[Frontiers in Bioscience E3, 1541-1555, June 1, 2011]

EPO reduces reactive gliosis and stimulates neurotrophin expression in Muller cells

Liu-Mei Hu1, Yan Luo1, Jingfa Zhang2, 3, Xia Lei3, Jianfeng Shen3, Yalan Wu3, Mei Qin1, Yaprak Banu Unver4, Yong Zhong1, Guo-Tong Xu2, 3, Weiye Li1, 2, 3, 5

1Department of Ophthalmology, Peking Union Medical College Hospital, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China,2Tongji Eye Institute and Department of Regenerative Medicine, Tongji University School of Medicine, Shanghai, China, 3Laboratory of Clinical Visual Sciences, Institute of Health Sciences, Shanghai Institutes for Biological Sciences and Shanghai Jiaotong University School of Medicine, Chinese Academy of Sciences, Shanghai, China, 4Beyoglu Eye Research and Training Hospital, Istanbul, Turkey, 5Department of Ophthalmology, Drexel University College of Medicine, Philadelphia, PA, USA


1. Abstract
2. Introduction
3. Materials and methods
3.1. Experimental animals and intravitreal injection
3.2. Cell culture of retinal Müller cells
3.3. Sample preparations and immunofluorescence study
3.4. Western blotting analysis for GFAP, phospho- and total-CREB
3.5. Enzyme-linked immunosorbent assay (ELISA)
3.6. RNA isolation and gene expression determined by real-time PCR
3.7. Primary retinal neuronal cell culture and neurite outgrowth assay
3.8. Statistic analysis
4. Results
4.1. Müller cells become reactive with the progression of diabetes
4.2. Glial localization of EPO receptor
4.3. EPO down-regulates GFAP and vimentin expressions in diabetic rat retinas
4.4. EPO enhances the expressions of BDNF and CNTF by Müller cells in diabetic rat retinas
4.5. EPO up-regulates the production of BDNF and CNTF by rMC-1 cells
4.6. EPO promotes neurite outgrowth in part via BDNF
4.7. ERK1/2 and PI3K pathways are involved in EPO's neurotrophic effect
5. Discussion
6. Acknowledgement
7. References


To characterize Müller cell-mediated neuroprotective and neurotrophic functions of the erythropoietin (EPO)/EPO receptor (EpoR) system in diabetic rat retina. A single intravitreal injection of EPO (8 mU/eye) was administered in rats 4 or 24 weeks after diabetes onset. The results showed that intravitreal EPO ameliorated the up-regulation of GFAP and vimentin in the diabetic retina evaluated by immunofluorescence and Western blotting; but up-regulated BDNF and CNTF expressions, quantified by real-time PCR and ELISA, in the 24-week diabetic rat retinas. In vitro, BDNF and CNTF expressions were stimulated by EPO through both extracellular signal-regulated kinase1/2 (ERK1/2) and Akt pathways. The neuro-regenerative function of EPO, as indicated by promotion of neurite outgrowth, was corroborated in vitro. BDNF was involved in EPO-induced neurite outgrowth of primary rat retinal neurons. Exogenous EPO exerts neuroprotective and neurotrophic functions by attenuating reactive gliosis and promoting neurotrophic factors in Müller cells in diabetic retina. Signaling pathways that are responsible for these Müller cell-mediated EPO/EpoR functions may be therapeutic targets for diabetic retinopathy.