Changes in gene expression of ARPE-19 cells in response to vitreous treatment.
OBJECTIVE
The purpose of this study is to determine the changes in gene expression by a human retinal pigment epithelium (RPE) cell line (ARPE-19) in response to vitreous treatment, which induces RPE proliferation and phenotypic changes in vitro that mimic the repair response observed in vivo during proliferative vitreoretinopathy.


METHODS
ARPE-19 cells were grown for more than 4 weeks and their gene expression studied in: (1) subconfluent cultures treated with 50% human vitreous for 72 h; (2) subconfluent cultures without vitreous treatment, and (3) a confluent, nondividing monolayer. Total RNA was extracted from RPE cells and differential gene expression between each condition was determined using gene arrays (Clontech, Palo Alto, Calif., USA). Semiquantitative RT-PCR was used to confirm the upregulation of 4 genes related to vitreous treatment. In addition, the secretion of 1 of these upregulated gene products, monocyte chemotactic protein 1 (MCP-1), was confirmed by ELISA.


RESULTS
A greater than threefold increase in the expression of mRNA for cell cycle regulators, intracellular transducers, cell adhesion proteins, growth factors and chemokines was observed following vitreous treatment of the RPE cell line and a corresponding decrease in the expression of genes related to apoptosis. RT-PCR confirmed the increased gene expression of MCP-1, intercellular adhesion molecule-1, vascular cell adhesion protein 1 precursor and vascular endothelial growth factor in vitreous-treated cells. Immunoassays further showed an increased MCP-1 secretion by vitreous-treated RPE.


CONCLUSIONS
These findings suggest that in this in vitro vitreous treatment model, ARPE-19 cells participate in a mock repair response by upregulating genes encoding for proteins associated with inflammation and wound healing.
Fan
Wei
W
Department of Ophthalmology & Visual Sciences, Kentucky Lions Eye Center, University of Louisville School of Medicine, Louisville, KY 40202, USA.
Zheng
Jing Juan
JJ
Peiper
Stephen C
SC
McLaughlin
Barbara J
BJ
D002460:Cell Line; D002478:Cells, Cultured; D018932:Chemokine CCL2 / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D015536:Down-Regulation; D005136:Eye Proteins / Q000502:physiology; D015870:Gene Expression / Q000502:physiology*; D020869:Gene Expression Profiling; D006801:Humans; D010857:Pigment Epithelium of Eye / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D015854:Up-Regulation; D014822:Vitreous Body / Q000502:physiology*
D018932:Chemokine CCL2; D017931:DNA Primers; D005136:Eye Proteins; D012333:RNA, Messenger