Effect of erythropoietin axotomy-induced apoptosis in rat retinal ganglion cells.
OBJECTIVE
Erythropoietin (EPO) modulates erythropoiesis by inhibiting apoptosis in erythrocyte progenitors. Recently, EPO has been shown to be protective in experimental models of mechanical trauma, neuroinflammation, cerebral and retinal ischemia, and even in a human stroke trial. However, little is known about EPO signal transduction in vivo and the usefulness of EPO in the prevention of the chronic, purely apoptotic neuronal cell death that contributes to vision loss in glaucoma and the progression of neurodegenerative diseases.


METHODS
EPO's effects and signaling in the retinal ganglion cell axotomy paradigm were studied by Western blot analysis and immunohistochemistry, receptor expression was characterized in the retina before and after lesion. EPO was injected into the vitreous body to investigate neuroprotection of axotomized rat RGCs. Moreover, EPO's effects were studied in cultures of immunopurified retinal ganglion cells. Signal-transduction pathways transmitting neuroprotective EPO effects in vivo were characterized by the use of specific kinase inhibitors, immunohistochemistry, and Western blot analysis.


RESULTS
EPO receptors (EPORs) were expressed on RGC somata and dendrites in vivo. EPOR expression did not significantly change after axotomy. Application of EPO prevented death of neurotrophic-factor-deprived immunopurified rat RGCs in vitro, rescued axotomized RGCs in vivo, and prevented caspase-3 activation. EPO-induced Akt phosphorylation and survival-promoting EPO effects were completely abolished by inhibition of PI-3-kinase. EPO neuroprotection followed a bell-shaped dose-response curve in vitro and in vivo, whereas toxic EPO effects were never observed, even at high concentrations.


CONCLUSIONS
These data support a potential role for EPO as a therapeutic molecule against predominantly apoptotic neuronal cell death in the context of glaucoma or neurodegenerative diseases and delineate the PI-3-K/Akt pathway as the predominant mediator of EPO neuroprotection in this in vivo paradigm of neuronal cell death.
Weishaupt
Jochen H
JH
Department of Neurology, University of Göttingen, Göttingen, Germany.
Rohde
Gundula
G
Pölking
Esther
E
Siren
Anna-Leena
AL
Ehrenreich
Hannelore
H
Bähr
Mathias
M
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D019771:Axotomy; D015153:Blotting, Western; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004921:Erythropoietin / Q000494:pharmacology*; D019084:Fluorescent Antibody Technique, Indirect; D018696:Neuroprotective Agents / Q000494:pharmacology*; D009900:Optic Nerve / Q000601:surgery; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D017207:Rats, Sprague-Dawley; D017467:Receptors, Erythropoietin / Q000378:metabolism; D011994:Recombinant Proteins; D012165:Retinal Ganglion Cells / Q000378:metabolism / Q000473:pathology*; D015398:Signal Transduction
D004791:Enzyme Inhibitors; D018696:Neuroprotective Agents; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011518:Proto-Oncogene Proteins; D017467:Receptors, Erythropoietin; D011994:Recombinant Proteins; D004921:Erythropoietin; C494920:Akt1 protein, rat; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; C505407:CASP3 protein, human; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases