title: Negative regulation of RPE cell attachment by carbohydrate-dependent cell surface binding of galectin-3 and inhibition of the ERK-MAPK pathway.
abstract: Adhesion and spreading of retinal pigment epithelial (RPE) cells on fibronectin-rich extracellular matrices is a crucial event in the pathogenesis of proliferative vitreoretinopathy (PVR). In the present study we explored the capacity of galectin-3, a β-galactoside-binding endogenous lectin, to inhibit early PVR-associated cellular events from a therapeutic perspective. We assessed the relative expression levels of galectin-3 in native RPE and dedifferentiated, cultured RPE. Galectin-3 was constitutively expressed under in vivo and in vitro conditions and was abundant in cultured cells. Treatment of human RPE cells with soluble galectin-3 disclosed no toxicity within control limits up to 250 μg/ml. When added to the medium, galectin-3 dose-dependently inhibited attachment and spreading of the cells on fibronectin by more than 75%. When coated on the plastic surface, galectin-3 alone impaired attachment and spreading of RPE cells, and reduced attachment but not spreading on fibronectin. Galectin-3 bound to the cell surface, and, as determined by the use of the competing sugar β-lactose, galectin-3-mediated effects were dependent on carbohydrate binding. To ascertain the role of the ability of galectin-3 to form pentamers, we proteolytically removed the N-terminal, cross-linking section. The remaining C-terminal carbohydrate-binding domain alone failed to bind to cells and was functionally inactive. These results emphasize the relevance of both properties, i.e., glycan-binding and cross-linking of glycan moieties, for the inhibitory activity of galectin-3. Incubation of mobilized RPE cells with galectin-3 significantly disturbed microfilament assembly and, in correlation with decreased attachment, inhibited ERK phosphorylation. Therefore, galectin-3, acting as a cross-linking lectin on the cell surface, negatively regulates attachment and spreading of RPE cells in vitro. This effect, at least in part, is attributed to an inhibition of the ERK-MAPK pathway, which prevents cytoskeletal rearrangements needed for RPE cell attachment and spreading. Further investigation at this pathway may disclose a promising nouveau perspective for treatment and prophylaxis of early PVR.
authors.affiliation: Department of Ophthalmology, Ludwig-Maximilians-University, Mathildenstrasse 8, Munich, Germany. siegfried.priglinger@med.uni-muenchen.de
authors.forename: Claudia S
authors.initials: CS
authors.lastname: Alge-Priglinger
authors.forename: Sabine
authors.initials: S
authors.lastname: André
authors.forename: Harald
authors.initials: H
authors.lastname: Schoeffl
authors.forename: Anselm
authors.initials: A
authors.lastname: Kampik
authors.forename: Rupert W
authors.initials: RW
authors.lastname: Strauss
authors.forename: Marcus
authors.initials: M
authors.lastname: Kernt
authors.forename: Hans-Joachim
authors.initials: HJ
authors.lastname: Gabius
authors.forename: Siegfried G
authors.initials: SG
authors.lastname: Priglinger
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000368:Aged; D001667:Binding, Competitive; D050260:Carbohydrate Metabolism; D002448:Cell Adhesion; D048430:Cell Shape; D002470:Cell Survival; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D005260:Female; D037483:Galectin 1 / Q000378:metabolism; D037502:Galectin 3 / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D020935:MAP Kinase Signaling System; D008297:Male; D008875:Middle Aged; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D055213:Retinal Pigment Epithelium / Q000166:cytology* / Q000201:enzymology / Q000378:metabolism*
chemical_list: D037483:Galectin 1; D037502:Galectin 3; D048049:Extracellular Signal-Regulated MAP Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases