Abstract
It was previously shown that EphB/ephrinB reverse signaling in retinal ganglion cells (RGCs) is activated and involved in RGC apoptosis in a rat chronic ocular hypertension (COH) model. In the present work, we first show that ephrinB/EphB forward signaling was activated in COH retinas, and RGC apoptosis in COH retinas was reduced by PP2, an inhibitor of ephrinB/EphB forward signaling. We further demonstrate that treatment of cultured Müller cells with ephrinB1-Fc, an EphB1 activator, or intravitreal injection of ephrinB1-Fc in normal rats induced an increase in phosphorylated EphB levels in these cells, indicating the activation of ephrinB/EphB forward signaling, similar to those in COH retinas. The ephrinB1-Fc treatment did not induce Müller cell gliosis, as evidenced by unchanged GFAP expression, but significantly up-regulated mRNA and protein levels of tumor necrosis factor-α (TNF-α) in Müller cells, thereby promoting RGC apoptosis. Production of TNF-α induced by the activation of ephrinB/EphB forward signaling was mediated by the NR2B subunit of NMDA receptors, which was followed by a distinct PI3K/Akt/NF-κB signaling pathway, as pharmacological interference of each step of this pathway caused a reduction of TNF-α production, thus attenuating RGC apoptosis. Functional analysis of forward and reverse signaling in such a unique system, in which ephrin and Eph exist respectively in a glial element and a neuronal element, is of theoretical importance. Moreover, our results also raise a possibility that suppression of ephrinB/EphB forward signaling may be a new strategy for ameliorating RGC apoptosis in glaucoma.
Highlights
Erythropoietin-producing hepatocyte receptor (Eph) is the largest family of transmembrane receptor tyrosine kinases [35, 51]
EphrinB/EphB forward signaling is activated in chronic ocular hypertension (COH) retinas As shown in Fig. 1, in COH rats the average Intraocular pressure (IOP) of operated eyes were kept at high levels from G1w to G4w (19.2 ± 0.5 mmHg to 17.2 ± 1.1 mmHg, n = 6~ 24), which was significantly higher than that at 0d (9.1 ± 0.2 mmHg, n = 24), and of those of unoperated eyes (9.1 ± 0.2 mmHg to 9.3 ± 0.2 mmHg, n = 6~ 24) (P all < 0.001)
To find out whether ephrinB/EphB forward signaling was activated in rat COH retinas, EphB1 and phosphorylated EphB (p-EphB) protein levels of retinal extracts obtained from COH rats were determined at different post-operational times by Western blotting
Summary
Erythropoietin-producing hepatocyte receptor (Eph) is the largest family of transmembrane receptor tyrosine kinases [35, 51]. Eph receptors interacting with their ligands (ephrins, Eph receptor-interacting proteins) that are expressed in the membrane of adjacent cells, initiate forward signaling acting on receptor-expressing cells and reverse signaling acting on ephrin-expressing cells [1, 37] Both types of signaling play crucial roles in physiological processes, such as developmental axonal guidance, cell migration, synaptic plasticity [32, 38, 48, 49]. This system is involved in pathological conditions, such as inflammatory neuropathic pain, hyperalgesic condition [2, 12, 35]. Our recent study shows that EphB/ephrinB reverse signaling is activated in retinal ganglion cells (RGCs) in chronic ocular hypertension (COH) rat retina, which contributes to
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