Senior Scholars Program

Title

Polyplex-mediated gene transfer into human retinal pigment epithelial cells in vitro

UMMS Affiliation

Department of Medicine, Division of Ophthalmology; Department of Cell Biology

Date

12-1-1999

Document Type

Article

Medical Subject Headings

DNA; Gene Expression; *Gene Transfer Techniques; Humans; In Vitro Techniques; Pigment Epithelium of Eye; Transgenes

Disciplines

Biochemistry | Cell Biology | Eye Diseases | Ophthalmology

Abstract

The human retinal pigment epithelium (RPE) is a potential target tissue for directed transfer of candidate genes to treat age-related macular degeneration (AMD). The RPE is uniquely suited to gene therapy protocols that use liposome-mediated DNA transfer because of its high intrinsic phagocytic function in vivo. In these studies, we examined the efficacy of human RPE cell uptake and expression of the green fluorescent protein (GFP) and neomycin resistance marker genes by polyplex-mediated gene transfer in vitro. The effects of varying DNA and polyplex concentration and ratios on GFP transgene expression were examined. A narrow range of experimental conditions were found to maximize transgene expression; most important were the DNA concentration and the DNA:polyplex ratio. The transfection efficiency for human RPE cells was reproducibly 20% in vitro by this method and reached a maximum level of expression after 48 h. There was a rapid decline in gene expression over 2 weeks following polyplex-mediated gene transfer, but stable integration does occur at low frequencies with and without selection.

Rights and Permissions

Citation: J Cell Biochem. 1999 Nov;76(1):153-60. DOI:10.1002/(SICI)1097-4644(20000101)76:1%3C153::AID-JCB15%3E3.0.CO;2-%23

Related Resources

Link to Article in PubMed

Comments

Medical student Mark P. Hatton participated in this study as part of the Senior Scholars research program at the University of Massachusetts Medical School.

PubMed ID

10581009