title: Intrinsic and extrinsic erythropoietin enhances neuroprotection against ischemia and reperfusion injury in vitro.
abstract: This study was designed to investigate the neuroprotective effect of intrinsic and extrinsic erythropoietin (EPO) against hypoxia/ischemia, and determine the optimal time-window with respect to the EPO-induced neuroprotection. Experiments were conducted using primary mixed neuronal/astrocytic cultures and neuron-rich cultures. Hypoxia (2%) induces hypoxia-inducible factor-1alpha (HIF-1alpha) activity followed by strong EPO expression in mixed cultures and weak expression in neuron-rich cultures as documented by both western blot and RT-PCR. Immunoreactive EPO was strongly detected in astrocytes, whereas EPOR was only detected in neurons. Neurons were significantly damaged in neuron-rich cultures but were distinctly rescued in mixed cultures. Application of recombinant human EPO (rhEPO) (0.1 U/mL) within 6 h before or after hypoxia significantly increased neuronal survival compared with no rhEPO treatment. Application of rhEPO after onset of reoxygenation achieved the maximal neuronal protection against ischemia/reperfusion injury (6 h hypoxia followed 24 h reoxygenation). Our results indicate that HIF-1alpha induces EPO gene released by astrocytes and acts as an essential mediator of neuroprotection, prove the protective role of intrinsic astrocytic-neuronal signaling pathway in hypoxic/ischemic injury and demonstrate an optimal therapeutic time-window of extrinsic rhEPO in ischemia/reperfusion injury in vitro. The results point to the potential beneficial effects of HIF-1alpha and EPO for the possible treatment of stroke.
authors.affiliation: Department of Neurology, Juntendo University of School of Medicine, Bunkyo-ku, Tokyo, Japan.
authors.forename: Ruiqin
authors.initials: R
authors.lastname: Liu
authors.forename: Asuka
authors.initials: A
authors.lastname: Suzuki
authors.forename: Zhiwei
authors.initials: Z
authors.lastname: Guo
authors.forename: Yoshikuni
authors.initials: Y
authors.lastname: Mizuno
authors.forename: Takao
authors.initials: T
authors.lastname: Urabe
mesh_terms: D000818:Animals; D002545:Brain Ischemia / Q000517:prevention & control*; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000502:physiology; D004921:Erythropoietin / Q000494:pharmacology / Q000502:physiology*; D006801:Humans; D009474:Neurons / Q000166:cytology* / Q000187:drug effects / Q000502:physiology; D018696:Neuroprotective Agents / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D017467:Receptors, Erythropoietin / Q000502:physiology; D011994:Recombinant Proteins; D015427:Reperfusion Injury / Q000517:prevention & control*
chemical_list: D018696:Neuroprotective Agents; D017467:Receptors, Erythropoietin; D011994:Recombinant Proteins; D004921:Erythropoietin