title: The erythropoietin receptor is not required for the development, function, and aging of rods and cells in the retinal periphery.
abstract: OBJECTIVE
  Erythropoietin (EPO) was originally described for its antiapoptotic effects on erythroid progenitor cells in bone marrow. In recent years, however, EPO has also been shown to be cytoprotective in several tissues, including the retina. There, exogenous application of EPO was reported to exert neuro- and vasoprotection in several models of retinal injury. EPO and the erythropoietin receptor (EPOR) are expressed in the retina, but the role of endogenous EPO-EPOR signaling in this tissue remains elusive. Here, we investigated the consequences for cell physiology and survival when EpoR is ablated in rod photoreceptors or in the peripheral retina.
  
  
  METHODS
  Two mouse lines were generated harboring a cyclization recombinase (CRE)-mediated knockdown of EpoR in rod photoreceptors (EpoR(flox/flox);Opn-Cre) or in a heterogeneous cell population of the retinal periphery (EpoR(flox/flox);α-Cre). The function of the retina was measured with electroretinography. Retinal morphology was analyzed in tissue sections. The vasculature of the retina was investigated on flatmount preparations, cryosections, and fluorescein angiography. Retinal nuclear layers were isolated by laser capture microdissection to test for EpoR expression. Gene expression analysis was performed with semiquantitative real-time PCR. To test if the absence of EPOR potentially increases retinal susceptibility to hypoxic stress, the knockdown mice were exposed to hypoxia.
  
  
  RESULTS
  Newborn mice had lower retinal expression levels of EpoR and soluble EpoR (sEpoR) than the adult wild-type mice. In the adult mice, the EpoR transcripts were elevated in the inner retinal layers, while expression in the photoreceptors was low. CRE-mediated deletion in the EpoR(flox/flox);Opn-Cre mice led to a decrease in EpoR mRNA expression in the outer nuclear layer. A significant decrease in EpoR expression was measured in the retina of the EpoR(flox/flox);α-Cre mice, accompanied by a strong and significant decrease in sEpoR expression. Analysis of the retinal morphology in the two knockdown lines did not reveal any developmental defects or signs of accelerated degeneration in the senescent tissue. Similarly, retinal function was not altered under scotopic and photopic conditions. In addition, EpoR knockdown had no influence on cell viability under acute hypoxic conditions. Retinal angiogenesis and vasculature were normal in the absence of EPOR. However, expression of some EPOR-signaling target genes was significantly altered in the retinas of the EpoR(flox/flox);α-Cre mice.
  
  
  CONCLUSIONS
  Our data suggest that expression of EPOR in rod photoreceptors, Müller cells, and amacrine, horizontal, and ganglion cells of the peripheral retina is not required for the maturation, function, and survival of these cells in aging tissue. Based on the expression of the EPOR-signaling target genes, we postulate that expression of soluble EPOR in the retina may modulate endogenous EPO-EPOR signaling.
authors.affiliation: Lab for Retinal Cell Biology, Department of Ophthalmology, University of Zurich, Switzerland.
authors.forename: Christian
authors.initials: C
authors.lastname: Caprara
authors.affiliation: Lab for Retinal Cell Biology, Department of Ophthalmology, University of Zurich, Switzerland.
authors.forename: Corinne
authors.initials: C
authors.lastname: Britschgi
authors.affiliation: Lab for Retinal Cell Biology, Department of Ophthalmology, University of Zurich, Switzerland.
authors.forename: Marijana
authors.initials: M
authors.lastname: Samardzija
authors.affiliation: Lab for Retinal Cell Biology, Department of Ophthalmology, University of Zurich, Switzerland ; Zurich Center for Integrative Human Physiology, University of Zurich, Switzerland ; Neuroscience Center Zurich, University of Zurich and ETH Zurich, Switzerland.
authors.forename: Christian
authors.initials: C
authors.lastname: Grimm
mesh_terms: D000818:Animals; D015687:Cell Hypoxia; D002470:Cell Survival; D016922:Cellular Senescence / Q000235:genetics; D004921:Erythropoietin / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D019426:Integrases / Q000378:metabolism; D051379:Mice; D020033:Protein Isoforms / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D017467:Receptors, Erythropoietin / Q000235:genetics / Q000378:metabolism*; D015861:Retinal Neovascularization / Q000378:metabolism / Q000473:pathology; D017948:Retinal Rod Photoreceptor Cells / Q000166:cytology* / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D020033:Protein Isoforms; D017467:Receptors, Erythropoietin; D004921:Erythropoietin; C045073:Cre recombinase; D019426:Integrases