Inhibition of retinal pigment epithelial cell-induced tractional retinal detachment by disintegrins, a group of Arg-Gly-Asp-containing peptides from viper venom.
OBJECTIVE
Integrin-mediated extracellular matrix (ECM) attachment plays an important role in vitreous contraction of retinal pigment epithelial (RPE) cells. Disintegrins, a group of Arg-Gly-Asp (RGD)-containing peptides from viper venom, are potential anti-adhesion agents that interfere with integrin-ECM binding. This study was performed to determine whether disintegrins were effective in inhibiting RPE cell-induced matrix attachment in vitro and tractional retinal detachment in a rabbit model in vivo.


METHODS
Two disintegrins, echistatin from viper Echis carinatus and flavoridin from Trimeresurus flavoviridis, were used. The expression of integrins on the surface of bovine and rabbit RPE cells was examined by indirect immunofluorescent stain with specific anti-integrin monoclonal antibodies. The inhibitory effect of disintegrins on RPE cell-mediated ECM attachment and vitreous contraction was evaluated with cell adhesion and vitreous contraction assays. In the in vivo model, rabbit eyes were injected intravitreously with either homologous rabbit RPE cells alone or together with disintegrins to induce tractional retinal detachment. The cytotoxic effect of disintegrins was examined with a cell proliferation assay using the alamar blue method. Retinal toxicity of disintegrins was evaluated with electroretinograms and histologic examination of the rabbit eyes.


RESULTS
Bovine and rabbit RPE cells showed the positive staining for the integrins alpha 2 beta 1 and alpha 5 beta 1 on cell surface. Disintegrins, echistatin, and flavoridin inhibited RPE cell attachment to the ECM. The potency of disintegrins was 150 to 300 times higher than that of Gly-Arg-Gly-Asp-Ser (GRGDS) peptide. The disintegrins also inhibited RPE cell-induced vitreous contraction in a dose-dependent manner, whereas the GRGDS peptide had no effect. In the in vivo experiment, echistatin (50 microgram/ml) or flavoridin (80 microgram/ml) significantly inhibited RPE cell-induced tractional retinal detachment compared with the control group at week 2 (P< 0.05) and week 4 (P< 0.01) after surgery. Disintegrins were nontoxic to RPE cells and rabbit retina as evaluated by cytotoxicity tests, electroretinograms, and histologic examinations.


CONCLUSIONS
The disintegrins were effective in inhibiting RPE cell attachment to the ECM and vitreous contraction in vitro. They also were effective in suppressing RPE cell-induced tractional retinal detachment in the rabbit eyes. They were nontoxic. Disintegrins and their analogs might be potential anti-adhesion therapeutic agents in the treatment of proliferative vitreoretinopathy.
Yang
C H
CH
Department of Ophthalmology, National Taiwan University Hospital, Taipei, Taiwan.
Huang
T F
TF
Liu
K R
KR
Chen
M S
MS
Hung
P T
PT
D000595:Amino Acid Sequence; D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology; D002417:Cattle; D002448:Cell Adhesion / Q000187:drug effects; D002478:Cells, Cultured; D003435:Crotalid Venoms; D004305:Dose-Response Relationship, Drug; D005109:Extracellular Matrix / Q000378:metabolism*; D019084:Fluorescent Antibody Technique, Indirect; D016023:Integrins / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry / Q000494:pharmacology*; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology; D010857:Pigment Epithelium of Eye / Q000378:metabolism*; D010975:Platelet Aggregation Inhibitors / Q000737:chemistry / Q000494:pharmacology; D011817:Rabbits; D012163:Retinal Detachment / Q000209:etiology / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D014757:Viper Venoms / Q000737:chemistry / Q000494:pharmacology*; D018630:Vitreoretinopathy, Proliferative / Q000209:etiology / Q000473:pathology; D014822:Vitreous Body / Q000378:metabolism
D000970:Antineoplastic Agents; D003435:Crotalid Venoms; D016023:Integrins; D036341:Intercellular Signaling Peptides and Proteins; D009842:Oligopeptides; D010455:Peptides; D010975:Platelet Aggregation Inhibitors; D014757:Viper Venoms; C057986:echistatin; C069044:flavoridin protein, Trimeresurus; C047981:arginyl-glycyl-aspartic acid; C049554:glycyl-arginyl-glycyl-aspartyl-serine