Involvement of EphB/ephrin-B signaling in axonal survival in mouse experimental glaucoma

Abstract

PURPOSE. To examine the functional significance of EphB/ephrin-B upregulation in mouse experimental glaucoma. METHODS. In a loss-of-function approach, mouse mutants lacking EphB2 (EphB2 -/-) or EphB3 (EphB3 -/-) protein, and mutants expressing EphB2 truncated in the C-terminus (EphB2 lacZ/lacZ) were subjected to laser-induced ocular hypertension (LIOH), an experimental mouse model of glaucoma. The number of optic nerve axons was counted in paraphenylenediamine (PPD)-stained sections and compared between EphB mutants and wild type littermates. In a gain-of-function approach, retina/optic nerve explants obtained from LIOHtreated animals were exposed to EphB2-Fc recombinant proteins or Fc control proteins. Tissue sections through the optic nerve head (ONH) were labeled with neuron-specific antitubulin β-III antibody to determine axonal integrity. RESULTS. Both EphB2 and EphB3 null mutant mice exhibited more severe axonal degeneration than wild type littermates after treatment with LIOH. Mutant mice in which the C-terminal portion of EphB2 is truncated had an intermediate phenotype. Application of EphB2-Fc recombinant protein to LIOHtreated optic nerve explants resulted in greater sparing of tubulin β-III- containing retinal ganglion cell (RGC) axons. CONCLUSIONS. These results provide genetic evidence in mice that both EphB/ephrin-B forward and reverse signaling feed into an endogenous pathway to moderate the effects of glaucomatous insult on RGC axons. LIOH-induced axon loss is maintained in retina/optic nerve explants after removal from an ocular hypertensive environment. Exogenous application of EphB2 protein enhances RGC axon survival in explants, suggesting that modulation of Eph/ephrin signaling may be of therapeutic interest. © 2012 The Association for Research in Vision and Ophthalmology, Inc.