Appropriately Differentiated ARPE-19 Cells Regain a Native Phenotype and Similar Gene Expression Profile

The retinal pigment epithelial (RPE) cell line ARPE-19 provides a widely-used alternative to native RPE. However, retention of the native RPE phenotype becomes problematic after multiple passages. We wished to determine if suitable culture conditions and differentiation could restore RPE-appropriate gene expression to ARPE-19. ARPE-19 cells at passages p9 to p12, grown in DMEM containing high glucose and pyruvate with 1% fetal bovine serum, were differentiated for up to 4 months. Using RNA-Seq, we compared the transcriptome of ARPE-19 cells kept in long-term culture with those cultured for 4 days. The 4 month cells developed the classic native RPE phenotype with heavy pigmentation. RNA-Seq analysis provided a comprehensive view of the relative abundance and differential expression of genes in the 4 month cells. Of the 16,757 genes with detectable signals, nearly 2435 genes were upregulated, and 931 genes were down-regulated with a fold change differences of 2 or more. Genes characteristic of RPE, including RPE65, RDH5 and RDH10, were greatly increased in ARPE-19 cells maintained at confluence for 4 months. Comparison with microarray data sets from human primary cell lines revealed important overall similarities in expression of "signature" genes. The results of this study demonstrate that ARPE-19 cells can express genes specific to native human RPE cells when appropriately cultured, and thus, can provide a relevant system to study differentiated cellular functions of RPE in vitro. Overall design: RNA-Seq profiles of ARPE-19 cells grown for 4 days or 4 months; triplicate replicates were sequenced.

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Samuel W, Jaworski C, Postnikova OA, Kutty RK, Duncan T, Tan LX, Poliakov E, Lakkaraju A, Redmond TM