Effect of quiescence on integrin alpha5beta1 expression in human retinal pigment epithelium.
OBJECTIVE
The retinal pigment epithelium (RPE) is differentiated and mitotically inactive in the normal eye, but several pathologies such as proliferative vitreoretinopathy (PVR) cause RPE cells to dedifferentiate and resume proliferation. Integrins, a family of cell surface glycoproteins that mediate cell proliferation and differentiation, are thought to play fundamental roles in PVR. The aim of this study was to evaluate protein expression and gene regulation of the integrin alpha5 subunit in proliferating and quiescent RPE cells.


METHODS
Protein expression was studied in situ by immunohistochemistry and in vitro at different cell confluences by immunoprecipitation. Semi-quantitative RT-PCR and transient transfections were used to determine whether increasing cell confluence also affected alpha5 subunit mRNA levels and promoter activity, respectively.


RESULTS
We demonstrated that the integrin alpha5 subunit is present at the RPE cell surface both in situ and in vitro, and that alpha5 protein level is influenced by confluence. Levels of integrin alpha5 transcripts are similar for sub-confluent and confluent cells, and a small increase in the promoter activity was observed between sub-confluent and confluent cells. However, both the integrin alpha5 subunit transcript and the alpha5 promoter activity decreased when cells reached post-confluence.


CONCLUSIONS
We demonstrated that cell confluence affected protein and gene expression of the integrin alpha5 subunit. Proliferating RPE cells expressed high levels of both the alpha5 protein and mRNA transcripts and showed a high promoter activity. However, when cells reached quiescence, alpha5 gene expression was substantially reduced and RPE cells expressed little alpha5 protein at their cell surface.
Proulx
Stephanie
S
Université du Québec à Trois-Rivières, Département Chimie-Biologie, Trois-Rivières, Québec, Canada. Unité de Recherche en Ophtalmologie, Centre de Recherche du Centre Hospitalier de l'Université Laval, Ste-Foy, Québec, Canada
Guérin
Sylvain L
SL
Salesse
Christian
C
D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002452:Cell Count; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D002648:Child; D002675:Child, Preschool; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D039081:Integrin alpha5beta1 / Q000235:genetics* / Q000378:metabolism*; D008875:Middle Aged; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D039081:Integrin alpha5beta1; D012333:RNA, Messenger