The Efficacy And Mechanisms Of Erythropoietin Protects Retina From Intravitreal Injection Of Ceramide 2 Induced Retinal Damage In Rat

Purpose: To investigate the protective effect and possible mechanism of erythropoietin(EPO) in ceramide 2(C2) induced retinal damage in rat.Methods: Male Sprague–Dawley(SD) rats were randomly divided into 4 groups(n=6 in each group): normal control, DMSO treated group, C2 treated group, and C2+EPO treated group. Retinal damage was induced by intravitreal injection of C2. In C2 + EPO treated group, EPO was administered three days after C2 injection, both EPO and C2 were given intravitreally. The retinal function was evaluated by flash electroretinogram(f–ERG). Retinal cell death was measured by TUNEL. The transendothelial electrical resistance(TEER) was detected in the primary human retinal microvascular endothelial cells(HRMECs). The changes of glial fibrillary acidic protein(GFAP), plasmalemma vesicle–associated protein(PLVAP, also known as PV–1 or PAL–E),ecto–5’–nucleotidase(CD73) and intercellular adhesion molecule–1(ICAM–1) in R28 cells were studied with immunofluorescence and real–time PCR(Q–PCR).Results: C2 treated rats demonstrated significant vision loss as reflected by the reduction of b–wave amplitude in ERG recordings when compared with normal control and DMSO control. TUNEL positive cells, mainly in the retinal ganglion cell layer(GCL), and increased GFAP immunostaining were observed in C2 treated rats in comparison with the two control groups(normal control, DMSO treated group). EPO demonstrated obvious rescue effect in this retinal injury model as supported by the increased b–wave amplitude, decreased GFAP expression, and reduced retinal cell death. The TEER value was significantly reduced in C2 treated HRMECs, which could be reversed by EPO. C2 treated R28 cells in vitro showed evidently decreased cell viability in a dose–dependent manner, and significant increase in m RNA levels of PV–1, CD73 and ICAM–1. All these changes were significantly reversed after EPO treatment.Conclusions: C2 ceramide can induce severe retinal injury, i.e., retinal cell death and BRB breakdown. EPO can protect the function and structure of retina in such C2–induced retinal damage.