losartan-potassium and Nerve-Degeneration

In the United States, TBI remains a major cause of morbidity and mortality in children and young adults. A total of 1.5 million Americans experience head trauma every year, and the yearly economic cost of this exceeds $56 billion. The magnitude of this problem has generated a great deal of interest in elucidating the complex molecular mechanism underlying cell death and dysfunction after TBI and in the development of neuroprotective agents that will reduce morbidity and mortality.. A review of recent literature on EPO, TBI, and apoptosis is conducted with analysis of pathophysiologic mechanisms of TBI. In addition, animal experiments and clinical trials pertaining to mechanisms of cell death in TBI and EPO as a neuroprotective agent are reviewed.. The literature and evidence for EPO as a potent inhibitor of apoptosis and promising therapeutic agent in a variety of neurological insults, including trauma, are mounting. With the recent interest in clinical trials of EPO in human stroke, it is both timely and prudent to consider the use of this pharmaceutical avenue in TBI in man.

Topics: Animals; Apoptosis; Apoptosis Regulatory Proteins; Brain Injuries; Cytoprotection; Disease Models, Animal; Erythropoietin; Humans; Nerve Degeneration; Neurons; Neuroprotective Agents

Spinal cord injury (SCI) is a devastating condition for individual patients and costly for health care systems requiring significant long-term expenditures. Cytokine erythropoietin (EPO) is a glycoprotein mediating cytoprotection in a variety of tissues, including spinal cord, through activation of multiple signaling pathways. It has been reported that EPO exerts its beneficial effects by apoptosis blockage, reduction of inflammation, and restoration of vascular integrity. Neuronal regeneration has been also suggested. In the present review, the pathophysiology of SCI and the properties of endogenous or exogenously administered EPO are briefly described. Moreover, an attempt to present the current traumatic, ischemic and inflammatory animal models that mimic SCI is made. Currently, a clearly effective pharmacological treatment is lacking. It is highlighted that administration of EPO or other recently generated EPO analogues such as asialo-EPO and carbamylated-EPO demonstrate exceptional preclinical characteristics, rendering the evaluation of these tissue-protective agents imperative in human clinical trials.

Topics: Animals; Clinical Trials as Topic; Disease Models, Animal; Drug Evaluation, Preclinical; Erythropoietin; Humans; Myelitis; Nerve Degeneration; Neuroprotective Agents; Spinal Cord; Spinal Cord Injuries

Leptin is well known as a hormone important in the central control of appetitive behaviors via receptor-mediated actions in the hypothalamus, where leptin adjusts food intake to maintain homeostasis with the body's energy stores. Recent evidence has shown that leptin and its receptors are widespread in the CNS and may provide neuronal survival signals. This review summarizes our current knowledge of how leptin functions in the brain and then focuses on the ability of leptin to mitigate neuronal damage in experimental models of human neurological disorders. Damage to the brain by acute events such as stroke, or long-term loss of neurons associated with neurodegenerative diseases, including Parkinson's and Alzheimer's disease, may be amenable to treatment using leptin to limit death of susceptible cells. Leptin-mediated pro-survival signaling is now known to prevent the death of neurons in these models. The signaling cascades that leptin generates are shared by other neuroprotective molecules including insulin and erythropoietin, and are thus a component of the neurotrophic effects mediated by endogenous hormones. Coupled with evidence that leptin dysregulation in human disease also results in enhanced neuronal susceptibility to damage, development of leptin as a therapeutic methodology is an attractive and viable possibility.

Topics: Animals; Brain Diseases; Cell Death; Central Nervous System; Erythropoietin; Humans; Insulin; Leptin; Nerve Degeneration; Neuroprotective Agents; Signal Transduction

With the increased life expectancy in western industrialized countries, the incidence and prevalence of brain diseases dramatically increased. Stroke and a wide spectrum of neuropsychiatric illnesses such as Alzheimer's disease, Parkinson's disease, multiple sclerosis, traumatic head injury, and schizophrenia all lead to severe disability. However, targeted effective therapies for treatment of these diseases are lacking. Even more frustrating is the fact that we do not yet clearly understand the basic mechanisms underlying the disease processes in these conditions. We propose a hypothesis of loss of neuronal function via a final common deleterious pathway in this clinically very heterogeneous disease group. This review presents a novel neuroprotective concept for treatment of brain disease: Erythropoietin (EPO). EPO is a natural body-own-protein hormone that has been used for treatment of anemia for more than a decade. The neuroprotective approach using EPO in brain disease represents a totally new frontier. The "Göttingen EPO-stroke trial" represents the first effective use in man of a neuroprotective therapy in an acute brain disease while the experimental EPO therapy to combat cognitive decline in patients with schizophrenia will be introduced as an example of a neuroprotective strategy for a chronic brain disease.

Topics: Animals; Brain Diseases; Clinical Trials as Topic; Erythropoietin; Humans; Nerve Degeneration; Neuroprotective Agents; Regeneration; Schizophrenia; Stroke

Erythropoietin (EPO) has potent neuroprotective effects. The short-term delivery of high-dose EPO seemed to improve patients' neuromuscular functions; however, excessive EPO resulted in systematically high hematocrit and thrombotic risk. In our study, we established a cellular material for future in vivo studies of neurodegenerative diseases based on EPO provided regionally at a nontoxic level.. A mouse EPO cDNA was subcloned into the pCMS-EGFP vector and transfected into NIH/3T3 fibroblasts to design a biological provider that can regionally release EPO for the treatment of neurological diseases. After G418 selection, a stable EPO-overexpressing cell line, EPO-3T3-EGFP, was established. To further confirm the neuroprotective abilities of secreted EPO from EPO-3T3-EGFP cells, a cell model of neurodegeneration, PC12-INT-EGFP, was applied.. The expression level of EPO was highly elevated in EPO-3T3-EGFP cells, and an abundant amount of EPO secreted from EPO-3T3-EGFP cells was detected in the extracellular milieu. After supplementation with conditioned medium prepared from EPO-3T3-EGFP cells, the survival rate of PC12-INT-EGFP cells was significantly enhanced. Surprisingly, a fraction of aggregated cytoskeletal EGFP-tagged α-internexin in PC12-INT-EGFP cells was disaggregated and transported into neurites dynamically. The immunocytochemical distribution of IF proteins, including NF-M, phosphorylated-NF-M, and the α-INT-EGFP fusion protein, were less aggregated in the perikaryal region and transported into neurites after the EPO treatment.. The established EPO-overexpressing NIH/3T3 cell line, EPO-3T3-EGFP, may provide a material for future studies of cell-based therapies for neurodegenerative diseases via the secretion of EPO on a short-term, high-dose, regional basis.

Topics: Animals; Cloning, Molecular; DNA, Complementary; Erythropoietin; Humans; Mice; Nerve Degeneration; Neuroprotective Agents; NIH 3T3 Cells; Protein Engineering; Recombinant Proteins

So far, typical causes of presbycusis such as degeneration of hair cells and/or primary auditory (spiral ganglion) neurons cannot be treated. Because erythropoietin's (Epo) neuroprotective potential has been shown previously, we determined hearing thresholds of juvenile and aged mice overexpressing Epo in neuronal tissues. Behavioral audiometry revealed in contrast to 5 months of age, that 11-month-old Epo-transgenic mice had up to 35 dB lower hearing thresholds between 1.4 and 32 kHz, and at the highest frequencies (50-80 kHz), thresholds could be obtained in aged Epo-transgenic only but not anymore in old C57BL6 control mice. Click-evoked auditory brainstem response showed similar results. Numbers of spiral ganglion neurons in aged C57BL6 but not Epo-transgenic mice were dramatically reduced mainly in the basal turn, the location of high frequencies. In addition, there was a tendency to better preservation of inner and outer hair cells in Epo-transgenic mice. Hence, Epo's known neuroprotective action effectively suppresses the loss of spiral ganglion cells and probably also hair cells and, thus, development of presbycusis in mice.

Topics: Animals; Erythropoietin; Evoked Potentials, Auditory; Gene Expression; Gene Expression Regulation, Developmental; Hair Cells, Auditory; Mice, Inbred C57BL; Mice, Transgenic; Nerve Degeneration; Neuroprotective Agents; Presbycusis; Spiral Ganglion

Erythropoiesis must be tightly balanced to guarantee adequate oxygen delivery to all tissues in the body. This process relies predominantly on the hormone erythropoietin (EPO) and its transcription factor hypoxia inducible factor (HIF). Accumulating evidence suggests that oxygen-sensitive prolyl hydroxylases (PHDs) are important regulators of this entire system. Here, we describe a novel mouse line with conditional PHD2 inactivation (cKO P2) in renal EPO producing cells, neurons, and astrocytes that displayed excessive erythrocytosis because of severe overproduction of EPO, exclusively driven by HIF-2α. In contrast, HIF-1α served as a protective factor, ensuring survival of cKO P2 mice with HCT values up to 86%. Using different genetic approaches, we show that simultaneous inactivation of PHD2 and HIF-1α resulted in a drastic PHD3 reduction with consequent overexpression of HIF-2α-related genes, neurodegeneration, and lethality. Taken together, our results demonstrate for the first time that conditional loss of PHD2 in mice leads to HIF-2α-dependent erythrocytosis, whereas HIF-1α protects these mice, providing a platform for developing new treatments of EPO-related disorders, such as anemia.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Brain; Cells, Cultured; Erythropoietin; Female; Fibroblasts; Hematopoiesis, Extramedullary; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Hypoxia-Inducible Factor-Proline Dioxygenases; Keratinocytes; Kidney; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Nerve Degeneration; Polycythemia; Procollagen-Proline Dioxygenase; Severity of Illness Index; Thrombocytopenia

Iron regulatory proteins (Irps) 1 and 2 posttranscriptionally control the expression of transcripts that contain iron-responsive element (IRE) sequences, including ferritin, ferroportin, transferrin receptor, and hypoxia-inducible factor 2α (HIF2α). We report here that mice with targeted deletion of Irp1 developed pulmonary hypertension and polycythemia that was exacerbated by a low-iron diet. Hematocrits increased to 65% in iron-starved mice, and many polycythemic mice died of abdominal hemorrhages. Irp1 deletion enhanced HIF2α protein expression in kidneys of Irp1(-/-) mice, which led to increased erythropoietin (EPO) expression, polycythemia, and concomitant tissue iron deficiency. Increased HIF2α expression in pulmonary endothelial cells induced high expression of endothelin-1, likely contributing to the pulmonary hypertension of Irp1(-/-) mice. Our results reveal why anemia is an early physiological consequence of iron deficiency, highlight the physiological significance of Irp1 in regulating erythropoiesis and iron distribution, and provide important insights into the molecular pathogenesis of pulmonary hypertension.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Diet; Endothelial Cells; Endothelin-1; Erythropoietin; Gastrointestinal Hemorrhage; Gene Deletion; Hematopoiesis, Extramedullary; Hypertension, Pulmonary; Iron; Iron Regulatory Protein 1; Iron Regulatory Protein 2; Longevity; Mice; Models, Biological; Nerve Degeneration; Organ Specificity; Polycythemia; Protein Biosynthesis; Transcriptional Activation

We examined the preventive effect of human recombinant erythropoietin (HrEPO) on nitric oxide (NO)-mediated toxicity to neurons and cysteine protease release into cytoplasm, which is attributed to neuronal death in brain ischemia. Focal cerebral ischemia was induced by permanent occlusion of middle cerebral artery in two sets of rat. The first set was used to monitor NO concentration and cathepsin activity, while the second was used for histological examination with hematoxylin and eosin, and TUNEL staining. A group in both set was administered human recombinant erythropoietin (HrEPO). NO content, cathepsins B and L activity increased significantly in the post-ischemic cerebral tissue (p<0.05). HrEPO treatment reduced NO concentration and cathepsin activity to control level (p>0.05). A significant increase in the number of necrotic and apoptotic neurons was observed in the post-ischemic cerebral cortex (p<0.05). HrEPO treatment was markedly lowered both of these (p<0.05). It is concluded that HrEPO prevents neuronal death by protecting neuronal liposomes from NO-mediated toxicity and suppressing the release of cathepsins.

Topics: Animals; Brain Infarction; Brain Ischemia; Cathepsin B; Cathepsin L; Cell Death; Disease Models, Animal; Erythropoietin; Humans; Nerve Degeneration; Neuroprotective Agents; Rats; Rats, Sprague-Dawley

Hypoglycemia is a common complication for insulin treated people with diabetes. Severe hypoglycemia, which occurs in the setting of excess or ill-timed insulin administration, has been shown to cause brain damage. Previous pre-clinical studies have shown that memantine (an N-methyl-d-aspartate receptor antagonist) and erythropoietin can be neuroprotective in other models of brain injury. We hypothesized that these agents might also be neuroprotective in response to severe hypoglycemia-induced brain damage. To test this hypothesis, 9-week old, awake, male Sprague-Dawley rats underwent hyperinsulinemic (0.2 U kg(-1)min(-1)) hypoglycemic clamps to induce severe hypoglycemia (blood glucose 10-15 mg/dl for 90 min). Animals were randomized into control (vehicle) or pharmacological treatments (memantine or erythropoietin). One week after severe hypoglycemia, neuronal damage was assessed by Fluoro-Jade B and hematoxylin and eosin staining of brain sections. Treatment with both memantine and erythropoietin significantly decreased severe hypoglycemia-induced neuronal damage in the cortex by 35% and 39%, respectively (both p<0.05 vs. controls). These findings demonstrate that memantine and erythropoietin provide a protective effect against severe hypoglycemia-induced neuronal damage.

Topics: Animals; Cell Count; Cerebral Cortex; Erythropoietin; Hippocampus; Hypoglycemia; Insulin; Male; Memantine; Nerve Degeneration; Neurons; Neuroprotective Agents; Rats; Rats, Sprague-Dawley

The design of peptide mimetics offers interesting opportunities to selectively include beneficial and exclude undesirable effects of a parent molecule. Epotris represents a novel erythropoietin mimetic, which lacks an erythropoietic activity. The present study evaluates the potential of this peptide to interfere with the histopathological consequences of electrical-induced status epilepticus in rats. The peptide attenuated status epilepticus-associated expansion of the neuronal progenitor cell population in a significant manner. Moreover, Epotris affected the number of persistent basal dendrites exhibited by neuronal progenitor cells. In contrast, hippocampal cell loss remained unaffected by administration of this peptide mimetic. Status epilepticus resulted in obvious microglial activation in different brain regions involved in seizure generation and spread. Epotris diminished the microglial response caused by prolonged seizure activity in the thalamus but not in other brain regions. The study renders support that the Epotris' sequences from binding site 2 in helix C of Epo play a role in receptor interaction and cytokine function. In addition, the data demonstrate that Epotris can exert limited in vivo effects on the cellular consequences of prolonged seizure activity. When considering further testing it should be taken in mind that Epotris administration only attenuated selected cellular consequences of status epilepticus and did not completely prevent cellular alterations.

Topics: Animals; Doublecortin Domain Proteins; Electric Stimulation; Electrodes, Implanted; Erythropoietin; Female; Microglia; Microtubule-Associated Proteins; Molecular Mimicry; Nerve Degeneration; Neural Stem Cells; Neurogenesis; Neuropeptides; Peptide Fragments; Rats; Rats, Sprague-Dawley; Status Epilepticus

To examine whether recombinant erythropoietin (rEPO) attenuates neurodegeneration and the learning disability induced by isoflurane with the postnatal day 7 (P7) mice.. Some of general anesthetic agents induce neurodegeneration in developing brain. Several drugs, but not rEPO, were reported as candidates for the prevention of or treatment for neurodegeneration.. We divided P7 mice into three groups at random. One group (IE group) was exposed to 6-h isoflurane (1.0%) after 50,000 IU·kg(-1) rEPO administered subcutaneously. The second group (I) was exposed to isoflurane in the same manner as IE group except saline instead of rEPO. The third group (E) was exposed to air after rEPO administered. The mice were assigned to the radial arm maze on four consecutive days from P56 (day 1) to P59 (day 4). We divided the number of errors each day by that of day 1 to establish each-day performance ratio. After the test, neurodegenerative change in the hilus of dentate gyrus was assessed using Nissl staining.. In radial maze test, the performance ratios of day 3 (mean ± sd) were 0.3 ± 0.2 (P < 0.05, vs I group), 0.8 ± 0.5, and 0.6 ± 0.2 in IE, I, and E groups, respectively, while those of day 4 were 0.3 ± 0.1 (P < 0.05), 0.8 ± 0.5, and 0.3 ± 0.2 (P < 0.05), respectively. The histopathological study revealed that in IE group the degenerative neuronal change was attenuated compared with I group.. These results suggested that rEPO attenuated isoflurane-induced neurodegeneration.

Topics: Air; Analysis of Variance; Anesthetics, Inhalation; Animals; Animals, Newborn; Apoptosis; Brain; Disease Models, Animal; Erythropoietin; Isoflurane; Maze Learning; Mice; Mice, Inbred C57BL; Nerve Degeneration; Sodium Chloride

The Akita mouse is a robust model of diabetic autonomic neuropathy which develops severe diabetes following beta cell death, which occurs reproducibly at 3-4 weeks of age, and maintains the diabetic state without therapy for as long as 11 additional months. Neuritic dystrophy and neuronopathy involving prevertebral sympathetic superior mesenteric and celiac ganglia begin to develop within the first two months of onset of diabetes and are progressive with time. We have examined the effect of insulin implants resulting in normoglycemia and injections of ARA290, a small erythropoietin peptide which has no effect on glycemic parameters, on the reversal of established neuritic dystrophy and neuronopathy. We have found that 4 weeks of insulin therapy beginning at 2 months of diabetes resulted in normalization of blood glucose, body weight and HbA1c. Insulin therapy successfully reversed established neuritic dystrophy and neuronopathy to control levels. Numbers of sympathetic neurons were not significantly changed in either 3 month diabetic or insulin-treated Akita mice. Treatment with ARA290 for 7 weeks beginning at 4 months of diabetes did not result in altered metabolic severity of diabetes as measured by blood glucose, body weight or HbA1c levels. ARA290 treatment was able to decrease neuritic dystrophy by 55-74% compared to untreated diabetics or in comparison to a separate group of diabetic animals representing the 4 month treatment onset point. Surprisingly, there was no effect of ARA290 on ganglionic neuron number or ongoing neuronopathy (pale/degenerating neurons) in diabetic Akita mice during this time period. The development of neuroprotective EPO-like peptides may provide a possible future therapy for this debilitating complication of diabetes; however, it appears that discrete elements may be differentially targeted by the diabetic state and may require selective therapy.

Topics: Animals; Blood Glucose; Body Weight; Diabetes Mellitus, Type 1; Diabetic Neuropathies; Disease Models, Animal; Erythropoietin; Ganglia, Sympathetic; Hypoglycemic Agents; Insulin; Male; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Microscopy, Electron; Nerve Degeneration; Neurites; Peptides

Erythropoietin (EPO) may be protective for early stage diabetic retinopathy, although there are concerns that it could exacerbate retinal angiogenesis and thrombosis. A peptide based on the EPO helix-B domain (helix B-surface peptide [pHBSP]) is nonerythrogenic but retains tissue-protective properties, and this study evaluates its therapeutic potential in diabetic retinopathy.. After 6 months of streptozotocin-induced diabetes, rats (n = 12) and age-matched nondiabetic controls (n = 12) were evenly split into pHBSP and scrambled peptide groups and injected daily (10 μg/kg per day) for 1 month. The retina was investigated for glial dysfunction, microglial activation, and neuronal DNA damage. The vasculature was dual stained with isolectin and collagen IV. Retinal cytokine expression was quantified using real-time RT-PCR. In parallel, oxygen-induced retinopathy (OIR) was used to evaluate the effects of pHBSP on retinal ischemia and neovascularization (1-30 μg/kg pHBSP or control peptide).. pHBSP or scrambled peptide treatment did not alter hematocrit. In the diabetic retina, Müller glial expression of glial fibrillary acidic protein was increased when compared with nondiabetic controls, but pHBSP significantly reduced this stress-related response (P < 0.001). CD11b+ microglia and proinflammatory cytokines were elevated in diabetic retina responses, and some of these responses were attenuated by pHBSP (P < 0.01-0.001). pHBSP significantly reduced diabetes-linked DNA damage as determined by 8-hydroxydeoxyguanosine and transferase-mediated dUTP nick-end labeling positivity and also prevented acellular capillary formation (P < 0.05). In OIR, pHBSP had no effect on preretinal neovascularization at any dose.. Treatment with an EPO-derived peptide after diabetes is fully established can significantly protect against neuroglial and vascular degenerative pathology without altering hematocrit or exacerbating neovascularization. These findings have therapeutic implications for disorders such as diabetic retinopathy.

Topics: Animals; Animals, Newborn; Apoptosis; Cytokines; Diabetic Retinopathy; DNA Damage; Erythropoietin; Gene Expression Regulation; Ischemia; Male; Mice; Mice, Inbred C57BL; Nerve Degeneration; Neuroglia; Peptide Fragments; Protein Interaction Domains and Motifs; Random Allocation; Rats; Rats, Sprague-Dawley; Retina; Retinal Degeneration; Retinal Vessels

Erythropoietin (EPO), essential for erythropoiesis, provides neuroprotection. The EPO receptor (EPOR) is expressed in both neural and non-neural cells in the brain. This study was designed to test the hypothesis that EPO provides beneficial therapeutic effects, even in the absence of the neural EPOR. In this study, EPOR-null mice were rescued with selective EpoR expression driven by the endogenous EpoR promoter in hematopoietic tissue, but not in the neural cells. Anesthetized young adult female EPOR-null and wild-type mice were subjected to traumatic brain injury (TBI) induced by controlled cortical impact. EPO (5000 U/kg) or saline was intraperitoneally administered at 6 h and 3 and 7 days post-injury. Sensorimotor and spatial learning functions were assessed. Expression of EPOR and its downstream signal proteins were evaluated by Western blot analysis. Our data demonstrated that EPO treatment significantly reduced cortical tissue damage and hippocampal cell loss, and improved spatial learning following TBI in both the wild-type and EPOR-null mice. EPO treatment significantly improved sensorimotor functional recovery, with better outcomes in the wild-type mice. EPO treatment upregulated anti-apoptotic proteins (p-Akt and Bcl-XL) in the ipsilateral hippocampus and cortex of the injured wild-type and EPOR-null mice. These data demonstrate that EPO significantly provides neuroprotection following TBI, even in the absence of EPOR in the neural cells, suggesting that its therapeutic benefits may be mediated through vascular protection.

Topics: Animals; Apoptosis Regulatory Proteins; Brain; Brain Injuries; Cerebral Cortex; Disease Models, Animal; Erythropoietin; Female; Hippocampus; Humans; Memory Disorders; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Nerve Degeneration; Neuroprotective Agents; Promoter Regions, Genetic; Receptors, Erythropoietin; Treatment Outcome

Erythropoietin (EPO) is a chemokine hormone that is widely distributed throughout the body including nervous system. For last years its role as cytokine involved in many physiological processes out of the bone marrow has been suggested. Moreover, it plays a very important role in CNS as potential neuroprotective agent. There is much evidence that EPO protects neuronal cells in vitro and in vivo models of brain injury, independently of its erythropoietic action. The aim of this study was to determine the potential neuroprotective effects of erythropoietin on the glutamate-mediated injury of motor neurons (MNs) in vitro. The study was performed on organotypic cultures of the rat lumbar spinal cord subjected to glutamate uptake blocker, DL-threo-beta-hydroxyaspartate (THA) and pretreated with EPO. Ultrastructural study evidenced that the spinal cord cultures pretreated with EPO exhibited less severe neuronal injury. The cultures exposed to EPO + THA showed inhibition of early MNs degeneration, including various mode of degenerative changes caused by THA, whereas in the later period the typical postsynaptic necrotic changes of neuronal cells occurred. However, the ultrastructural characteristics of apoptotic MNs changes were not observed during the whole period of observation. The results of this study indicate that, in the model of chronic glutamate excitotoxicity, EPO exhibits the neuroprotective ability mainly through prevention of apoptotic neuronal changes.

Topics: Amyotrophic Lateral Sclerosis; Animals; Aspartic Acid; Cell Membrane; Cytoplasm; Endoplasmic Reticulum; Erythropoietin; Nerve Degeneration; Neurons; Neuropil; Neuroprotective Agents; Organ Culture Techniques; Rats; Spinal Cord; Vacuoles

Exposure to Gamma-aminobutyric-acid (GABA)(A)-receptor agonists and N-Methyl-D-Aspartate (NMDA)-antagonists has been demonstrated to induce neurodegeneration in newborn rats. Exogenous erythropoietin (EPO) protects against NMDA antagonist-mediated neuronal death. In this study we evaluated whether EPO is also effective in limiting neurodegeneration of the GABA(A)-mimetic agent propofol in newborn rats. 6 day old rats were randomized to one of four groups and treated with intraperitoneal applications of 3 x 30 mg/kg propofol at 0, 90 and 180 min, propofol in combination with 5000 IU/kg rEPO, propofol in combination with 20,000 IU/kg rEPO or sham injections of PAD II solution as controls. After 24h, brains of the animals were histopathologically examined and a summation score of degenerated cells was calculated for every brain. Propofol increased neuronal degeneration scores from 16,090+/-4336 to 28,860+/-6569 (p<0.01). This effect was completely abolished by low-dose rEPO (14,270+/-4542, p<0.001 versus propofol only; p>0.05 versus controls). In contrast, high-dose rEPO was not protective (23 930+/-8896, p>0.05 versus propofol only). Propofol may cause neuronal death in newborn rat brains, which is prevented by low-dose rEPO but not high-dose rEPO.

Topics: Age Factors; Animals; Animals, Newborn; Dose-Response Relationship, Drug; Drug Interactions; Erythropoietin; Hypnotics and Sedatives; Injections, Intraperitoneal; Nerve Degeneration; Neuroprotective Agents; Propofol; Rats; Treatment Outcome

Chemotherapy-induced peripheral neurotoxicity (CIPN) is a side effect limiting cisplatin (CDDP) and docetaxel (DOCE) treatment. Erythropoietin (EPO) is a hematopoietic growth factor also displaying neurotrophic properties. Evidence suggests that EPO's neuroprotective action may rely on PI3K/AKT pathway activation; however, data regarding the EPO neuroprotective mechanism are still limited. This study evaluated the effect of EPO on organotypic cultures of rat dorsal root ganglia (DRG) and in primary cultures of DRG-dissociated sensory neurons exposed to CDDP- and DOCE-induced neurotoxicity, aiming to investigate EPO's neuroprotective mechanism. Subsequently, the levels of AKT expression and activation were analyzed by Western blot in neurons exposed to CDDP or DOCE; AKT's role was further evaluated by using a chemical inhibitor of AKT activation, wortmannin. In these models EPO, was protective against both CDDP- and DOCE-induced cell death and against CDDP-induced neurite elongation reduction. A modulation of AKT activation was observed in CDDP-treated neurons, and the presence of wortmannin prevented EPO's neuroprotective action against CDDP toxicity but did not have any effect on EPO's protection against DOCE-induced toxicity, thus ruling out the PI3K-AKT pathway as the mechanism of EPO's effect in neuronal death prevention after DOCE exposure. Our results confirm in vitro the effectiveness of EPO as a neuroprotectant against both CDDP- and DOCE-induced neurotoxicity. In addition, a role of PI3K/AKT in EPO's protection against CDDP, but not against DOCE, neurotoxicity was shown, suggesting that alternative pathways could be involved in EPO's neuroprotective activity.

Topics: Animals; Antineoplastic Agents; Cells, Cultured; Cisplatin; Docetaxel; Erythropoietin; Ganglia, Spinal; Nerve Degeneration; Neuroprotective Agents; Peripheral Nervous System Diseases; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Sensory Receptor Cells; Signal Transduction; Taxoids

Using magnetic resonance imaging (MRI) protocols of T(2)-, T(2)*-, diffusion- and susceptibility-weighted imaging (T2WI, T2*WI, DWI, and SWI, respectively) with a 7T system, we tested the hypothesis that treatment of embolic stroke with erythropoietin (EPO) initiated at 24 hr and administered daily for 7 days after stroke onset has benefit in repairing ischemic cerebral tissue. Adult Wistar rats were subjected to embolic stroke by means of middle cerebral artery occlusion (MCAO) and were randomly assigned to a treatment (n = 11) or a control (n = 11) group. The treated group was given EPO intraperitoneally at a dose of 5,000 IU/kg daily for 7 days starting 24 hr after MCAO. Controls were given an equal volume of saline. MRI was performed at 24 hr and then weekly for 6 weeks. MRI and histological measurements were compared between groups. Serial T2WI demonstrated that expansion of the ipsilateral ventricle was significantly reduced in the EPO-treated rats. The volume ratio of ipsilateral parenchymal tissue relative to the contralateral hemisphere was significantly increased after EPO treatment compared with control animals, indicating that EPO significantly reduces atrophy of the ipsilateral hemisphere, although no significant differences in ischemic lesion volume were observed between the two groups. Angiogenesis and white matter remodeling were significantly increased and occurred earlier in EPO-treated animals than in the controls, as evident from T2*WI and diffusion anisotropy maps, respectively. These data indicate that EPO treatment initiated 24 hr poststroke promotes angiogenesis and axonal remodeling in the ischemic boundary, which may potentially reduce atrophy of the ipsilateral hemisphere.

Topics: Animals; Atrophy; Disease Models, Animal; Erythropoietin; Intracranial Embolism; Magnetic Resonance Imaging; Male; Nerve Degeneration; Neuroprotective Agents; Random Allocation; Rats; Rats, Wistar; Recombinant Proteins; Stroke

Deferoxamine (DFO) and erythropoietin (EPO) have each been shown to provide neuroprotection in neonatal rodent models of brain injury. In view of the described anti-oxidative actions of DFO and the anti-apoptotic and anti-inflammatory effects of EPO, we hypothesized that the combination of DFO and EPO would increase neuroprotection after neonatal hypoxic-ischemic brain injury as compared to single DFO or EPO treatment. At postnatal day 7 rats underwent right common carotid artery occlusion followed by a 90-min exposure to 8% oxygen. Rats were treated intraperitoneally with DFO (200mg/kg), recombinant human EPO (1 kU/kg), a combination of DFO-EPO or vehicle at 0, 24 and 48 h after hypoxia-ischemia (HI) and were sacrificed at 72 h. DFO-EPO administration reduced the number of cleaved caspase 3-positive cells in the ipsilateral cerebral cortex. Early neuronal damage was assessed by staining for microtubuli-associated protein (MAP)-2. In our model 63+/-9% loss of ipsilateral MAP-2 was observed after HI, indicating extensive brain injury. DFO, EPO or DFO-EPO treatment did not improve neuronal integrity as defined by MAP-2. Cerebral white matter tracts were stained for myelin basic protein (MBP), a constituent of myelin. Hypoxia-ischemia strongly reduced MBP staining which suggests white matter damage. However, DFO, EPO and DFO-EPO treatment had no effect on the loss of MBP staining. Finally, HI-induced loss of striatal tyrosine hydroxylase staining was not attenuated by DFO, EPO or DFO-EPO. Although DFO-EPO treatment reduced the number of cleaved caspase 3(+) cells, treatment with DFO, EPO, or with the combination of DFO and EPO did not protect against gray or white matter damage in the experimental setting applied.

Topics: Animals; Animals, Newborn; Anti-Inflammatory Agents; Antioxidants; Brain Infarction; Caspase 3; Cerebral Cortex; Cytoprotection; Deferoxamine; Disease Models, Animal; Drug Combinations; Drug Synergism; Erythropoietin; Hypoxia-Ischemia, Brain; Microtubule-Associated Proteins; Myelin Basic Protein; Nerve Degeneration; Nerve Fibers, Myelinated; Neurons; Rats; Rats, Wistar

Topics: Animals; Apoptosis; Brain Injuries; Cytoprotection; Erythropoietin; Free Radicals; Humans; Nerve Degeneration; Neuroprotective Agents; Oxidative Stress; Signal Transduction

Chemotherapy induced peripheral neuropathy is a common and dose-limiting side effect of anticancer drugs. Studies aimed at understanding the underlying mechanism of neurotoxicity of chemotherapeutic drugs have been hampered by lack of suitable culture systems that can differentiate between neuronal cell body, axon or associated glial cells. Here, we have developed an in vitro compartmentalized microfluidic culture system to examine the site of toxicity of chemotherapeutic drugs. To test the culture platform, we used paclitaxel, a widely used anticancer drug for breast cancer, because it causes sensory polyneuropathy in a large proportion of patients and there is no effective treatment. In previous in vitro studies, paclitaxel induced distal axonal degeneration but it was unclear if this was due to direct toxicity on the axon or a consequence of toxicity on the neuronal cell body. Using microfluidic channels that allow compartmentalized culturing of neurons and axons, we demonstrate that the axons are much more susceptible to toxic effects of paclitaxel. When paclitaxel was applied to the axonal side, there was clear degeneration of axons; but when paclitaxel was applied to the soma side, there was no change in axon length. Furthermore, we show that recombinant human erythropoietin, which had been shown to be neuroprotective against paclitaxel neurotoxicity, provides neuroprotection whether it is applied to the cell body or the axons directly. This observation has implications for development of neuroprotective drugs for chemotherapy induced peripheral neuropathies as dorsal root ganglia do not possess blood-nerve-barrier, eliminating one of the cardinal requirements of drug development for the nervous system. This compartmentalized microfluidic culture system can be used for studies aimed at understanding axon degeneration, neuroprotection and development of the nervous system.

Topics: Animals; Antineoplastic Agents, Phytogenic; Axons; Cell Count; Cells, Cultured; Embryo, Mammalian; Erythropoietin; Fluoresceins; Ganglia, Spinal; Microfluidics; Nerve Degeneration; Paclitaxel; Rats; Recombinant Proteins; Sensory Receptor Cells

Inspired from preconditioning studies, ischemic postconditioning, consisting of the application of intermittent interruptions of blood flow shortly after reperfusion, has been described in cardiac ischemia and recently in stroke. It is well known that ischemic tolerance can be achieved in the brain not only by ischemic preconditioning, but also by hypoxic preconditioning. However, the existence of hypoxic postconditioning has never been reported in cerebral ischemia.. Adult mice subjected to transient middle cerebral artery occlusion underwent chronic intermittent hypoxia starting either 1 or 5 days after ischemia and brain damage was assessed by T2-weighted MRI at 43 days. In addition, we investigated the potential neuroprotective effect of hypoxia applied after oxygen glucose deprivation in primary neuronal cultures.. The present study shows for the first time that a late application of hypoxia (5 days) after ischemia reduced delayed thalamic atrophy. Furthermore, hypoxia performed 14 hours after oxygen glucose deprivation induced neuroprotection in primary neuronal cultures. We found that hypoxia-inducible factor-1alpha expression as well as those of its target genes erythropoietin and adrenomedullin is increased by hypoxic postconditioning. Further studies with pharmacological inhibitors or recombinant proteins for erythropoietin and adrenomedullin revealed that these molecules participate in this hypoxia postconditioning-induced neuroprotection.. Altogether, this study demonstrates for the first time the existence of a delayed hypoxic postconditioning in cerebral ischemia and in vitro studies highlight hypoxia-inducible factor-1alpha and its target genes, erythropoietin and adrenomedullin, as potential effectors of postconditioning.

Topics: Adrenomedullin; Animals; Atrophy; Brain; Cells, Cultured; Cytoprotection; Disease Models, Animal; Energy Metabolism; Erythropoietin; Hypoxia-Inducible Factor 1, alpha Subunit; Hypoxia-Ischemia, Brain; Hypoxia, Brain; Infarction, Middle Cerebral Artery; Male; Mice; Nerve Degeneration; Oxidative Stress; Time Factors

Oxygen toxicity has been identified as a risk factor for adverse neurological outcome in survivors of preterm birth. In infant rodent brains, hyperoxia induces disseminated apoptotic neurodegeneration. Because a tissue-protective effect has been observed for recombinant erythropoietin (rEpo), widely used in neonatal medicine for its hematopoietic effect, we examined the effect of rEpo on hyperoxia-induced brain damage.. Six-day-old C57Bl/6 mice or Wistar rats were exposed to hyperoxia (80% O(2)) or normoxia for 24 hours and received rEpo or normal saline injections intraperitoneally. The amount of degenerating cells in their brains was determined by DeOlmos cupric silver staining. Changes of their brain proteome were determined through two-dimensional electrophoresis and mass spectrometry. Western blot, enzyme activity assays and real-time polymerase chain reaction were performed for further analysis of candidate proteins.. Systemic treatment with 20,000 IE/kg rEpo significantly reduced hyperoxia-induced apoptosis and caspase-2, -3, and -8 activity in the brains of infant rodents. In parallel, rEpo inhibited most brain proteome changes observed in infant mice when hyperoxia was applied exclusively. Furthermore, brain proteome changes after a single systemic rEpo treatment point toward a number of mechanisms through which rEpo may generate its protective effect against oxygen toxicity. These include reduction of oxidative stress and restoration of hyperoxia-induced increased levels of proapoptotic factors, as well as decreased levels of neurotrophins.. These findings are highly relevant from a clinical perspective because oxygen administration to neonates is often inevitable, and rEpo may serve as an adjunctive neuroprotective therapy.

Topics: Animals; Apoptosis; Asphyxia Neonatorum; Brain; Caspases; Cell Death; Disease Models, Animal; Erythropoietin; Humans; Hyperoxia; Infant, Newborn; Mice; Mice, Inbred C57BL; Nerve Degeneration; Neurons; Neuroprotective Agents; Oxidative Stress; Oxygen; Oxygen Inhalation Therapy; Proteome; Rats; Rats, Wistar

Both heat acclimation (HA) and post-injury treatment with recombinant human erythropoietin (Epo, rhEpo, exogenous Epo) are neuroprotective against traumatic brain injury (TBI). Our previous data demonstrated that HA-induced neuroprotection includes improved functional recovery and reduced cerebral edema formation. Additionally, in earlier Western-blot analyses, we found that HA mice display increased expression of the specific erythropoietin receptor (EpoR) and of hypoxia-inducible factor-1 alpha (HIF-1 alpha), the inducible subunit of the transcription factor, which regulates Epo gene expression, but not of Epo itself. In light of this, the aim of the current study was threefold: (1) to assess Epo expression in the trauma area and hippocampus following HA, rhEpo administration, or combined HA-rhEpo treatment, using immunohistochemical methods that offer enhanced anatomical resolution; (2) to examine the effects of endogenous and exogenous Epo on edema formation in normothermic (NT) mice; and (3) to evaluate the effects of exogenous Epo administration on neuroprotective outcome measures in HA animals. HA induced enhanced expression of endogenous Epo in the trauma area and the hippocampus. Treatment with anti-Epo antibody given to NT mice increased edema formation, whereas rhEpo induced no beneficial effect. Cognitive performance testing and immunohistochemical findings reinforced HA and rhEpo as separate protective interventions but showed no advantage to combining the two strategies. We therefore suggest that HA-induced neuroprotection is shaped by pre-existing mediators but cannot be modified by post-injury treatment aimed at increasing the levels of neuroprotective agents.

Topics: Acclimatization; Animals; Brain Edema; Brain Injuries; Cognition; Erythropoietin; Fluoresceins; Fluorescent Dyes; Hippocampus; Hot Temperature; Humans; Immunohistochemistry; Immunotherapy; Male; Mice; Nerve Degeneration; Neuroprotective Agents; Organic Chemicals; Recognition, Psychology; Recombinant Proteins

To investigate whether desferoxamine (DFO) preconditioning can induce tolerance against cerebral ischemia and its effect on the expression of hypoxia inducible factor 1alpha (HIF-1alpha) and erythropoietin (EPO) in vivo and in vitro.. Rat model of cerebral ischemia was established by middle cerebral artery occlusion with or without DFO administration. Infarct size was examined by TTC staining, and the neurological severity score was evaluated according to published method. Cortical neurons were cultured under ischemia stress which was mimicked by oxygen-glucose deprivation (OGD), and the neuron damage was assessed by MTT assay. Immunofluorescent staining was employed to detect the expressions of HIF-1alpha and EPO.. The protective effect induced by DFO (decreasing the infarction volume and ameliorating the neurological function) appeared at 2 d after administration of DFO (post-DFO), lasted until 7 d and disappeared at 14 d (P < 0.05); the most effective action was observed at 3 d post-DFO. DFO induced tolerance of cultured neurons against OGD: neuronal viability was increased 23%, 34%, 40%, 48% and 56% at 8 h, 12 h, 24 h, 36 h, and 48 h, respectively, post-DFO (P < 0.05). Immunofluorescent staining found that HIF-1alpha and EPO were upregulated in the neurons of rat brain at 3 d and 7 d post-DFO; increase of HIF-1alpha and EPO appeared in cultured cortex neurons at 36 h and 48 h post-DFO.. DFO induced tolerance against focal cerebral ischemia in rats, and exerted protective effect on OGD cultured cortical neurons. DFO significant induced the expression of HIF-1alpha and EPO both in vivo and in vitro. DFO preconditioning can protect against cerebral ischemia, which may be associated with the synthesis of HIF-1alpha and EPO.

Topics: Animals; Brain Ischemia; Cells, Cultured; Cerebral Infarction; Deferoxamine; Disease Models, Animal; Erythropoietin; Fluorescent Antibody Technique; Hypoxia-Inducible Factor 1, alpha Subunit; Hypoxia-Ischemia, Brain; Infarction, Middle Cerebral Artery; Iron; Ischemic Preconditioning; Nerve Degeneration; Neurons; Rats; Rats, Sprague-Dawley; Siderophores; Time Factors; Treatment Outcome; Up-Regulation

Metallothioneins (MTs) are small cysteine-rich proteins found widely throughout the mammalian body, including the CNS. MT-1 and -2 protect against reactive oxygen species and free radicals. We investigated the role of MT-1 and -2 using MT-1,-2 knockout (KO) mice. MT-1,-2 KO mice exhibited greater neuronal damage after permanent middle cerebral artery occlusion (MCAO) than wild-type mice. MT-2 mRNA was significantly increased at 6, 12, and 24 h after MCAO in the wild-type mouse brain [as detected by real-time reverse-transcription polymerase chain reaction (RT-PCR)], while MT-1 and MT-3 were decreased at 12 and 24 h. In an immunohistochemical study, MT expression displayed colocalization with glial fibrillary acidic protein (GFAP)-positive cells (astrocytes) in the penumbra area in wild-type mice. Since erythropoietin (EPO) has been reported to induce MT-1 and -2 gene expression in vitro, we examined its effect after permanent MCAO, and explored the possible underlying mechanism by examining MT-1 and -2 induction in vivo. In wild-type mice, EPO significantly reduced both infarct area and volume at 24 h after the ischemic insult. However, in MT-1,-2 KO mice EPO-treatment did not alter infarct volume (vs. vehicle-treatment). In wild-type mice at 6 h after EPO administration, real-time RT-PCR revealed increased MT-1 and -2 mRNA expression in the cerebral cortex (without MCAO). Further, MT-1 and -2 immunoreactivity was increased in the cortex of EPO-treated mice. These findings indicate that MTs are induced, and may be neuroprotective against neuronal damage, after MCAO. Furthermore, EPO is neuroprotective in vivo during permanent MCAO, and this may be at least partly mediated by MTs.

Topics: Animals; Astrocytes; Brain; Brain Infarction; Brain Ischemia; Cytoprotection; Erythropoietin; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Infarction, Middle Cerebral Artery; Male; Metallothionein; Metallothionein 3; Mice; Mice, Inbred C57BL; Mice, Knockout; Nerve Degeneration; Neuroprotective Agents; RNA, Messenger; Up-Regulation

Hypoxia is a common cause of cell death and is implicated in many disease processes including stroke and chronic degenerative disorders. In response to hypoxia, cells express a variety of genes which allow adaptation to altered metabolic demands, decreased oxygen demands, and the removal of irreversibly damaged cells. Hypoxia-inducible factor-1 (HIF-1) is a transcription factor that regulates the adaptive response to hypoxia in cells. In this study, we reported an early, time-related, gradual up-regulation of HIF-1alpha, and a moderate increase in vascular endothelial growth factor (VEGF)- and erythropoietin (Epo)-levels following transient focal ischemia. Moreover, we demonstrated, for the first time a specific localization of the pro-apoptotic regulator BNIP3 in striatal and cortical neurons after transient focal ischemia in rats. Prolonged intranuclear BNIP3 immunoreactivity was associated with delayed neuronal death. Experiments showed protein increases on Western blots of brain tissue with peaks at 48h after ischemia. Epo responds to ischemia in an early stage, whereas VEGF and BNIP3 accumulate in cells at later times after ischemia. This suggests the possibility that BH3-only proteins might be one of the major downstream effectors of HIF-1alpha in hypoxic cell death. These findings open the possibility that the hypoxia-regulated pro-apoptotic protein BNIP3 enters the nucleus and could interact with other proteins involved in DNA structure, transcription or mRNA splicing after focal brain ischemia.

Topics: Active Transport, Cell Nucleus; Animals; Apoptosis; Brain Ischemia; Cell Nucleus; Cerebral Cortex; Cerebral Infarction; Corpus Striatum; Disease Models, Animal; Erythropoietin; Gene Expression Regulation; Hypoxia-Inducible Factor 1, alpha Subunit; Infarction, Middle Cerebral Artery; Male; Membrane Proteins; Mitochondrial Proteins; Nerve Degeneration; Neurons; Proto-Oncogene Proteins; Rats; Rats, Wistar; Signal Transduction; Time Factors; Up-Regulation; Vascular Endothelial Growth Factor A

Erythropoietin (EPO), a pleiotropic cytokine involved in erythropoiesis, is tissue-protective in ischemic, traumatic, toxic and inflammatory injuries. In this study, we investigated the effect of EPO in experimental intracerebral hemorrhage (ICH). Two hours after inducing ICH via the stereotaxic infusion of collagenase, recombinant human EPO (500 or 5000 IU/kg, ICH + EPO group) or PBS (ICH + vehicle group) was administered intraperitoneally, then once daily afterwards for 1 or 3 days. ICH + EPO showed the better functional recovery in both rotarod and modified limb placing tests. The brain water content was decreased in ICH + EPO dose-dependently, as compared with ICH + vehicle. The effect of EPO on the brain water content was inhibited by N(omega)-Nitro-L-arginine methyl ester hydrochloride (L-NAME, 10 mg/kg). Mean hemorrhage volume was also decreased in ICH + EPO. EPO reduced the numbers of TUNEL +, myeloperoxidase + or OX-42 + cells in the perihematomal area. In addition, EPO reduced the mRNA level of TNF-alpha, Fas and Fas-L, as well as the activities of caspase-8, 9 and 3. EPO treatment showed up-regulations of endothelial nitric oxide synthase (eNOS) and p-eNOS, pAkt, pSTAT3 and pERK levels. These data suggests that EPO treatment in ICH induces better functional recovery with reducing perihematomal inflammation and apoptosis, coupled with activations of eNOS, STAT3 and ERK.

Topics: Animals; Apoptosis; Biomarkers; Body Water; Brain; Brain Edema; Cell Death; Cerebral Hemorrhage; Disease Models, Animal; Dose-Response Relationship, Drug; Encephalitis; Enzyme Activation; Enzyme Inhibitors; Erythropoietin; Male; Nerve Degeneration; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase Type III; Rats; Rats, Sprague-Dawley; Recovery of Function; Signal Transduction; STAT3 Transcription Factor; Treatment Outcome

To investigate the consequences of inborn excessive erythrocytosis, we made use of our transgenic mouse line (tg6) that constitutively overexpresses erythropoietin (Epo) in a hypoxia-independent manner, thereby reaching hematocrit levels of up to 0.89. We detected expression of human Epo in the brain and, to a lesser extent, in the lung but not in the heart, kidney, or liver of tg6 mice. Although no acute cardiovascular complications are observed, tg6 animals have a reduced lifespan. Decreased swim performance was observed in 5-mo-old tg6 mice. At about 7 mo, several tg6 animals developed spastic contractions of the hindlimbs followed by paralysis. Morphological analysis by light and electron microscopy showed degenerative processes in liver and kidney characterized by increased vascular permeability, chronic progressive inflammation, hemosiderin deposition, and general vasodilatation. Moreover, most of the animals showed severe nerve fiber degeneration of the sciatic nerve, decreased number of neuromuscular junctions, and degeneration of skeletal muscle fibers. Most probably, the developing demyelinating neuropathy resulted in muscular degeneration demonstrated in the extensor digitorum longus muscle. Taken together, chronically increased Epo levels inducing excessive erythrocytosis leads to multiple organ degeneration and reduced life expectancy. This model allows investigation of the impact of excessive erythrocytosis in individuals suffering from polycythemia vera, chronic mountain sickness, or in subjects tempted to abuse Epo by means of gene doping.

Topics: Altitude Sickness; Animals; Disease Models, Animal; Doping in Sports; Erythropoietin; Female; Hematocrit; Humans; Kidney; Liver; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Muscle, Skeletal; Nerve Degeneration; Nervous System; Physical Endurance; Polycythemia

Using an established mouse model of human periventricular leukomalacia, we investigated whether EPO could reduce excitotoxic damage. When administered 1 h following intracerebral injection of 10 microg ibotenic acid at day 5 of life, both a single injection of EPO (5000 IU/kg bw) and repetitive administrations of EPO reduced white and gray matter lesion size. The therapeutic window for protection was small as the protective effect of EPO was lost when EPO administration was delayed to 4 h post-insult. EPO-mediated upregulation of EPO-R, but not EPO, mRNA was observed within 4 h of the excitotoxic insult. The EPO effect was gender independent. Minor hematopoetic effects were observed following EPO treatment. We conclude that a single dose of EPO is sufficient to reduce excitotoxic brain injury and may therefore possess therapeutic relevance in the clinical setting.

Topics: Animals; Animals, Newborn; Brain; Cytoprotection; Disease Models, Animal; Drug Administration Schedule; Erythropoietin; Female; Glutamic Acid; Humans; Ibotenic Acid; Infant, Newborn; Injections, Intraventricular; Leukomalacia, Periventricular; Male; Mice; Nerve Degeneration; Neuroprotective Agents; Neurotoxins; Receptors, Erythropoietin; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Time Factors

Paclitaxel causes a sensory polyneuropathy with characteristic features of distal axonal degeneration. Although the exact mechanisms underlying distal axonal degeneration are unknown, paclitaxel-induced axonal degeneration has been shown to be associated with an increase in detyrosinated tubulin. Here we show that recombinant human erythropoietin prevents axonal degeneration in sensory neurons in vitro and this effect is associated with downregulation of detyrosinated tubulin. Furthermore, in an animal model of paclitaxel-induced distal sensory polyneuropathy, recombinant human erythropoietin protects against distal axonal degeneration. These findings suggest that recombinant human erythropoietin may be useful as a therapy to prevent paclitaxel-induced sensory polyneuropathy in patients undergoing chemotherapy.

Topics: Animals; Antineoplastic Agents, Phytogenic; Axons; Cells, Cultured; Dose-Response Relationship, Drug; Drug Carriers; Erythropoietin; Female; Ganglia, Spinal; Glycerol; Humans; Injections, Intraperitoneal; Mice; Nerve Degeneration; Neurons, Afferent; Neuroprotective Agents; Paclitaxel; Recombinant Proteins; Tubulin; Tyrosine

Apart from its hematopoietic function, erythropoietin (Epo) exerts neuroprotective activity upon reduced oxygenation or ischemia of brain, retina, and spinal cord. To examine whether Epo has an impact on the retrograde degeneration of retinal ganglion cells (RGCs) following optic nerve transection in vivo, we made use of our transgenic mouse line tg21 that constitutively expresses human Epo preferentially in neuronal cells without inducing polycythemia. We show that the tg21 retina expresses human Epo and that RGCs in this mouse line carry the Epo receptor. Upon axotomy, the RGCs of Epo transgenic tg21 mice were protected against degeneration, as compared with wild-type control animals. Western blot analysis revealed decreased phosphorylation levels of STAT-5 and reduced expression of Bcl-XL in RGCs of axotomized tg21 animals, suggesting that the corresponding pathways are not crucial for Epo's neuroprotective activity. Increased phosphorylation levels of ERK-1/-2 and Akt, as well as decreased caspase-3 activity, however, were observed in injured tg21 retinae. Injection of selective inhibitors of ERK-1/-2 (PD98059) or Akt (Wortmannin) pathways into the vitreous space revealed that transgenic Epo protected the RGCs by a pathway involving ERK-1/-2 but not Akt. In view that axotomy-induced degeneration of RGC occurs slowly, and considering the earlier data on the safety and efficacy of Epo in human stroke patients, we predict the clinical implementation of recombinant human Epo not only in patients with acute ischemic stroke, but also with more delayed degenerative neurological diseases.

Topics: Animals; Axotomy; bcl-X Protein; Caspase 3; Caspases; DNA-Binding Proteins; Enzyme Activation; Erythropoietin; Humans; Janus Kinase 2; JNK Mitogen-Activated Protein Kinases; Male; Mice; Mice, Transgenic; Milk Proteins; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Nerve Degeneration; Neuroprotective Agents; Optic Nerve; Phosphatidylinositol 3-Kinases; Protein Serine-Threonine Kinases; Protein-Tyrosine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-bcl-2; Receptors, Erythropoietin; Retina; Retinal Ganglion Cells; Signal Transduction; STAT5 Transcription Factor; Trans-Activators

Both mild hypoxia and exogenous erythropoietin may protect the brain against subsequent severe hypoxia, and the conditioning effect of transient hypoxia is partly mediated by hypoxia-induced endogenous erythropoietin. We now observed in several experimental models that combining transient hypoxia and exogenous erythropoietin may cause neuronal damage. High-dose erythropoietin (40 IU/ml) profoundly impeded synaptic transmission of rat hippocampal slice cultures when used in conjunction with moderate hypoxia (10% O2 for two 8-h periods). Addition of erythropoietin increased viability of cultured rat embryonic cortical neurons at 21% O2 but decreased viability under hypoxic conditions (2% O2) in a dose-dependent fashion. Death of human neuronal precursor cells challenged by oxygen and glucose deprivation was increased by erythropoietin when cells were cultured under hypoxic but not under normoxic conditions. In neonatal rats exposed to moderate hypoxia plus erythropoietin, numbers of degenerating cerebral neurons were increased, as compared to controls or rats subjected to either hypoxia or erythropoietin alone. Thus, erythropoietin may aggravate rather than ameliorate neuronal damage when administered during transient hypoxia.

Topics: Age Factors; Animals; Animals, Newborn; Brain Infarction; Cell Line; Cell Survival; Cells, Cultured; Dose-Response Relationship, Drug; Entorhinal Cortex; Erythropoietin; Glucose; Hippocampus; Humans; Hypoxia-Ischemia, Brain; Infant, Newborn; Nerve Degeneration; Neurons; Neurotoxins; Organ Culture Techniques; Rats; Rats, Wistar; Reperfusion Injury; Stem Cells; Synaptic Transmission

Erythropoietin (EPO) is a hematopoietic growth factor with tissue-protective properties, and can protect animals from cerebral ischemic injury. However, the central nervous effects of EPO as a glycoprotein is limited by the potential complication resulted from its erythropoietic activity and the problem of the penetration through blood-brain barrier (BBB). To avoid these limitations, in this study we administered recombinant human EPO (rhEPO) intranasally (i.n.) to evaluate its neuroprotective effect in the rats with focal cerebral ischemia induced by middle cerebral artery occlusion (MCAO). We found that rhEPO i.n. at doses of 4.8, 12 and 24 U (administered 10 min after MCAO and 1h after reperfusion) reduced infarct volume, brain swelling and cell damage in the ischemic hemispheres, and improved behavioral dysfunction 24 h after cerebral ischemia. Intraperitoneal rhEPO (5000 U/kg) also showed the protective effect, but the heat-inactivated rhEPO did not show any effect. Thus, intranasal administration of relatively small doses of rhEPO protects rats from acute injury after focal cerebral ischemia, suggesting that intranasal rhEPO may be a more effective and safer administration route for treatments of ischemic or other brain diseases.

Topics: Administration, Intranasal; Animals; Brain Edema; Brain Ischemia; Cerebral Infarction; Disease Models, Animal; Dose-Response Relationship, Drug; Erythropoietin; Humans; Infarction, Middle Cerebral Artery; Male; Nerve Degeneration; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Recovery of Function; Treatment Outcome

The developing central nervous system is extremely sensitive to ethanol, with well-defined temporal periods of vulnerability. Recent studies have shown that administration of ethanol to infant rats during the synaptogenesis period triggers extensive apoptotic neurodegeneration throughout many regions of the developing brain. Furthermore, acute ethanol administration produces lipid peroxidation in the brain as an indicator of oxidative stress. In recent years, it has been shown that erythropoietin (EPO) has a critical role in the development, maintenance, protection, and repair of the nervous system. In the present study, we investigated the effect of EPO against ethanol-induced neurodegeneration and oxidative stress in the developing C57BL/6 mouse brain. Seven-day-old C57BL/6 mice were divided into three groups: control group, saline-treated group, EPO-treated group. Ethanol was administered to mice at a dosage of 2.5 g/kg for two times with a 2-h interval. Recombinant human EPO (rhEPO) was given 1000 U/kg. Twenty-four hours after the first dose of ethanol, all the animals were killed. Neuronal cell death, apoptosis, thiobarbituric acid substance (TBARS) levels, superoxide dismutase (SOD), and glutathione peroxidase (Gpx) enzymes activities were evaluated. Histopathological evaluation demonstrated that EPO significantly diminished apoptosis in the cerebellum, prefrontal cortex, and hippocampus and also spared hippocampal CA1, CA2, and CA3 neurons. Simultaneous administration of EPO along with ethanol attenuated the lipid peroxidation process and restored the levels of antioxidants. Regarding the wide use of erythropoietin in premature newborns, this agent may be potentially beneficial in treating ethanol-induced brain injury in the perinatal period.

Topics: Analysis of Variance; Animals; Animals, Newborn; Apoptosis; Brain; Cell Count; Central Nervous System Depressants; Drug Interactions; Erythropoietin; Ethanol; Glutathione Peroxidase; In Situ Nick-End Labeling; Lipid Peroxidation; Mice; Mice, Inbred C57BL; Nerve Degeneration; Neuroprotective Agents; Organ Size; Oxidative Stress; Superoxide Dismutase; Thiobarbituric Acid Reactive Substances; Time Factors

No longer considered exclusive for the function of the hematopoietic system, erythropoietin (EPO) is now considered as a viable agent to address central nervous system injury in a variety of cellular systems that involve neuronal, vascular, and inflammatory cells. Yet, it remains unclear whether the protective capacity of EPO may be effective for chronic neurodegenerative disorders such as Alzheimer's disease (AD) that involve beta-amyloid (Abeta) apoptotic injury to hippocampal neurons. We therefore investigated whether EPO could prevent both early and late apoptotic injury during Abeta exposure in primary hippocampal neurons and assessed potential cellular pathways responsible for this protection. Primary hippocampal neuronal injury was evaluated by trypan blue dye exclusion, DNA fragmentation, membrane phosphatidylserine (PS) exposure, and nuclear factor-kappaB (NF-kappaB) expression with subcellular translocation. We show that EPO, in a concentration specific manner, is able to prevent the loss of both apoptotic genomic DNA integrity and cellular membrane asymmetry during Abeta exposure. This blockade of Abeta generated neuronal apoptosis by EPO is both necessary and sufficient, since protection by EPO is completely abolished by co-treatment with an anti-EPO neutralizing antibody. Furthermore, neuroprotection by EPO is closely linked to the expression of NF-kappaB p65 by preventing the degradation of this protein by Abeta and fostering the subcellular translocation of NF-kappaB p65 from the cytoplasm to the nucleus to allow the initiation of an anti-apoptotic program. In addition, EPO intimately relies upon NF-kappaB p65 to promote neuronal survival, since gene silencing of NF-kappaB p65 by RNA interference removes the protective capacity of EPO during Abeta exposure. Our work illustrates that EPO is an effective entity at the neuronal cellular level against Abeta toxicity and requires the close modulation of the NF-kappaB p65 pathway, suggesting that either EPO or NF-kappaB may be used as future potential therapeutic strategies for the management of chronic neurodegenerative disorders, such as AD.

Topics: Active Transport, Cell Nucleus; Alzheimer Disease; Amyloid beta-Peptides; Animals; Antibodies; Apoptosis; Cell Nucleus; Cells, Cultured; Dose-Response Relationship, Drug; Erythropoietin; Hippocampus; Male; Nerve Degeneration; Neurons; Phosphatidylserines; Protein Transport; Rats; RNA Interference; Transcription Factor RelA

Pharmacological blockade of NMDA receptor function induces apoptotic neurodegeneration in the developing rat brain. However, the use of NMDA receptor antagonists as anesthetics and sedatives represents a difficult-to-avoid clinical practice in pediatrics. This warrants the search for adjunctive neuroprotective measures that will prevent or ameliorate neurotoxicity of NMDA receptor antagonists. The NMDA receptor antagonist MK801 triggered apoptosis in the neonatal rat forebrain, most notably in cortex and thalamus. MK801 exposure reduced mRNA levels of erythropoietin (EPO) and the EPO receptor, suggesting that loss of endogenous EPO activity may contribute to MK801-induced apoptosis. Coadministration of recombinant EPO (rEPO) conferred 50% neuroprotection, partially restored MK801-induced reduction of brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) mRNA, and prevented decreased phosphorylation levels of extracellular signal-regulated protein kinase-1/2 (ERK1/2) and Akt. These observations indicate that rEPO partly rescues newborn rats from MK801-mediated brain damage by enhancing neurotrophin-associated signaling pathways.

Topics: Animals; Animals, Newborn; Apoptosis; Brain; Brain-Derived Neurotrophic Factor; Dizocilpine Maleate; Erythropoietin; Excitatory Amino Acid Antagonists; Glial Cell Line-Derived Neurotrophic Factor; Mice; Mice, Transgenic; Mitogen-Activated Protein Kinases; Nerve Degeneration; Nerve Growth Factors; Neuroprotective Agents; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Rats; Rats, Wistar; Receptors, Erythropoietin; Receptors, N-Methyl-D-Aspartate; Recombinant Fusion Proteins; RNA, Messenger; Signal Transduction

Recent experimental evidence indicates that erythropoietin (Epo), in addition to its hormonal role in regulating red cell production, operates as a neuroprotective agent. So far, the neuroprotective effect of human recombinant Epo (rhEpo) has been mainly demonstrated in models of cerebral ischemia/hypoxia and in selected in vivo studies of traumatic neuronal injury. To further investigate the potential role of this multifunctional trophic factor in post-traumatic cell death, we examined the protective effects of rhEpo in a newly developed model of mechanical trauma in organotypic hippocampal slices. Organotypic rat hippocampal slices were subjected to traumatic injury by allowing a stylus to impact on the CA1 area with an energy of 6 microJ. Hippocampal damage was identified and measured 24 and 48 h later with the fluorescent dye propidium iodide (PI). In untreated slices, the impact induced a significant increase in the mean hippocampal PI fluorescence, co-localized with the area of impact at 24 h (primary post-traumatic injury) and progressively spread to the whole slice between 24 and 48 h (secondary post-traumatic injury). Addition of rhEpo (1-100 UI/mL) or of the NMDA antagonist MK-801 (30 microM) immediately after the traumatic injury reduced hippocampal damage by approximately 30% when observed 24 h later. At 48 h after trauma, the protective effect of rhEpo was greater (by about 47%) and significantly more pronounced than that of MK-801 (28%). Our results suggest that the neuroprotective activity of rhEpo is particularly effective against delayed, secondary post-traumatic damage. This well tolerated agent could provide a therapeutic benefit in pathologies involving post-traumatic neurodegeneration.

Topics: Animals; Animals, Newborn; Brain Injuries; Dizocilpine Maleate; Down-Regulation; Erythropoietin; Excitatory Amino Acid Antagonists; Hippocampus; In Vitro Techniques; Models, Biological; Nerve Degeneration; Neurons; Neuroprotective Agents; Propidium; Rats; Rats, Wistar; Recombinant Fusion Proteins; Recombinant Proteins; Treatment Outcome

Clinically relevant peripheral neuropathies (such as diabetic and human immunodeficiency virus sensory neuropathies) are characterized by distal axonal degeneration, rather than neuronal death. Here, we describe a novel, endogenous pathway that prevents axonal degeneration. We show that in response to axonal injury, periaxonal Schwann cells release erythropoietin (EPO), which via EPO receptor binding on neurons, prevents axonal degeneration. We demonstrate that the relevant axonal injury signal that stimulates EPO production from surrounding glial cells is nitric oxide. In addition, we show that this endogenous pathway can be therapeutically exploited by administering exogenous EPO. In an animal model of distal axonopathy, systemic EPO administration prevents axonal degeneration, and this is associated with a reduction in limb weakness and neuropathic pain behavior. Our in vivo and in vitro data suggest that EPO prevents axonal degeneration and therefore may be therapeutically useful in a wide variety of human neurological diseases characterized by axonopathy.

Topics: Animals; Axons; Cells, Cultured; Erythropoietin; Nerve Degeneration; Rats; Rats, Sprague-Dawley; Receptors, Erythropoietin; Schwann Cells; Signal Transduction

Using immunohistochemistry, expression of erythropoietin (EPO), a hypoxia-inducible neuroprotective factor, and its receptor (EPOR) were investigated in human brain tissue after ischemia/hypoxia. Autopsy brains of neuropathologically normal subjects were compared to those with ischemic infarcts or hypoxic damage. In normal brain, weak EPO/EPOR immunoreactivity was mainly neuronal. In fresh infarcts, EPO immunoreactivity appeared in vascular endothelium, EPOR in microvessels and neuronal fibers. In older infarcts reactive astrocytes exhibited EPO/EPOR immunoreactivity. Acute hypoxic brain damage was associated with vascular EPO expression, older hypoxic damage with EPO/EPOR immunoreactivity in reactive astrocytes. The pronounced up-regulation of EPO/EPOR in human ischemic/hypoxic brains underlines their role as an endogenous neuroprotective system and suggests a novel therapeutic potential in cerebrovascular disease for EPO, a clinically well-characterized and safe compound.

Topics: Adult; Aged; Astrocytes; Brain; Encephalitis; Endothelium, Vascular; Erythropoietin; Female; Fluorescent Antibody Technique; Glial Fibrillary Acidic Protein; Humans; Hypoxia-Ischemia, Brain; Male; Middle Aged; Nerve Degeneration; Neurofilament Proteins; Neurons; Receptors, Erythropoietin; von Willebrand Factor

Erythropoietin (EPO) prevents the ischemia-induced delayed neuronal death in the hippocampal CA1 field in gerbils. EPO receptor (EPOR) is also expressed in the cerebral cortex but its function is not known. To examine whether EPO has a neuroprotective action in the cortex, EPO was infused into the cerebroventricles of stroke-prone spontaneously hypertensive rats with permanent occlusion of the left middle cerebral artery. Morris water maze test indicated that EPO infusion alleviated the ischemia-induced place navigation disability. The left (ischemic)-to-right (contralateral nonischemic) (L/R) ratio of cerebrocortical area in the EPO-infused ischemic group was larger than that in the vehicle-infused ischemic group. The occlusion caused secondary thalamic degeneration but infusion of EPO prevented the decrease in the L/R ratio of thalamic area and supported neuron survival in the ventroposterior thalamic nucleus. In situ hybridization indicated that EPOR mRNA was upregulated in the periphery (ischemic penumbra) of a cerebrocortical infarct after occlusion of the middle cerebral artery, suggesting that an increased number of EPOR in neurons facilitates the EPO signal transmission, thereby preventing the damaged area from enlarging.

Topics: Animals; Arterial Occlusive Diseases; Cell Count; Cerebral Arteries; Cerebral Infarction; Cerebrovascular Disorders; Erythropoietin; Infusion Pumps, Implantable; Maze Learning; Nerve Degeneration; Neurons; Rats; Receptors, Erythropoietin; RNA, Messenger; Spatial Behavior; Thalamus