Erythropoietin preconditioning in neuronal cultures: signaling, protection from in vitro ischemia, and proteomic analysis.
In this study we confirmed the presence of the erythropoietin (EPO) receptor on both cultured cortical neurons and PC12 cells and showed that EPO can induce changes in p38, ERK, and JNK signaling molecules in these cells. We induced EPO preconditioning in cortical neuronal cultures that protected neurons from a subsequent in vitro ischemic insult (transient oxygen-glucose deprivation). To investigate downstream changes in protein expression in EPO-preconditioned cortical neuronal cultures, we used two-dimensional gel electrophoresis. Overall, EPO preconditioning resulted in protein up-regulation, and, from 84 of the most differentially expressed proteins selected for identification, the proteins or tentative proteins were identified in 57 cases, representing 40 different proteins. Different protein spots representing the same or closely related protein(s) occurred for 13 of the identified proteins and are likely to represent posttranslational modifications or proteolytic fragments of the protein. Two proteins (78-kD glucose-regulated protein and tropomyosin, fibroblast isoform 1) were detected in control neuronal cultures, but not following EPO preconditioning treatment, whereas one protein (40S ribosomal protein SA) was detected only following EPO preconditioning. Most of the other proteins identified had not previously been associated with EPO preconditioning and will aid in the understanding of EPO's neuroprotective response and possibly the development of new therapeutic interventions to inhibit neuronal death in acute and chronic neurodegenerative diseases.
Meloni
Bruno P
BP
Department of Neurosurgery/Sir Charles Gairdner Hospital, Centre for Neuromuscular and Neurological Disorders/The University of Western Australia, Perth, Western Australia, Australia. meloni@cyllene.uwa.edu.au
Tilbrook
Peta A
PA
Boulos
Sherif
S
Arthur
Peter G
PG
Knuckey
Neville W
NW
D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects; D015180:Electrophoresis, Gel, Two-Dimensional; D004921:Erythropoietin / Q000494:pharmacology*; D007091:Image Processing, Computer-Assisted; D015151:Immunoblotting; D007150:Immunohistochemistry; D007511:Ischemia / Q000473:pathology; D009419:Nerve Tissue Proteins / Q000096:biosynthesis* / Q000235:genetics*; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D018696:Neuroprotective Agents / Q000494:pharmacology; D016716:PC12 Cells; D040901:Proteomics; D051381:Rats; D011994:Recombinant Proteins; D015398:Signal Transduction / Q000187:drug effects*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013268:Stimulation, Chemical; D014357:Trypsin
D009419:Nerve Tissue Proteins; D018696:Neuroprotective Agents; D011994:Recombinant Proteins; D004921:Erythropoietin; D014357:Trypsin