This Article Full Text Full Text (PDF) Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this link to a friend Similar articles in this journal Similar articles in PubMed Add article to my folders Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Tian, J Articles by Handa, J T Search for Related Content PubMed PubMed Citation Articles by Tian, J Articles by Handa, J T

The expression of native and cultured human retinal pigment epithelial cells grown in different culture conditions

¹ Michael Panitch Macular Degeneration Research Laboratory, Wilmer Ophthalmological Institute, Johns Hopkins Medical Institutes, Baltimore, MD, USA
² Departments of Ophthalmology and Molecular and Cellular Biology, University of California, Davis, CA, USA

Correspondence to:
Correspondence to:
James T Handa
MD; 3-109 Jefferson Street Building; Johns Hopkins Hospital; 600 N Wolfe Street; Baltimore, MD 21287, USA; jthanda{at}jhmi.edu

Aim: To determine the transcriptional proximity of retinal pigment epithelium (RPE) cells grown under different culture conditions and native RPE.

Methods: ARPE-19 cells were grown under five conditions in 10% CO₂: "subconfluent" in DMEM/F12 + 10% FBS, "confluent" in serum and serum withdrawn, and "differentiated" for 2.5 months in serum and serum withdrawn medium. Native RPE was laser microdissected. Total RNA was extracted, reverse transcribed, and radiolabelled probes were hybridised to an array containing 5353 genes. Arrays were evaluated by hierarchical cluster analysis and significance analysis of microarrays.

Results: 78% of genes were expressed by native RPE while 45.3–47.7% were expressed by ARPE-19 cells, depending on culture condition. While the most abundant genes were expressed by native and cultured cells, significant differences in low abundance genes were seen. Hierarchical cluster analysis showed that confluent and differentiated, serum withdrawn cultures clustered closest to native RPE, and that serum segregated cultured cells from native RPE. The number of differentially expressed genes and their function, and profile of expressed and unexpressed genes, demonstrate differences between native and cultured cells.

Conclusions: While ARPE-19 cells have significant value for studying RPE behaviour, investigators must be aware of how culture conditions can influence the mRNA phenotype of the cell.

Abbreviations: BSA, bovine serum albumin; CS, confluent serum; CSW, confluent, serum withdrawn; DMEM, Dulbecco’s Modified Eagle medium; DS, differentiated serum; DSW, differentiated serum withdrawn; FBS, fetal bovine serum; FDR, false discovery rate; RPE, retinal pigment epithelium

Keywords: cell culture density; laser capture microdissection; microarray analysis; retinal pigment epithelium; serum

This article has been cited by other articles:

				D. M. Gamm, J. N. Melvan, R. L. Shearer, I. Pinilla, G. Sabat, C. N. Svendsen, and L. S. Wright A Novel Serum-Free Method for Culturing Human Prenatal Retinal Pigment Epithelial Cells Invest. Ophthalmol. Vis. Sci., February 1, 2008; 49(2): 788 - 799. [Abstract] [Full Text] [PDF]

				Y. F. Leung, P. Ma, and J. E. Dowling Gene Expression Profiling of Zebrafish Embryonic Retinal Pigment Epithelium In Vivo Invest. Ophthalmol. Vis. Sci., February 1, 2007; 48(2): 881 - 890. [Abstract] [Full Text] [PDF]

				A. Maminishkis, S. Chen, S. Jalickee, T. Banzon, G. Shi, F. E. Wang, T. Ehalt, J. A. Hammer, and S. S. Miller Confluent monolayers of cultured human fetal retinal pigment epithelium exhibit morphology and physiology of native tissue. Invest. Ophthalmol. Vis. Sci., August 1, 2006; 47(8): 3612 - 3624. [Abstract] [Full Text] [PDF]

				Y. Luo, Y. Zhuo, M. Fukuhara, and L. J. Rizzolo Effects of Culture Conditions on Heterogeneity and the Apical Junctional Complex of the ARPE-19 Cell Line. Invest. Ophthalmol. Vis. Sci., August 1, 2006; 47(8): 3644 - 3655. [Abstract] [Full Text] [PDF]