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Michael Böhm, Karina Hadrian, Stefan Schlatt, Joachim Wistuba, Solon Thanos, Sven Rahmann, Ludger Klein-Hitpass; The age-related transcriptome of the retinal pigment epithelium of the macular-bearing Callithrix jacchus. Invest. Ophthalmol. Vis. Sci. 2019;60(9):2345.
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© ARVO (1962-2015); The Authors (2016-present)
Alterations of metabolic requirements within the macula compared to peripheral regions of the vitreoretinal compartment suspect differences in the aging process of the retinal pigment epithelium (RPE). The underlying molecular alterations of the RPE under topographical conditions during lifetime are not well known. The aim of this study was to analyze the transcriptome of the RPE of common marmosets Callithrix jacchus(C. jacchus) using Next-Generation-Sequencing (NGS).
The RPE was removed from eyecups of newborn (0-3 days), adult (10-22 months), and senile (96-108 months) C. jacchus (n=3 each). The RPE of the macula and peripheral regions were separated. 100 ng of total RNA was processed for RNA sequencing using the Ovation Human FFPE RNA-Seq Multiplex System Kit (NuGEN) and libraries were sequenced on an Illumina HiSeq 2500. The sequences were aligned to the genome of C. jacchus and quantified using the RPKM (Reads Per Kilobase Million) method. Significantly differentially expressed genes under topographical and age-related conditions were identified using the ANOVA test (p<0.05). Gene Set Enrichment Analysis (GSEA) was performed using gene sets derived from the Molecular Signatures Database (MsigDB, v6.0). Results of promising candidate genes were further verified using real-time quantitative PCR (qRT-PCR).
The transcriptome of the studied RPE revealed differences under topological and age-related conditions. The differences in mRNA levels are low in neonate samples and increase with age. GESA revealed genes regarding angiogenesis (e.g. LUM, STC1), inflammation (e.g. HMOX1), extracellular matrix remodeling (e.g. FBN1) and oxidative stress (e.g. PRDX) were found to be significantly altered in senile compared to neonate tissue. Validation of selected genes like HMOX1, CTSB and LUM using qRT-PCR confirm recent findings of NGS.
The presented study shows significant alterations of the transcriptome within the aging RPE in the macula compared to the periphery. The number of differentially expressed genes increases with age, which possibly reflects endorsed light exposures as well as the degenerative state of the tissue. Further studies area mandatory to expand the understanding of the aging process in the RPE and its progression to pathological conditions. Supported by the German Research Foundation (DFG; BO 4556/1-1).
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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