Galectin-1 and -3 in high amounts inhibit angiogenic properties of human retinal microvascular endothelial cells in vitro

by Anna Hillenmayer, Christian M. Wertheimer, Arie Geerlof, Kirsten H. Eibl, Siegfried Priglinger, Claudia Priglinger, Andreas Ohlmann

Purpose

Galectin-1 and -3 are β-galactoside binding lectins with varying effects on angiogenesis and apoptosis. Since in retinal pigment epithelial cells high amounts of human recombinant galectin (hr-GAL)1 and 3 inhibit cell adhesion, migration and proliferation, we investigated if hr-GAL1 and 3 have homologous effects on human retinal microvascular endothelial cells (HRMEC) in vitro.

Methods

To investigate the effect of galectin-1 and -3 on HRMEC, proliferation, apoptosis and viability were analyzed after incubation with 30, 60 and 120 μg/ml hr-GAL1 or 3 by BrdU-ELISA, histone-DNA complex ELISA, live/dead staining and the WST-1 assay, respectively. Further on, a cell adhesion as well as tube formation assay were performed on galectin-treated HRMEC. Migration was investigated by the scratch migration assay and time-lapse microscopy. In addition, immunohistochemical staining on HRMEC for β-catenin, galectin-1 and -3 were performed and β-catenin expression was investigated by western blot analysis.

Results

Incubation with hr-GAL1 or 3 lead to a decrease in proliferation, migration, adhesion and tube formation of HRMEC compared to the untreated controls. No toxic effects of hr-GAL1 and 3 on HRMEC were detected. Intriguingly, after treatment of HRMEC with hr-GAL1 or 3, an activation of the proangiogenic Wnt/β-catenin signaling pathway was observed. However, incubation of HRMEC with hr-GAL1 or 3 drew intracellular galectin-1 and -3 out of the cells, respectively.

Conclusion

Exogenously added hr-GAL1 or 3 inhibit angiogenic properties of HRMEC in vitro, an effect that might be mediated via a loss of intracellular endogenous galectins.