystallins, immunoglobulins, and complement components. Half of the affected monkeys had single or multiple autoantibodies against 38, 40, 50, and 60 kDa retinal proteins. The reacting antigens of 38 and 40 kDa were identified as annexin II and μ‐crystallin, respectively. Relative antibody titer against annexin II in affected monkeys was significantly higher than control animals (P<0.01). Significant difference was not observed in antibody titer against μ‐crystallin; however, several affected monkeys showed considerably elevated titer (360–610%) compared with the mean for unaffected animals. Monkey drusen both in late and early onset forms of macular degeneration had common components with drusen in human AMD patients, indicating that chronic inflammation mediated by complement activation might also be involved in the formation of drusen in these affected monkeys. The high prevalence of anti‐retinalautoantibodies in sera from affected monkeys demonstrated an autoimmune aspect of the pathogenesis of the disease. Although further analyses are required to determine whether and how autoantibodies against annexin II or μ‐crystallin relate to the pathogenesis of the disease, it could be hypothesized that immune responses directed against these antigens might trigger chronic activation of the complement cascade at the site of drusen formation.

Dominant Mutations in RP1L1 Are Responsible for Occult Macular Dystrophy

Occult macular dystrophy (OMD) is an inherited macular dystrophy characterized by progressive loss of macular function but normal ophthalmoscopic appearance. Typical OMD is characterized by a central cone dysfunction leading to a loss of vision despite normal ophthalmoscopic appearance, normal fluorescein angiography, and normal full-field electroretinogram (ERGs), but the amplitudes of the focal macular ERGs and multifocal ERGs are significantly reduced at the central retina. Linkage analysis of two OMD families was performed by the SNP High Throughput Linkage analysis system (SNP HiTLink), localizing the disease locus to chromosome 8p22-p23. Among the 128 genes in the linkage region, 22 genes were expressed in the retina, and four candidate genes were selected. No mutations were found in the first three candidate genes, methionine sulfoxide reductase A (MSRA), GATA binding 4 (GATA4), and pericentriolar material 1 (PCM1). However, amino acid substitution of p.Arg45Trp in retinitis pigmentosa 1-like 1 (RP1L1) was found in three OMD families and p.Trp960Arg in a remaining OMD family. These two mutations were detected in all affected individuals but in none of the 876 controls. Immunohistochemistry of RP1L1 in the retina section of cynomolgus monkey revealed expression in the rod and cone photoreceptor, supporting a role of RP1L1 in the photoreceptors that, when disrupted by mutation, leads to OMD. Identification of RP1L1 mutations as causative for OMD has potentially broader implications for understanding the differential cone photoreceptor functions in the fovea and the peripheral retina.

Genetic analysis of typical wet-type age-related macular degeneration and polypoidal choroidal vasculopathy in Japanese population

Age-related macular degeneration (AMD) is a common cause of blindness in the elderly. Caucasian patients are predominantly affected by the dry form of AMD, whereas Japanese patients have predominantly the wet form of AMD and/or polypoidal choroidal vasculopathy (PCV). Although genetic association in the 10q26 (ARMS2/HTRA1) region has been established in many ethnic groups for dry-type AMD, typical wet-type AMD, and PCV, the contribution of the 1q32 (CFH) region seem to differ among these groups. Here we show a single nucleotide polymorphism (SNP) in the ARMS2/HTRA1 locus is associated in the whole genome for Japanese typical wet-type AMD (rs10490924:

Complement factor H polymorphisms in Japanese population with age-related macular degeneration.

p = 4.1 \times 10 ^{ - 4}

Early-Onset Macular Degeneration with Drusen in a Cynomolgus Monkey (Macaca fascicularis) Pedigree: Exclusion of 13 Candidate Genes and Loci

, OR = 4.16) and PCV (rs10490924:

Proteomic and Transcriptomic Analyses of Retinal Pigment Epithelial Cells Exposed to REF-1/TFPI-2

p = 3.7 \times 10 ^{ -8}