In vivo gene transfer as a means to study the physiology and morphogenesis of the retinal pigment epithelium in the rat

Alan D Marmorstein, Neal S. Peachey, Karl G. Csaky

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Our understanding of the morphogenesis of epithelial phenotypes has been greatly advanced by the use of in vitro cell culture systems. However, cell cultures often do not faithfully reconstitute many of the differentiated properties of the cell from which they are derived and cannot be used to examine complex physiologic interactions between adjacent tissues. This is particularly true of the retinal pigment epithelium (RPE). Many plasma membrane proteins, in vivo, exhibit a reversed polarity with respect to other epithelia, and RPE-derived cell lines seldom exhibit these same polarity properties. Furthermore, the interaction between the RPE cell and the neuorsensory retina, or the underlying blood supply, the choroid, is absent in cell culture. Most epithelia are difficult to isolate and study in vivo. The RPE is an exception to this. We have explored several aspects of RPE protein transport properties, vision-related physiology, and disease-related pathophysiology in the eye using in vivo gene transfer and electrophysiologic techniques. By injecting replication-defective adenoviruses into the subretinal space of rat eyes, we have been able to easily direct the expression of a test protein and follow its sorting and physiologic effects on RPE cells and adjacent tissues. Due to binding and internalization of adenoviral vectors to integrins found on the RPE apical plasma membrane, expression in a healthy eye is essentially confined to the RPE cell, even under control of a cytomegalovirus promotor. The use of varying amounts of adenoviral vector allows for determination of dose-responsive effects and the comparison of multiple mutants of a protein. In addition, there are substantial savings with respect to time and money in comparison to standard transgenic approaches.

Original languageEnglish (US)
Pages (from-to)277-285
Number of pages9
JournalMethods
Volume30
Issue number3
DOIs
StatePublished - Jul 1 2003
Externally publishedYes

Fingerprint

Gene transfer
Retinal Pigments
Retinal Pigment Epithelium
Physiology
Morphogenesis
Rats
Genes
Cell culture
Cell Culture Techniques
Cell membranes
Epithelium
Cell Membrane
Tissue
Gene Transfer Techniques
Proteins
Choroid
Protein Transport
Mutant Proteins
Cytomegalovirus
Sorting

Keywords

  • Adenovirus
  • Electroretinogram
  • Gene therapy
  • Photoreceptor
  • Retina
  • Retinal degeneration
  • Retinal pigment epithelium

ASJC Scopus subject areas

  • Molecular Biology

Cite this

In vivo gene transfer as a means to study the physiology and morphogenesis of the retinal pigment epithelium in the rat. / Marmorstein, Alan D; Peachey, Neal S.; Csaky, Karl G.

In: Methods, Vol. 30, No. 3, 01.07.2003, p. 277-285.

Research output: Contribution to journalArticle

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