Inhibition of RPE cell-mediated matrix adhesion and collagen gel contraction by crovidisin, a collagen-binding snake venom protein.
OBJECTIVE
Cell-mediated collagen gel contraction plays an important role in the pathogenesis of proliferative vitreoretinopathy (PVR). Anti-adhesion therapy has been suggested as a promising strategy in the treatment of PVR. Crovidisin, a snake venom protein isolated from Crotalus viridis, has been shown to bind selectively to collagen and to inhibit collagen-induced platelet aggregation. In the present study, the effectiveness of crovidisin in inhibiting the attachment of retinal pigment epithelial (RPE) cells to collagen, and RPE cell-mediated collagen gel contraction, was evaluated.


METHODS
Fluorescein isothiocyanate (FITC)-conjugated crovidisin was prepared and used to evaluate its binding affinity for collagen type I, fibronectin, vitronectin, and laminin. The inhibitory effect of crovidisin on RPE cell-mediated extracellular matrix attachment and collagen gel contraction was evaluated by cell adhesion and type I collagen gel contraction assays. The cytotoxic effect of crovidisin was examined with a cell proliferation assay, using the Alamar blue method. Flavoridin, an Arg-Gly-Asp-containing peptide from viper venom, was used for comparison.


RESULTS
FITC-conjugated crovidisin bound selectively to collagen type I with high affinity. It did not bind to other matrix proteins, including fibronectin, vitronectin and laminin, nor to RPE cells. Crovidisin inhibited RPE cell attachment to type I collagen in a dose-dependent manner. This inhibitory effect was enhanced by the presence of flavoridin. Crovidisin also dose-dependently inhibited RPE cell-mediated type I collagen gel contraction. Crovidisin was non-toxic to RPE cells.


CONCLUSIONS
Crovidisin, a snake venom-derived collagen-binding protein, possessing an inhibitory activity on RPE cell-collagen interaction and RPE cell-mediated collagen gel contraction, may be a useful tool for studying cell-collagen interaction, and a potential anti-adhesion therapeutic agent for ocular disorders in which cell-collagen interaction in involved, such as PVR.
Yang
C H
CH
Department of Ophthalmology, National Taiwan University Hospital, Taipei, Taiwan.
Liu
C Z
CZ
Huang
T F
TF
Yang
C M
CM
Lui
K R
KR
Chen
M S
MS
Hung
P T
PT
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism / Q000494:pharmacology*; D002417:Cattle; D002448:Cell Adhesion / Q000187:drug effects*; D002455:Cell Division; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism*; D004396:Coloring Agents; D003435:Crotalid Venoms / Q000378:metabolism / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005109:Extracellular Matrix / Q000378:metabolism*; D016650:Fluorescein-5-isothiocyanate; D036341:Intercellular Signaling Peptides and Proteins; D045726:Metalloproteases; D010078:Oxazines; D010455:Peptides / Q000494:pharmacology; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000378:metabolism*; D010975:Platelet Aggregation Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D030162:Reptilian Proteins; D014966:Xanthenes
D002352:Carrier Proteins; D004396:Coloring Agents; D003435:Crotalid Venoms; D036341:Intercellular Signaling Peptides and Proteins; D010078:Oxazines; D010455:Peptides; D010975:Platelet Aggregation Inhibitors; D030162:Reptilian Proteins; D014966:Xanthenes; C069044:flavoridin protein, Trimeresurus; C005843:resazurin; D003094:Collagen; D045726:Metalloproteases; C103975:crovidisin protein, Crotalus viridis; D016650:Fluorescein-5-isothiocyanate