Gene expression profile of cultured adult compared to immortalized human RPE.
OBJECTIVE
ARPE19 is a spontaneously immortalized cell line of human retinal pigment epithelium (RPE) that is used widely to draw inferences about the behavior of adult human RPE (ahRPE). We used DNA microarray analysis to compare the gene expression profiles of these two cell types.


METHODS
Second-passage cultured ahRPE from four human donors (age range 48-82 years) and ARPE19 cultured to confluence in five dishes were used for this DNA microarray study. Total RNA was isolated and first- and second-strand complimentary DNA was synthesized using standard techniques. Biotin-labeled antisense complimentary RNA was produced by an in vitro transcription reaction. Target hybridization, washing, staining, and scanning probe arrays were done following an Affymetrix GeneChip Expression Analysis Manual. Microarray data were normalized and statistical techniques were used to determine the presence or absence of expression of individual genes within ARPE19 and ahRPE, and their relative expression levels.


RESULTS
Hierarchic clustering analysis demonstrated that the gene expression profile of ahRPE and ARPE19 samples cluster into two distinct groups with no discernable overlap. The expression of 5,634+/-65 gene probes (out of 12,600 on microarray Human U95Av2 chip) was detected in ARPE19 cells compared to 5,580+/-84 genes in ahRPE cells from four human donor eyes. Thirty-five genes are expressed exclusively in ahRPE and nine genes exclusively in ARPE19 cells. Fifty additional genes have a threefold increase and 40 genes have a threefold decrease in expression level in ahRPE compared to ARPE19. There was no clear difference in the global expression level of genes known to be related to phagocytosis, angiogenesis, or apoptosis.


CONCLUSIONS
There are significant differences in the gene expression profile of ahRPE compared to ARPE19, and with some genes exclusively being expressed in one group and other genes being upregulated or downregulated by threefold. Caution should be exercised when generalizing results obtained from ARPE19 to the behavior of ahRPE.
Cai
Hui
H
Department of Ophthalmology, Harkness Eye Institute, Columbia University, New York, NY, USA.
Del Priore
Lucian V
LV
D000368:Aged; D000369:Aged, 80 and over; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D016000:Cluster Analysis; D015536:Down-Regulation; D005260:Female; D020869:Gene Expression Profiling; D006801:Humans; D008297:Male; D008875:Middle Aged; D020411:Oligonucleotide Array Sequence Analysis; D010857:Pigment Epithelium of Eye / Q000378:metabolism*; D015854:Up-Regulation