Erythropoietin, a kidney cytokine regulating haematopoiesis (the production of blood cells), is also produced in the brain after oxidative or nitrosative stress. The transcription factor hypoxia-inducible factor-1 (HIF-1) upregulates EPO following hypoxic stimuli. Here we show that preconditioning with EPO protects neurons in models of ischaemic and degenerative damage due to excitotoxins and consequent generation of free radicals, including nitric oxide (NO). Activation of neuronal EPO receptors (EPORs) prevents apoptosis induced by NMDA (N-methyl-d-aspartate) or NO by triggering cross-talk between the signalling pathways of Janus kinase-2 (Jak2) and nuclear factor-kappaB (NF-kappaB). We show that EPOR-mediated activation of Jak2 leads to phosphorylation of the inhibitor of NF-kappaB (IkappaB), subsequent nuclear translocation of the transcription factor NF-kappaB, and NF-kappaB-dependent transcription of neuroprotective genes. Transfection of cerebrocortical neurons with a dominant interfering form of Jak2 or an IkappaBalpha super-repressor blocks EPO-mediated prevention of neuronal apoptosis. Thus neuronal EPORs activate a neuroprotective pathway that is distinct from previously well characterized Jak and NF-kappaB functions. Moreover, this EPO effect may underlie neuroprotection mediated by hypoxic-ischaemic preconditioning.

The haematopoietic growth factor erythropoietin is the primary regulator of mammalian erythropoiesis and is produced by the kidney and the liver in an oxygen-dependent manner. We and others have recently demonstrated erythropoietin gene expression in the rodent brain. In this work, we show that cerebral erythropoietin gene expression is not restricted to rodents but occurs also in the primate brain. Erythropoietin mRNA was detected in biopsies from the human hippocampus, amygdala and temporal cortex and in various brain areas of the monkey Macaca mulatta. Exposure to a low level of oxygen led to elevated erythropoietin mRNA levels in the monkey brain, as did anaemia in the mouse brain. In addition, erythropoietin receptor mRNA was detected in all brain biopsies tested from man, monkey and mouse. Analysis of primary cerebral cells isolated from newborn mice revealed that astrocytes, but not microglia cells, expressed erythropoietin. When incubated at 1% oxygen, astrocytes showed >100-fold time-dependent erythropoietin mRNA accumulation, as measured with the quantitative reverse transcription-polymerase chain reaction. The specificity of hypoxic gene induction in these cells was confirmed by quantitative Northern blot analysis showing hypoxic up-regulation of mRNA encoding the vascular endothelial growth factor, but not of other genes. These findings demonstrate that erythropoietin and its receptor are expressed in the brain of primates as they are in rodents, and that, at least in mice, primary astrocytes are a source of cerebral erythropoietin expression which can be up-regulated by reduced oxygenation.

Bax inhibitor-1 (BI-1) is an evolutionarily conserved endoplasmic reticulum (ER) protein that suppresses cell death in both animal and plant cells. We characterized mice in which the bi-1 gene was ablated. Cells from BI-1-deficient mice, including fibroblasts, hepatocytes, and neurons, display selective hypersensitivity to apoptosis induced by ER stress agents (thapsigargin, tunicamycin, brefeldin A), but not to stimulators of mitochondrial or TNF/Fas-death receptor apoptosis pathways. Conversely, BI-1 overexpression protects against apoptosis induced by ER stress. BI-1-mediated protection from apoptosis induced by ER stress correlated with inhibition of Bax activation and translocation to mitochondria, preservation of mitochondrial membrane potential, and suppression of caspase activation. BI-1 overexpression also reduces releasable Ca(2+) from the ER. In vivo, bi-1(-/-) mice exhibit increased sensitivity to tissue damage induced by stimuli that trigger ER stress, including stroke and tunicamycin injection. Thus, BI-1 regulates a cell death pathway important for cytopreservation during ER stress.