Role of inhibitors of isoprenylation in proliferation, phenotype and apoptosis of human retinal pigment epithelium.
BACKGROUND
Isoprenoid biosynthesis is known to be essential for diverse cellular functions, including cell proliferation. The aim of this work was to study the effects caused by the addition of different inhibitors of isoprenylation (lovastatin, manumycin A, farnesyltransferase inhibitor III and N-acetyl-S-farnesyl-L-cysteine) to human retinal pigment epithelium (RPE) in culture, as potential coadjunctive-to-surgery treatments applicable to proliferative vitreoretinopathy.


METHODS
Human RPE cell cultures were established from adult corneal donors. Proliferation levels were evaluated using the incorporation of 5-bromo-2'-deoxyuridine into the DNA. Cell viability was measured by tetrazolium bromide transformation. Apoptosis was determined by DNA fragmentation assay, TdT-mediated d-UTP-X nick-end labeling (TUNEL) and phosphatidylserine exposure assessment. Changes in cell morphology and actin cytoskeleton were evaluated using a phase-contrast microscope and by fluorescent staining of actin cables with TRITC-phalloidin.


RESULTS
We found that lovastatin showed an important antiproliferative effect on human RPE cells in culture. This effect was clearly dose-dependent, and adding mevalonate could reverse it. We also found that lovastatin induced changes in the distribution of actin cytoskeleton and, finally, that it also induced RPE apoptosis. Manumycin A, farnesyltransferase inhibitor III and N-acetyl-S-farnesyl-L-cysteine also showed antiproliferative effects in RPE. However, they do not have any effect on cell morphology or induction of apoptosis.


CONCLUSIONS
We identified various effects of lovastatin on human RPE cultures: inhibition of cell proliferation, modifications of the phenotype and induction of apoptosis. Interestingly, the addition of different inhibitors of protein isoprenylation only affected the proliferation of the cells. There was no evidence that isoprenylated proteins inhibition is related to lovastatin-induced RPE apoptosis.
Capeáns
C
C
Instituto Gallego de Oftalmología (INGO), Santiago de Compostela, Spain.
Piñeiro
A
A
Pardo
M
M
Carneiro
C
C
Blanco
M J
MJ
Viñuela
J M
JM
Sánchez Salorio
M
M
Domínguez
F
F
D000199:Actins / Q000378:metabolism; D000328:Adult; D019883:Alkyl and Aryl Transferases / Q000037:antagonists & inhibitors; D017209:Apoptosis / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects*; D002478:Cells, Cultured; D004261:DNA Replication; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D051231:Farnesyltranstransferase; D006801:Humans; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors / Q000494:pharmacology*; D020287:In Situ Nick-End Labeling; D008148:Lovastatin / Q000494:pharmacology*; D010641:Phenotype; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D011090:Polyenes / Q000494:pharmacology; D053284:Polyunsaturated Alkamides; D017368:Protein Prenylation / Q000187:drug effects*
D000199:Actins; D004791:Enzyme Inhibitors; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D011090:Polyenes; D053284:Polyunsaturated Alkamides; D008148:Lovastatin; D019883:Alkyl and Aryl Transferases; D051231:Farnesyltranstransferase; C054474:manumycin