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Mol Vis. 2007 Jul 23;13:1259-73.

Analysis of the RPE transcriptome reveals dynamic changes during the development of the outer blood-retinal barrier.

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  • 1Department of Surgery, Yale University School of Medicine, New Haven, CT 06520-8062, USA. lawrence.rizzolo@yale.edu

Abstract

PURPOSE:

The morphology of the RPE shows minimal change as the neural retina and choriocapillaris differentiate. Nonetheless, initial studies of proteins related to the outer blood-retinal barrier suggest extensive remodeling of the retinal pigment epithelium (RPE) in response to this changing environment. A genomic approach was used to investigate the extent of this remodeling.

METHODS:

RPE was isolated from E7, E10, E14, and E18 chick embryos and total RNA extracted for probing the entire genome on Affymetrix microarray chips. Statistical parameters using ANOVA were adjusted to yield a theoretical false discovery rate of 5%. STEM software was used to cluster genes into statistically related patterns of expression. Gene ontology clustering, using Affymetrix software was used for functional clustering. The proteinlounge.com database was used as a source of known biological pathways.

RESULTS:

Of the 37,694 probesets on the microarray, 17,199 were absent. Of the 20,495 expressed probes, the expression of 8,889 was developmentally regulated. 4,814 of these could be clustered into 12 patterns of expression that were statistically significant. Minimal contamination by surrounding tissues was detected. The developmental patterns of 22 tight and adherens junction proteins were compared using hybridization to the microarray and quantitative PCR. Only two showed small variations from the patterns revealed by the microarray. The data indicate extensive remodeling of the extracellular matrix, cell surface receptors, cell-cell junctions, transcellular ion transport, and signal transduction pathways throughout development. Notably, the appearance of the mRNAs for claudin 20, ZO-3, and cadherins 13 and 20 very late in development suggest barrier properties continue to change after functional junctions are formed.

CONCLUSIONS:

The data reveal a far more dynamic view of the RPE and its interactions with its environment than would be expected from morphological examination. The remodeling of junctional complexes, extracellular matrix interactions and transcellular transport capabilities indicates a continuous remodeling of the blood-retinal barrier as the retina develops. These data provide a standard whereby culture models of RPE function and regulation may be judged.

PMID:
17679949
[PubMed - indexed for MEDLINE]
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