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Zai-Long Chi, Yuko Katakai, Nobuhiro Shimozawa, Michihiro T Suzuki, Takeshi Iwata; Microarray Analysis Of RPE Cells Isolated From Cynomolgus Monkey With Early-onset Drusen Formation. Invest. Ophthalmol. Vis. Sci. 2011;52(14):964.
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© ARVO (1962-2015); The Authors (2016-present)
We have previously reported a cynomolgus (Macaca fascicularis) pedigree with early-onset drusen formation. These monkeys show cardinal features of early stage of age-related maculopathy (ARM) such as pigmentary disorders and/or drusen-like spots at two years after birth (Umeda et al., IOVS, FASEB J, 2005) with ERG abnormality. The RPE cells isolated from affected monkeys had significant reduction of cell proliferation, tight junction loss, and low phagocytosis activity (Chi et al., ARVO 2008, 2009). Here we report microarray analysis of retinal pigment epithelium (RPE) cells isolated from two affected monkeys in comparison with two controls.
Total RNAs were isolated from primary cultured RPE cells from normal and affected monkeys. The microarray analysis was performed using Rhesus Macaque (V2) Gene Expression Microarray (Agilent Technologies) and Filgen Array for Macaca fascicularis. The Agilent microarray contains over 45,000 sixty-mer oligonucleotide representing approximately 22,000 genes of known gene sequence. Filgen microarrays containing 12,535 oligonucleotide probes (EST). Deferentially expressed mRNAs and derived proteins were further confirmed by RT-PCR, flow cytometry, and immunohistochemistry analysis. The functions of these genes were also analyzed in laser-induced choroidal neovascularization (CNV) mice models, affected monkeys, and in vitro experiments.
mRNA expression in RPE cells significantly changed between disease severities. Expression of numerous chemokines, complement component factors, and other immune response related genes were increased in affected monkey RPE cells. These finding mimics the human ARM reported previously by others. Co-culture of fibroblast cells and endothelium cells (Toyobo) with supernatant of cultured affected RPE cells promoted tube-formation, while control RPE did not. Inhibition of proteasome and complement C3 reduced disease progression in animal models.
Significant change of gene expression leading to loss of tight junction and reduction of phagocytosis activity in RPE cells from affected monkeys. A new therapeutic strategy for ARM will be suggested at the meeting.
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