Erythropoietin is a paracrine mediator of ischemic tolerance in the brain: evidence from an in vitro model

J Neurosci. 2002 Dec 1;22(23):10291-301. doi: 10.1523/JNEUROSCI.22-23-10291.2002.

Abstract

In an in vitro model of cerebral ischemia (oxygen glucose deprivation, OGD) we investigated whether erythropoietin (EPO) plays a critical role in ischemic preconditioning. We found that EPO time and dose-dependently induced protection against OGD in rat primary cortical neurons. Protection was significant at 5 min and reached a maximum at 48 hr after EPO application. Protection was blocked by the coapplication of a soluble Epo receptor (sEpoR) or an antibody against EpoR (anti-EpoR). Medium transfer from OGD-treated astrocytes to untreated neurons induced protection against OGD in neurons, which was attenuated strongly by the application of sEpoR and anti-EpoR. In contrast, medium transfer from OGD-treated neurons to untreated neurons induced protection against OGD that did not involve EPO. In astrocytes the OGD enhanced the nuclear translocation of hypoxia-inducible factor 1 (HIF-1), the major transcription factor regulating EPO expression. Consequently, transcription of EPO-mRNA was increased in astrocytes after OGD. Cultured neurons express EpoR, and the Janus kinase-2 (JAK-2) inhibitor AG490 abolished EPO-induced tolerance against OGD. Furthermore, EPO-induced neuroprotection as well as phosphorylation of the proapoptotic Bcl family member Bad was reduced by the phosphoinositide-3 kinase (PI3K) inhibitor LY294002. The results suggest that astrocytes challenged with OGD provide paracrine protective signals to neurons. We provide evidence for the following signaling cascade: HIF-1 is activated rapidly by hypoxia in astrocytes. After HIF-1 activation the astrocytes express and release EPO. EPO activates the neuronal EPO receptor and, subsequently, JAK-2 and thereby PI3K. PI3K deactivates BAD via Akt-mediated phosphorylation and thus may inhibit hypoxia-induced apoptosis in neurons. Our results establish EPO as an important paracrine neuroprotective mediator of ischemic preconditioning.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies / pharmacology
  • Apoptosis
  • Astrocytes / cytology
  • Astrocytes / metabolism
  • Brain / blood supply*
  • Brain / cytology
  • Brain / metabolism*
  • Carrier Proteins / metabolism
  • Cells, Cultured
  • DNA-Binding Proteins / metabolism
  • Dose-Response Relationship, Drug
  • Erythropoietin / genetics
  • Erythropoietin / metabolism*
  • Erythropoietin / pharmacology
  • Glucose / deficiency
  • Glucose / metabolism
  • Hypoxia-Inducible Factor 1
  • Hypoxia-Inducible Factor 1, alpha Subunit
  • Hypoxia-Ischemia, Brain / metabolism*
  • Ischemic Preconditioning
  • Janus Kinase 2
  • Models, Biological
  • Neurons / cytology
  • Neurons / drug effects
  • Neurons / metabolism
  • Nuclear Proteins / metabolism
  • Paracrine Communication / drug effects
  • Paracrine Communication / physiology*
  • Phosphatidylinositol 3-Kinases / metabolism
  • Phosphorylation / drug effects
  • Protein-Tyrosine Kinases / metabolism
  • Proto-Oncogene Proteins*
  • RNA, Messenger / metabolism
  • Rats
  • Rats, Wistar
  • Receptors, Erythropoietin / antagonists & inhibitors
  • Receptors, Erythropoietin / metabolism
  • Signal Transduction / drug effects
  • Signal Transduction / physiology
  • Transcription Factors*
  • bcl-Associated Death Protein

Substances

  • Antibodies
  • Bad protein, rat
  • Carrier Proteins
  • DNA-Binding Proteins
  • Hif1a protein, rat
  • Hypoxia-Inducible Factor 1
  • Hypoxia-Inducible Factor 1, alpha Subunit
  • Nuclear Proteins
  • Proto-Oncogene Proteins
  • RNA, Messenger
  • Receptors, Erythropoietin
  • Transcription Factors
  • bcl-Associated Death Protein
  • Erythropoietin
  • Phosphatidylinositol 3-Kinases
  • Protein-Tyrosine Kinases
  • Jak2 protein, rat
  • Janus Kinase 2
  • Glucose