Retinal pigment epithelium rescues vascular endothelium from retinoic acid-induced apoptosis.
OBJECTIVE
To determine whether retinoids are capable of inducing vascular endothelial cell apoptosis and whether the presence of an intact RPE monolayer can block retinoid-induced vascular endothelial cell death.


METHODS
Confluent fetal bovine aortic endothelial (FBAE) cells were incubated with various concentrations of all-trans or 9-cis retinoic acid (an analogue of 11-cis retinoic acid). Apoptosis rates were determined at 24 hours, and the effect of inhibition of protein synthesis and activation of protein kinase C on apoptosis was investigated by supplying culture medium with 0.1 mg/mL cycloheximide and 10 nM phorbol myristate acetate. To investigate the impact of RPE on retinoid-induced apoptosis, confluent FBAE cells were cultured with a confluent layer of RPE in inserts where retinoids were added to the upper compartment. A confluent bovine corneal endothelium monolayer was used as the control. The permeabilities of the RPE and bovine corneal endothelium monolayers to fluorescein (20 microg/mL) and 9-cis retinoic acid (3 x 10(-4) M) were also determined.


RESULTS
9-cis Retinoic acid induced higher rates of apoptosis in FBAE cells than did all-trans retinoic acid and the control (P = 0.004). This effect was dose-dependent, with an ED(50) of 1.4 microM (r = 0.99, P = 0.004). Cycloheximide did not inhibit 9-cis retinoic acid-induced apoptosis, but phorbol myristate acetate significantly decreased the apoptosis rate (P = 0.005). The presence of a confluent RPE monolayer reduced the 9-cis retinoic acid-induced apoptosis rate (P = 0.002), but the presence of a bovine corneal endothelial monolayer did not (P > 0.05). Both cell types established a similar diffusion barrier against fluorescein and 9-cis retinoic acid.


CONCLUSIONS
9-cis Retinoic acid is an important mediator of vascular endothelial apoptosis. A confluent monolayer of RPE can prevent endothelial cell apoptosis, and this effect is not due simply to establishment of a diffusion barrier by the RPE.
Tezel
Tongalp H
TH
Department of Ophthalmology and Visual Sciences, Kentucky Lions Eye Center, University of Louisville School of Medicine, 301 E. Muhammad Ali Boulevard, Louisville, KY 40202, USA. tongalp.tezel@louisville.edu
Geng
Lijun
L
Kaplan
Henry J
HJ
Del Priore
Lucian V
LV
D000328:Adult; D000368:Aged; D000077556:Alitretinoin; D000818:Animals; D001013:Aorta, Thoracic / Q000166:cytology; D017209:Apoptosis / Q000187:drug effects*; D002417:Cattle; D018929:Cell Culture Techniques; D002470:Cell Survival / Q000502:physiology; D018920:Coculture Techniques; D003513:Cycloheximide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D008856:Microscopy, Fluorescence; D010857:Pigment Epithelium of Eye / Q000502:physiology*; D011493:Protein Kinase C / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014212:Tretinoin / Q000494:pharmacology*
D000077556:Alitretinoin; D014212:Tretinoin; D003513:Cycloheximide; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate